WorldWideScience

Sample records for selective image cytometry

  1. Multi-channel imaging cytometry with a single detector

    Science.gov (United States)

    Locknar, Sarah; Barton, John; Entwistle, Mark; Carver, Gary; Johnson, Robert

    2018-02-01

    Multi-channel microscopy and multi-channel flow cytometry generate high bit data streams. Multiple channels (both spectral and spatial) are important in diagnosing diseased tissue and identifying individual cells. Omega Optical has developed techniques for mapping multiple channels into the time domain for detection by a single high gain, high bandwidth detector. This approach is based on pulsed laser excitation and a serial array of optical fibers coated with spectral reflectors such that up to 15 wavelength bins are sequentially detected by a single-element detector within 2.5 μs. Our multichannel microscopy system uses firmware running on dedicated DSP and FPGA chips to synchronize the laser, scanning mirrors, and sampling clock. The signals are digitized by an NI board into 14 bits at 60MHz - allowing for 232 by 174 pixel fields in up to 15 channels with 10x over sampling. Our multi-channel imaging cytometry design adds channels for forward scattering and back scattering to the fluorescence spectral channels. All channels are detected within the 2.5 μs - which is compatible with fast cytometry. Going forward, we plan to digitize at 16 bits with an A-toD chip attached to a custom board. Processing these digital signals in custom firmware would allow an on-board graphics processing unit to display imaging flow cytometry data over configurable scanning line lengths. The scatter channels can be used to trigger data buffering when a cell is present in the beam. This approach enables a low cost mechanically robust imaging cytometer.

  2. Microfluidic Imaging Flow Cytometry by Asymmetric-detection Time-stretch Optical Microscopy (ATOM).

    Science.gov (United States)

    Tang, Anson H L; Lai, Queenie T K; Chung, Bob M F; Lee, Kelvin C M; Mok, Aaron T Y; Yip, G K; Shum, Anderson H C; Wong, Kenneth K Y; Tsia, Kevin K

    2017-06-28

    Scaling the number of measurable parameters, which allows for multidimensional data analysis and thus higher-confidence statistical results, has been the main trend in the advanced development of flow cytometry. Notably, adding high-resolution imaging capabilities allows for the complex morphological analysis of cellular/sub-cellular structures. This is not possible with standard flow cytometers. However, it is valuable for advancing our knowledge of cellular functions and can benefit life science research, clinical diagnostics, and environmental monitoring. Incorporating imaging capabilities into flow cytometry compromises the assay throughput, primarily due to the limitations on speed and sensitivity in the camera technologies. To overcome this speed or throughput challenge facing imaging flow cytometry while preserving the image quality, asymmetric-detection time-stretch optical microscopy (ATOM) has been demonstrated to enable high-contrast, single-cell imaging with sub-cellular resolution, at an imaging throughput as high as 100,000 cells/s. Based on the imaging concept of conventional time-stretch imaging, which relies on all-optical image encoding and retrieval through the use of ultrafast broadband laser pulses, ATOM further advances imaging performance by enhancing the image contrast of unlabeled/unstained cells. This is achieved by accessing the phase-gradient information of the cells, which is spectrally encoded into single-shot broadband pulses. Hence, ATOM is particularly advantageous in high-throughput measurements of single-cell morphology and texture - information indicative of cell types, states, and even functions. Ultimately, this could become a powerful imaging flow cytometry platform for the biophysical phenotyping of cells, complementing the current state-of-the-art biochemical-marker-based cellular assay. This work describes a protocol to establish the key modules of an ATOM system (from optical frontend to data processing and visualization

  3. Optofluidic fluorescent imaging cytometry on a cell phone.

    Science.gov (United States)

    Zhu, Hongying; Mavandadi, Sam; Coskun, Ahmet F; Yaglidere, Oguzhan; Ozcan, Aydogan

    2011-09-01

    Fluorescent microscopy and flow cytometry are widely used tools in biomedical sciences. Cost-effective translation of these technologies to remote and resource-limited environments could create new opportunities especially for telemedicine applications. Toward this direction, here we demonstrate the integration of imaging cytometry and fluorescent microscopy on a cell phone using a compact, lightweight, and cost-effective optofluidic attachment. In this cell-phone-based optofluidic imaging cytometry platform, fluorescently labeled particles or cells of interest are continuously delivered to our imaging volume through a disposable microfluidic channel that is positioned above the existing camera unit of the cell phone. The same microfluidic device also acts as a multilayered optofluidic waveguide and efficiently guides our excitation light, which is butt-coupled from the side facets of our microfluidic channel using inexpensive light-emitting diodes. Since the excitation of the sample volume occurs through guided waves that propagate perpendicular to the detection path, our cell-phone camera can record fluorescent movies of the specimens as they are flowing through the microchannel. The digital frames of these fluorescent movies are then rapidly processed to quantify the count and the density of the labeled particles/cells within the target solution of interest. We tested the performance of our cell-phone-based imaging cytometer by measuring the density of white blood cells in human blood samples, which provided a decent match to a commercially available hematology analyzer. We further characterized the imaging quality of the same platform to demonstrate a spatial resolution of ~2 μm. This cell-phone-enabled optofluidic imaging flow cytometer could especially be useful for rapid and sensitive imaging of bodily fluids for conducting various cell counts (e.g., toward monitoring of HIV+ patients) or rare cell analysis as well as for screening of water quality in

  4. BlobFinder, a tool for fluorescence microscopy image cytometry

    OpenAIRE

    Allalou, Amin; Wählby, Carolina

    2009-01-01

    Images can be acquired at high rates with modern fluorescence microscopy hardware, giving rise to a demand for high-speed analysis of image data. Digital image cytometry, i.e., automated measurements and extraction of quantitative data from images of cells, provides valuable information for many types of biomedical analysis. There exists a number of different image analysis software packages that can be programmed to perform a wide array of useful measurements. However, the multi-application ...

  5. Imaging cytometry in a plastic ultra-mobile system

    Science.gov (United States)

    Martínez Vázquez, R.; Trotta, G.; Paturzo, M.; Volpe, A.; Bernava, G.; Basile, V.; Ancona, A.; Ferraro, P.; Fassi, I.; Osellame, R.

    2017-03-01

    We present a cost-effective and highly-portable plastic prototype that can be interfaced with a cell phone to implement an optofluidic imaging cytometry platform. It is based on a PMMA microfluidic chip that fits inside an opto-mechanical platform fabricated by a 3D printer. The fluorescence excitation and imaging is performed using the LED and the CMOS from the cell phone increasing the compactness of the system. A custom developed application is used to analyze the images and provide a value of particle concentration.

  6. Image cytometry: nuclear and chromosomal DNA quantification.

    Science.gov (United States)

    Carvalho, Carlos Roberto; Clarindo, Wellington Ronildo; Abreu, Isabella Santiago

    2011-01-01

    Image cytometry (ICM) associates microscopy, digital image and software technologies, and has been particularly useful in spatial and densitometric cytological analyses, such as DNA ploidy and DNA content measurements. Basically, ICM integrates methodologies of optical microscopy calibration, standard density filters, digital CCD camera, and image analysis softwares for quantitative applications. Apart from all system calibration and setup, cytological protocols must provide good slide preparations for efficient and reliable ICM analysis. In this chapter, procedures for ICM applications employed in our laboratory are described. Protocols shown here for human DNA ploidy determination and quantification of nuclear and chromosomal DNA content in plants could be used as described, or adapted for other studies.

  7. Gross genomic damage measured by DNA image cytometry independently predicts gastric cancer patient survival

    NARCIS (Netherlands)

    Belien, J.A.M.; Buffart, T.E.; Gill, A.; Broeckaert, M.A.M.; Quirke, P.; Meijer, G.A.; Grabsch, H.

    2009-01-01

    BACKGROUND: DNA aneuploidy reflects gross genomic changes. It can be measured by flow cytometry (FCM-DNA) or image cytometry (ICM-DNA). In gastric cancer, the prevalence of DNA aneuploidy has been reported to range from 27 to 100%, with conflicting associations with clinicopathological variables.

  8. High resolution light-sheet based high-throughput imaging cytometry system enables visualization of intra-cellular organelles

    Science.gov (United States)

    Regmi, Raju; Mohan, Kavya; Mondal, Partha Pratim

    2014-09-01

    Visualization of intracellular organelles is achieved using a newly developed high throughput imaging cytometry system. This system interrogates the microfluidic channel using a sheet of light rather than the existing point-based scanning techniques. The advantages of the developed system are many, including, single-shot scanning of specimens flowing through the microfluidic channel at flow rate ranging from micro- to nano- lit./min. Moreover, this opens-up in-vivo imaging of sub-cellular structures and simultaneous cell counting in an imaging cytometry system. We recorded a maximum count of 2400 cells/min at a flow-rate of 700 nl/min, and simultaneous visualization of fluorescently-labeled mitochondrial network in HeLa cells during flow. The developed imaging cytometry system may find immediate application in biotechnology, fluorescence microscopy and nano-medicine.

  9. Wide-field fluorescent microscopy and fluorescent imaging flow cytometry on a cell-phone.

    Science.gov (United States)

    Zhu, Hongying; Ozcan, Aydogan

    2013-04-11

    Fluorescent microscopy and flow cytometry are widely used tools in biomedical research and clinical diagnosis. However these devices are in general relatively bulky and costly, making them less effective in the resource limited settings. To potentially address these limitations, we have recently demonstrated the integration of wide-field fluorescent microscopy and imaging flow cytometry tools on cell-phones using compact, light-weight, and cost-effective opto-fluidic attachments. In our flow cytometry design, fluorescently labeled cells are flushed through a microfluidic channel that is positioned above the existing cell-phone camera unit. Battery powered light-emitting diodes (LEDs) are butt-coupled to the side of this microfluidic chip, which effectively acts as a multi-mode slab waveguide, where the excitation light is guided to uniformly excite the fluorescent targets. The cell-phone camera records a time lapse movie of the fluorescent cells flowing through the microfluidic channel, where the digital frames of this movie are processed to count the number of the labeled cells within the target solution of interest. Using a similar opto-fluidic design, we can also image these fluorescently labeled cells in static mode by e.g. sandwiching the fluorescent particles between two glass slides and capturing their fluorescent images using the cell-phone camera, which can achieve a spatial resolution of e.g. - 10 μm over a very large field-of-view of - 81 mm(2). This cell-phone based fluorescent imaging flow cytometry and microscopy platform might be useful especially in resource limited settings, for e.g. counting of CD4+ T cells toward monitoring of HIV+ patients or for detection of water-borne parasites in drinking water.

  10. European symposium on cytometry

    International Nuclear Information System (INIS)

    1987-01-01

    This book of abstracts contains 59 contributions about cervical prescreening, expert systems, breast cancer, ploidy analysis, system and data evaluation, sampling, preparation and staining, image cytometry, general cytometry, cell kinetics with clinical applications. (AJ)

  11. Statistical performance of image cytometry for DNA, lipids, cytokeratin, & CD45 in a model system for circulation tumor cell detection.

    Science.gov (United States)

    Futia, Gregory L; Schlaepfer, Isabel R; Qamar, Lubna; Behbakht, Kian; Gibson, Emily A

    2017-07-01

    Detection of circulating tumor cells (CTCs) in a blood sample is limited by the sensitivity and specificity of the biomarker panel used to identify CTCs over other blood cells. In this work, we present Bayesian theory that shows how test sensitivity and specificity set the rarity of cell that a test can detect. We perform our calculation of sensitivity and specificity on our image cytometry biomarker panel by testing on pure disease positive (D + ) populations (MCF7 cells) and pure disease negative populations (D - ) (leukocytes). In this system, we performed multi-channel confocal fluorescence microscopy to image biomarkers of DNA, lipids, CD45, and Cytokeratin. Using custom software, we segmented our confocal images into regions of interest consisting of individual cells and computed the image metrics of total signal, second spatial moment, spatial frequency second moment, and the product of the spatial-spatial frequency moments. We present our analysis of these 16 features. The best performing of the 16 features produced an average separation of three standard deviations between D + and D - and an average detectable rarity of ∼1 in 200. We performed multivariable regression and feature selection to combine multiple features for increased performance and showed an average separation of seven standard deviations between the D + and D - populations making our average detectable rarity of ∼1 in 480. Histograms and receiver operating characteristics (ROC) curves for these features and regressions are presented. We conclude that simple regression analysis holds promise to further improve the separation of rare cells in cytometry applications. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

  12. Flow cytometry in diagnostic cytology.

    Science.gov (United States)

    O'Leary, T J

    1998-01-01

    Flow cytometry (FCM) is a useful adjunct to cytologic examination, because the quantitative biochemical information it provides complements the morphologic information gained during visual examination. It aids in the interpretation of bladder washings, and is particularly useful for the assessment of lymphoid lesions, whether they originate from fine-needle aspiration, cerebrospinal fluid, or effusions. Optimal use of FCM frequently requires assessment of more than one parameter; simultaneous use of cell differentiation markers and nuclear DNA quantitation is often significantly more useful than either alone. Despite the utility of FCM, however, the potential for future development appears to be limited. Improvements in image cytometry allow reasonable assessment of ploidy and S-fraction to be made from specimens prepared on glass slides. Multiparameter measurements may also be accomplished with imaging techniques, which allow the further advantage of visual identification of cells with equivocal morphologic changes. The development of artificial intelligence methods for use with imaging technology has also significantly exceeded that of FCM. Finally, image cytometry is often more useful for samples with few cells. Other challenges are posed by immunocytochemical methods which compete with flow cytometry as tools for assessment of proliferation. Given the relatively high cost of FCM instrumentation, survival of FCM as an ancillary technique in cytopathology will require further technical refinements to offset the advantages currently associated with image cytometry and immunocytochemistry.

  13. CytometryML and other data formats

    Science.gov (United States)

    Leif, Robert C.

    2006-02-01

    Cytology automation and research will be enhanced by the creation of a common data format. This data format would provide the pathology and research communities with a uniform way for annotating and exchanging images, flow cytometry, and associated data. This specification and/or standard will include descriptions of the acquisition device, staining, the binary representations of the image and list-mode data, the measurements derived from the image and/or the list-mode data, and descriptors for clinical/pathology and research. An international, vendor-supported, non-proprietary specification will allow pathologists, researchers, and companies to develop and use image capture/analysis software, as well as list-mode analysis software, without worrying about incompatibilities between proprietary vendor formats. Presently, efforts to create specifications and/or descriptions of these formats include the Laboratory Digital Imaging Project (LDIP) Data Exchange Specification; extensions to the Digital Imaging and Communications in Medicine (DICOM); Open Microscopy Environment (OME); Flowcyt, an extension to the present Flow Cytometry Standard (FCS); and CytometryML. The feasibility of creating a common data specification for digital microscopy and flow cytometry in a manner consistent with its use for medical devices and interoperability with both hospital information and picture archiving systems has been demonstrated by the creation of the CytometryML schemas. The feasibility of creating a software system for digital microscopy has been demonstrated by the OME. CytometryML consists of schemas that describe instruments and their measurements. These instruments include digital microscopes and flow cytometers. Optical components including the instruments' excitation and emission parts are described. The description of the measurements made by these instruments includes the tagged molecule, data acquisition subsystem, and the format of the list-mode and/or image data. Many

  14. DNA IMAGE CYTOMETRY IN PROGNOSTICATION OF COLORECTAL CANCER: PRACTICAL CONSIDERATIONS OF THE TECHNIQUE AND INTERPRETATION OF THE HISTOGRAMS

    Directory of Open Access Journals (Sweden)

    Abdelbaset Buhmeida

    2011-05-01

    Full Text Available The role of DNA content as a prognostic factor in colorectal cancer (CRC is highly controversial. Some of these controversies are due to purely technical reasons, e.g. variable practices in interpreting the DNA histograms, which is problematic particularly in advanced cases. In this report, we give a detailed account on various options how these histograms could be optimally interpreted, with the idea of establishing the potential value of DNA image cytometry in prognosis and in selection of proper treatment. Material consists of nuclei isolated from 50 ƒĘm paraffin sections from 160 patients with stage II, III or IV CRC diagnosed, treated and followed-up in our clinic. The nuclei were stained with the Feulgen stain. Nuclear DNA was measured using computer-assisted image cytometry. We applied 4 different approaches to analyse the DNA histograms: 1 appearance of the histogram (ABCDE approach, 2 range of DNA values, 3 peak evaluation, and 4 events present at high DNA values. Intra-observer reproducibility of these four histogram interpretation was 89%, 95%, 96%, and 100%, respectively. We depicted selected histograms to illustrate the four analytical approaches in cases with different stages of CRC, with variable disease outcome. In our analysis, the range of DNA values was the best prognosticator, i.e., the tumours with the widest histograms had the most ominous prognosis. These data implicate that DNA cytometry based on isolated nuclei is valuable in predicting the prognosis of CRC. Different interpretation techniques differed in their reproducibility, but the method showing the best prognostic value also had high reproducibility in our analysis.

  15. Cytometry metadata in XML

    Science.gov (United States)

    Leif, Robert C.; Leif, Stephanie H.

    2016-04-01

    Introduction: The International Society for Advancement of Cytometry (ISAC) has created a standard for the Minimum Information about a Flow Cytometry Experiment (MIFlowCyt 1.0). CytometryML will serve as a common metadata standard for flow and image cytometry (digital microscopy). Methods: The MIFlowCyt data-types were created, as is the rest of CytometryML, in the XML Schema Definition Language (XSD1.1). The datatypes are primarily based on the Flow Cytometry and the Digital Imaging and Communication (DICOM) standards. A small section of the code was formatted with standard HTML formatting elements (p, h1, h2, etc.). Results:1) The part of MIFlowCyt that describes the Experimental Overview including the specimen and substantial parts of several other major elements has been implemented as CytometryML XML schemas (www.cytometryml.org). 2) The feasibility of using MIFlowCyt to provide the combination of an overview, table of contents, and/or an index of a scientific paper or a report has been demonstrated. Previously, a sample electronic publication, EPUB, was created that could contain both MIFlowCyt metadata as well as the binary data. Conclusions: The use of CytometryML technology together with XHTML5 and CSS permits the metadata to be directly formatted and together with the binary data to be stored in an EPUB container. This will facilitate: formatting, data- mining, presentation, data verification, and inclusion in structured research, clinical, and regulatory documents, as well as demonstrate a publication's adherence to the MIFlowCyt standard, promote interoperability and should also result in the textual and numeric data being published using web technology without any change in composition.

  16. A widefield fluorescence microscope with a linear image sensor for image cytometry of biospecimens: Considerations for image quality optimization

    Energy Technology Data Exchange (ETDEWEB)

    Hutcheson, Joshua A.; Majid, Aneeka A.; Powless, Amy J.; Muldoon, Timothy J., E-mail: tmuldoon@uark.edu [Department of Biomedical Engineering, University of Arkansas, 120 Engineering Hall, Fayetteville, Arkansas 72701 (United States)

    2015-09-15

    Linear image sensors have been widely used in numerous research and industry applications to provide continuous imaging of moving objects. Here, we present a widefield fluorescence microscope with a linear image sensor used to image translating objects for image cytometry. First, a calibration curve was characterized for a custom microfluidic chamber over a span of volumetric pump rates. Image data were also acquired using 15 μm fluorescent polystyrene spheres on a slide with a motorized translation stage in order to match linear translation speed with line exposure periods to preserve the image aspect ratio. Aspect ratios were then calculated after imaging to ensure quality control of image data. Fluorescent beads were imaged in suspension flowing through the microfluidics chamber being pumped by a mechanical syringe pump at 16 μl min{sup −1} with a line exposure period of 150 μs. The line period was selected to acquire images of fluorescent beads with a 40 dB signal-to-background ratio. A motorized translation stage was then used to transport conventional glass slides of stained cellular biospecimens. Whole blood collected from healthy volunteers was stained with 0.02% (w/v) proflavine hemisulfate was imaged to highlight leukocyte morphology with a 1.56 mm × 1.28 mm field of view (1540 ms total acquisition time). Oral squamous cells were also collected from healthy volunteers and stained with 0.01% (w/v) proflavine hemisulfate to demonstrate quantifiable subcellular features and an average nuclear to cytoplasmic ratio of 0.03 (n = 75), with a resolution of 0.31 μm pixels{sup −1}.

  17. Application of image cytometry to characterize heterologous lipid flippases in yeast

    DEFF Research Database (Denmark)

    Jensen, Maria Stumph; Costa, Sara; Theorin, Lisa

    2016-01-01

    Lipid flippases are integral membrane proteins that play a central role in moving lipids across cellular membranes. Some of these transporters are ATPases that couple lipid translocation to ATP hydrolysis, whereas others function without any discernible metabolic energy input. A growing number...... is typically monitored by flow cytometry, a costly and maintenance-intensive method. Here, we have optimized a protocol to use an automated image-based cell counter to accurately measure lipid uptake by heterologous lipid flippases expressed in yeast. The method was validated by comparison with the classical...... for characterization of lipid flippase activity, and should be readily adaptable to analyze a variety of other transport systems in yeast, parasites, and mammalian cells. © 2016 International Society for Advancement of Cytometry....

  18. Microfluidic devices and methods for integrated flow cytometry

    Science.gov (United States)

    Srivastava, Nimisha [Goleta, CA; Singh, Anup K [Danville, CA

    2011-08-16

    Microfluidic devices and methods for flow cytometry are described. In described examples, various sample handling and preparation steps may be carried out within a same microfluidic device as flow cytometry steps. A combination of imaging and flow cytometry is described. In some examples, spiral microchannels serve as incubation chambers. Examples of automated sample handling and flow cytometry are described.

  19. Turn-on Fluorescent Probe for Exogenous and Endogenous Imaging of Hypochlorous Acid in Living Cells and Quantitative Application in Flow Cytometry.

    Science.gov (United States)

    Zhan, Zixuan; Liu, Rui; Chai, Li; Li, Qiuyan; Zhang, Kexin; Lv, Yi

    2017-09-05

    Hypochlorous acid (HClO) acts as a dominant microbicidal mediator in the natural immune system, and the excess production of hypochlorites is related to a series of diseases. Thus, it is vitally important and necessary to develop a highly sensitive and selective method for HClO detection in living systems, and most of fluorescent probes are mainly focused on cells imaging. Besides, accurate HClO quantitative information about individual cells in a large cell population is extremely important for understanding inflammation and cellular apoptosis as well. In our work, a turn-on fluorescent probe has been synthesized, which can selectively and sensitively detect HClO with fast response time. The probe is almost nonfluorescent possibly due to both the spirolactam form of fluorescein and unbridged C═N bonds which can undergo a nonradiative decay process in the excited state. Upon the addition of ClO - , the probe was oxidized to ring-opened fluorescent form and the fluorescence intensity was greatly enhanced. In live cell experiments, the probe was successfully applied to image exogenous ClO - in HeLa cells and endogenous HClO in RAW 264.7 macrophage cells. In particular, the quantitative information on exogenous and endogenous HClO can also be acquired in flow cytometry. Therefore, the probe not only can image exogenous and endogenous HClO but also provides a new and promising platform to quantitatively detect HClO in flow cytometry.

  20. Three-dimensional DNA image cytometry by optical projection tomographic microscopy for early cancer diagnosis.

    Science.gov (United States)

    Agarwal, Nitin; Biancardi, Alberto M; Patten, Florence W; Reeves, Anthony P; Seibel, Eric J

    2014-04-01

    Aneuploidy is typically assessed by flow cytometry (FCM) and image cytometry (ICM). We used optical projection tomographic microscopy (OPTM) for assessing cellular DNA content using absorption and fluorescence stains. OPTM combines some of the attributes of both FCM and ICM and generates isometric high-resolution three-dimensional (3-D) images of single cells. Although the depth of field of the microscope objective was in the submicron range, it was extended by scanning the objective's focal plane. The extended depth of field image is similar to a projection in a conventional x-ray computed tomography. These projections were later reconstructed using computed tomography methods to form a 3-D image. We also present an automated method for 3-D nuclear segmentation. Nuclei of chicken, trout, and triploid trout erythrocyte were used to calibrate OPTM. Ratios of integrated optical densities extracted from 50 images of each standard were compared to ratios of DNA indices from FCM. A comparison of mean square errors with thionin, hematoxylin, Feulgen, and SYTOX green was done. Feulgen technique was preferred as it showed highest stoichiometry, least variance, and preserved nuclear morphology in 3-D. The addition of this quantitative biomarker could further strengthen existing classifiers and improve early diagnosis of cancer using 3-D microscopy.

  1. Real-time Image Processing for Microscopy-based Label-free Imaging Flow Cytometry in a Microfluidic Chip.

    Science.gov (United States)

    Heo, Young Jin; Lee, Donghyeon; Kang, Junsu; Lee, Keondo; Chung, Wan Kyun

    2017-09-14

    Imaging flow cytometry (IFC) is an emerging technology that acquires single-cell images at high-throughput for analysis of a cell population. Rich information that comes from high sensitivity and spatial resolution of a single-cell microscopic image is beneficial for single-cell analysis in various biological applications. In this paper, we present a fast image-processing pipeline (R-MOD: Real-time Moving Object Detector) based on deep learning for high-throughput microscopy-based label-free IFC in a microfluidic chip. The R-MOD pipeline acquires all single-cell images of cells in flow, and identifies the acquired images as a real-time process with minimum hardware that consists of a microscope and a high-speed camera. Experiments show that R-MOD has the fast and reliable accuracy (500 fps and 93.3% mAP), and is expected to be used as a powerful tool for biomedical and clinical applications.

  2. Accurate measurement of peripheral blood mononuclear cell concentration using image cytometry to eliminate RBC-induced counting error.

    Science.gov (United States)

    Chan, Leo Li-Ying; Laverty, Daniel J; Smith, Tim; Nejad, Parham; Hei, Hillary; Gandhi, Roopali; Kuksin, Dmitry; Qiu, Jean

    2013-02-28

    Peripheral blood mononuclear cells (PBMCs) have been widely researched in the fields of immunology, infectious disease, oncology, transplantation, hematological malignancy, and vaccine development. Specifically, in immunology research, PBMCs have been utilized to monitor concentration, viability, proliferation, and cytokine production from immune cells, which are critical for both clinical trials and biomedical research. The viability and concentration of isolated PBMCs are traditionally measured by manual counting with trypan blue (TB) using a hemacytometer. One of the common issues of PBMC isolation is red blood cell (RBC) contamination. The RBC contamination can be dependent on the donor sample and/or technical skill level of the operator. RBC contamination in a PBMC sample can introduce error to the measured concentration, which can pass down to future experimental assays performed on these cells. To resolve this issue, RBC lysing protocol can be used to eliminate potential error caused by RBC contamination. In the recent years, a rapid fluorescence-based image cytometry system has been utilized for bright-field and fluorescence imaging analysis of cellular characteristics (Nexcelom Bioscience LLC, Lawrence, MA). The Cellometer image cytometry system has demonstrated the capability of automated concentration and viability detection in disposable counting chambers of unpurified mouse splenocytes and PBMCs stained with acridine orange (AO) and propidium iodide (PI) under fluorescence detection. In this work, we demonstrate the ability of Cellometer image cytometry system to accurately measure PBMC concentration, despite RBC contamination, by comparison of five different total PBMC counting methods: (1) manual counting of trypan blue-stained PBMCs in hemacytometer, (2) manual counting of PBMCs in bright-field images, (3) manual counting of acetic acid lysing of RBCs with TB-stained PBMCs, (4) automated counting of acetic acid lysing of RBCs with PI-stained PBMCs

  3. Cutting-edge analysis of extracellular microparticles using ImageStream(X) imaging flow cytometry.

    Science.gov (United States)

    Headland, Sarah E; Jones, Hefin R; D'Sa, Adelina S V; Perretti, Mauro; Norling, Lucy V

    2014-06-10

    Interest in extracellular vesicle biology has exploded in the past decade, since these microstructures seem endowed with multiple roles, from blood coagulation to inter-cellular communication in pathophysiology. In order for microparticle research to evolve as a preclinical and clinical tool, accurate quantification of microparticle levels is a fundamental requirement, but their size and the complexity of sample fluids present major technical challenges. Flow cytometry is commonly used, but suffers from low sensitivity and accuracy. Use of Amnis ImageStream(X) Mk II imaging flow cytometer afforded accurate analysis of calibration beads ranging from 1 μm to 20 nm; and microparticles, which could be observed and quantified in whole blood, platelet-rich and platelet-free plasma and in leukocyte supernatants. Another advantage was the minimal sample preparation and volume required. Use of this high throughput analyzer allowed simultaneous phenotypic definition of the parent cells and offspring microparticles along with real time microparticle generation kinetics. With the current paucity of reliable techniques for the analysis of microparticles, we propose that the ImageStream(X) could be used effectively to advance this scientific field.

  4. Development of on-chip multi-imaging flow cytometry for identification of imaging biomarkers of clustered circulating tumor cells.

    Directory of Open Access Journals (Sweden)

    Hyonchol Kim

    Full Text Available An on-chip multi-imaging flow cytometry system has been developed to obtain morphometric parameters of cell clusters such as cell number, perimeter, total cross-sectional area, number of nuclei and size of clusters as "imaging biomarkers", with simultaneous acquisition and analysis of both bright-field (BF and fluorescent (FL images at 200 frames per second (fps; by using this system, we examined the effectiveness of using imaging biomarkers for the identification of clustered circulating tumor cells (CTCs. Sample blood of rats in which a prostate cancer cell line (MAT-LyLu had been pre-implanted was applied to a microchannel on a disposable microchip after staining the nuclei using fluorescent dye for their visualization, and the acquired images were measured and compared with those of healthy rats. In terms of the results, clustered cells having (1 cell area larger than 200 µm2 and (2 nucleus area larger than 90 µm2 were specifically observed in cancer cell-implanted blood, but were not observed in healthy rats. In addition, (3 clusters having more than 3 nuclei were specific for cancer-implanted blood and (4 a ratio between the actual perimeter and the perimeter calculated from the obtained area, which reflects a shape distorted from ideal roundness, of less than 0.90 was specific for all clusters having more than 3 nuclei and was also specific for cancer-implanted blood. The collected clusters larger than 300 µm2 were examined by quantitative gene copy number assay, and were identified as being CTCs. These results indicate the usefulness of the imaging biomarkers for characterizing clusters, and all of the four examined imaging biomarkers-cluster area, nuclei area, nuclei number, and ratio of perimeter-can identify clustered CTCs in blood with the same level of preciseness using multi-imaging cytometry.

  5. MXS-Chaining: A Highly Efficient Cloning Platform for Imaging and Flow Cytometry Approaches in Mammalian Systems.

    Directory of Open Access Journals (Sweden)

    Hanna L Sladitschek

    Full Text Available The continuous improvement of imaging technologies has driven the development of sophisticated reporters to monitor biological processes. Such constructs should ideally be assembled in a flexible enough way to allow for their optimization. Here we describe a highly reliable cloning method to efficiently assemble constructs for imaging or flow cytometry applications in mammalian cell culture systems. We bioinformatically identified a list of restriction enzymes whose sites are rarely found in human and mouse cDNA libraries. From the best candidates, we chose an enzyme combination (MluI, XhoI and SalI: MXS that enables iterative chaining of individual building blocks. The ligation scar resulting from the compatible XhoI- and SalI-sticky ends can be translated and hence enables easy in-frame cloning of coding sequences. The robustness of the MXS-chaining approach was validated by assembling constructs up to 20 kb long and comprising up to 34 individual building blocks. By assessing the success rate of 400 ligation reactions, we determined cloning efficiency to be 90% on average. Large polycistronic constructs for single-cell imaging or flow cytometry applications were generated to demonstrate the versatility of the MXS-chaining approach. We devised several constructs that fluorescently label subcellular structures, an adapted version of FUCCI (fluorescent, ubiquitination-based cell cycle indicator optimized to visualize cell cycle progression in mouse embryonic stem cells and an array of artificial promoters enabling dosage of doxycyline-inducible transgene expression. We made publicly available through the Addgene repository a comprehensive set of MXS-building blocks comprising custom vectors, a set of fluorescent proteins, constitutive promoters, polyadenylation signals, selection cassettes and tools for inducible gene expression. Finally, detailed guidelines describe how to chain together prebuilt MXS-building blocks and how to generate new

  6. CytometryML: a markup language for analytical cytology

    Science.gov (United States)

    Leif, Robert C.; Leif, Stephanie H.; Leif, Suzanne B.

    2003-06-01

    Cytometry Markup Language, CytometryML, is a proposed new analytical cytology data standard. CytometryML is a set of XML schemas for encoding both flow cytometry and digital microscopy text based data types. CytometryML schemas reference both DICOM (Digital Imaging and Communications in Medicine) codes and FCS keywords. These schemas provide representations for the keywords in FCS 3.0 and will soon include DICOM microscopic image data. Flow Cytometry Standard (FCS) list-mode has been mapped to the DICOM Waveform Information Object. A preliminary version of a list mode binary data type, which does not presently exist in DICOM, has been designed. This binary type is required to enhance the storage and transmission of flow cytometry and digital microscopy data. Index files based on Waveform indices will be used to rapidly locate the cells present in individual subsets. DICOM has the advantage of employing standard file types, TIF and JPEG, for Digital Microscopy. Using an XML schema based representation means that standard commercial software packages such as Excel and MathCad can be used to analyze, display, and store analytical cytometry data. Furthermore, by providing one standard for both DICOM data and analytical cytology data, it eliminates the need to create and maintain special purpose interfaces for analytical cytology data thereby integrating the data into the larger DICOM and other clinical communities. A draft version of CytometryML is available at www.newportinstruments.com.

  7. FlowCam: Quantification and Classification of Phytoplankton by Imaging Flow Cytometry.

    Science.gov (United States)

    Poulton, Nicole J

    2016-01-01

    The ability to enumerate, classify, and determine biomass of phytoplankton from environmental samples is essential for determining ecosystem function and their role in the aquatic community and microbial food web. Traditional micro-phytoplankton quantification methods using microscopic techniques require preservation and are slow, tedious and very laborious. The availability of more automated imaging microscopy platforms has revolutionized the way particles and cells are detected within their natural environment. The ability to examine cells unaltered and without preservation is key to providing more accurate cell concentration estimates and overall phytoplankton biomass. The FlowCam(®) is an imaging cytometry tool that was originally developed for use in aquatic sciences and provides a more rapid and unbiased method for enumerating and classifying phytoplankton within diverse aquatic environments.

  8. The application of image cytometry to viability assessment in dual fluorescence-stained fish spermatozoa

    Czech Academy of Sciences Publication Activity Database

    Flajšhans, Martin; Cosson, J.; Rodina, Marek; Linhart, Otomar

    2004-01-01

    Roč. 28, č. 12 (2004), s. 955-959 ISSN 1065-6995 R&D Projects: GA MŠk ME 638; GA ČR GA524/03/0178; GA AV ČR KSK6005114 Institutional research plan: CEZ:AV0Z5045916 Keywords : image cytometry * dual fluorescent * staining Subject RIV: ED - Physiology Impact factor: 1.015, year: 2004

  9. Validation of image cytometry for sperm concentration measurement

    DEFF Research Database (Denmark)

    Egeberg Palme, Dorte L.; Johannsen, Trine Holm; Petersen, Jørgen Holm

    2017-01-01

    Sperm concentration is an essential parameter in the diagnostic evaluation of men from infertile couples. It is usually determined by manual counting using a hemocytometer, and is therefore both laborious and subjective. We have earlier shown that a newly developed image cytometry (IC) method may...... be used to determine sperm concentration. Here we present a validation of the IC method by analysis of 4010 semen samples. There was high agreement between IC and manual counting at sperm concentrations above 3 mill/ml and in samples with concentrations above 12 mill/ml the two methods can be used...... a lower coefficient of variation than the manual method (5% vs 10%), indicating a better precision of the IC method. In conclusion, measurement of sperm concentration by IC can be used at concentrations above 3 mill/ml and seems more accurate and precise than manual counting, making it an attractive...

  10. Imaging Flow Cytometry Analysis to Identify Differences of Survival Motor Neuron Protein Expression in Patients With Spinal Muscular Atrophy.

    Science.gov (United States)

    Arakawa, Reiko; Arakawa, Masayuki; Kaneko, Kaori; Otsuki, Noriko; Aoki, Ryoko; Saito, Kayoko

    2016-08-01

    Spinal muscular atrophy is a neurodegenerative disorder caused by the deficient expression of survival motor neuron protein in motor neurons. A major goal of disease-modifying therapy is to increase survival motor neuron expression. Changes in survival motor neuron protein expression can be monitored via peripheral blood cells in patients; therefore we tested the sensitivity and utility of imaging flow cytometry for this purpose. After the immortalization of peripheral blood lymphocytes from a human healthy control subject and two patients with spinal muscular atrophy type 1 with two and three copies of SMN2 gene, respectively, we used imaging flow cytometry analysis to identify significant differences in survival motor neuron expression. A bright detail intensity analysis was used to investigate differences in the cellular localization of survival motor neuron protein. Survival motor neuron expression was significantly decreased in cells derived from patients with spinal muscular atrophy relative to those derived from a healthy control subject. Moreover, survival motor neuron expression correlated with the clinical severity of spinal muscular atrophy according to SMN2 copy number. The cellular accumulation of survival motor neuron protein was also significantly decreased in cells derived from patients with spinal muscular atrophy relative to those derived from a healthy control subject. The benefits of imaging flow cytometry for peripheral blood analysis include its capacities for analyzing heterogeneous cell populations; visualizing cell morphology; and evaluating the accumulation, localization, and expression of a target protein. Imaging flow cytometry analysis should be implemented in future studies to optimize its application as a tool for spinal muscular atrophy clinical trials. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. An open-source solution for advanced imaging flow cytometry data analysis using machine learning.

    Science.gov (United States)

    Hennig, Holger; Rees, Paul; Blasi, Thomas; Kamentsky, Lee; Hung, Jane; Dao, David; Carpenter, Anne E; Filby, Andrew

    2017-01-01

    Imaging flow cytometry (IFC) enables the high throughput collection of morphological and spatial information from hundreds of thousands of single cells. This high content, information rich image data can in theory resolve important biological differences among complex, often heterogeneous biological samples. However, data analysis is often performed in a highly manual and subjective manner using very limited image analysis techniques in combination with conventional flow cytometry gating strategies. This approach is not scalable to the hundreds of available image-based features per cell and thus makes use of only a fraction of the spatial and morphometric information. As a result, the quality, reproducibility and rigour of results are limited by the skill, experience and ingenuity of the data analyst. Here, we describe a pipeline using open-source software that leverages the rich information in digital imagery using machine learning algorithms. Compensated and corrected raw image files (.rif) data files from an imaging flow cytometer (the proprietary .cif file format) are imported into the open-source software CellProfiler, where an image processing pipeline identifies cells and subcellular compartments allowing hundreds of morphological features to be measured. This high-dimensional data can then be analysed using cutting-edge machine learning and clustering approaches using "user-friendly" platforms such as CellProfiler Analyst. Researchers can train an automated cell classifier to recognize different cell types, cell cycle phases, drug treatment/control conditions, etc., using supervised machine learning. This workflow should enable the scientific community to leverage the full analytical power of IFC-derived data sets. It will help to reveal otherwise unappreciated populations of cells based on features that may be hidden to the human eye that include subtle measured differences in label free detection channels such as bright-field and dark-field imagery

  12. Hyperchromatic laser scanning cytometry

    Science.gov (United States)

    Tárnok, Attila; Mittag, Anja

    2007-02-01

    In the emerging fields of high-content and high-throughput single cell analysis for Systems Biology and Cytomics multi- and polychromatic analysis of biological specimens has become increasingly important. Combining different technologies and staining methods polychromatic analysis (i.e. using 8 or more fluorescent colors at a time) can be pushed forward to measure anything stainable in a cell, an approach termed hyperchromatic cytometry. For cytometric cell analysis microscope based Slide Based Cytometry (SBC) technologies are ideal as, unlike flow cytometry, they are non-consumptive, i.e. the analyzed sample is fixed on the slide. Based on the feature of relocation identical cells can be subsequently reanalyzed. In this manner data on the single cell level after manipulation steps can be collected. In this overview various components for hyperchromatic cytometry are demonstrated for a SBC instrument, the Laser Scanning Cytometer (Compucyte Corp., Cambridge, MA): 1) polychromatic cytometry, 2) iterative restaining (using the same fluorochrome for restaining and subsequent reanalysis), 3) differential photobleaching (differentiating fluorochromes by their different photostability), 4) photoactivation (activating fluorescent nanoparticles or photocaged dyes), and 5) photodestruction (destruction of FRET dyes). With the intelligent combination of several of these techniques hyperchromatic cytometry allows to quantify and analyze virtually all components of relevance on the identical cell. The combination of high-throughput and high-content SBC analysis with high-resolution confocal imaging allows clear verification of phenotypically distinct subpopulations of cells with structural information. The information gained per specimen is only limited by the number of available antibodies and by sterical hindrance.

  13. Flow cytometry and integrated imaging

    Directory of Open Access Journals (Sweden)

    V. Kachel

    2000-06-01

    Full Text Available It is a serious problem to relate the results of a flow cytometric analysis of a marine sample to different species. Images of particles selectively triggered by the flow cytometric analysis and picked out from the flowing stream give a valuable additional information on the analyzed organisms. The technical principles and problems of triggered imaging in flow are discussed, as well as the positioning of the particles in the plane of focus, freezing the motion of the quickly moving objects and what kinds of light sources are suitable for pulsed illumination. The images have to be stored either by film or electronically. The features of camera targets and the memory requirements for storing the image data and the conditions for the triggering device are shown. A brief explanation of the features of three realized flow cytometric imaging (FCI systems is given: the Macro Flow Planktometer built within the EUROMAR MAROPT project, the Imaging Module of the European Plankton Analysis System, supported by the MAST II EurOPA project and the most recently developed FLUVO VI universal flow cytometer including HBO 100- and laser excitation for fluorescence and scatter, Coulter sizing as well as bright field and and phase contrast FCI.

  14. Fully Automated On-Chip Imaging Flow Cytometry System with Disposable Contamination-Free Plastic Re-Cultivation Chip

    Directory of Open Access Journals (Sweden)

    Tomoyuki Kaneko

    2011-06-01

    Full Text Available We have developed a novel imaging cytometry system using a poly(methyl methacrylate (PMMA based microfluidic chip. The system was contamination-free, because sample suspensions contacted only with a flammable PMMA chip and no other component of the system. The transparency and low-fluorescence of PMMA was suitable for microscopic imaging of cells flowing through microchannels on the chip. Sample particles flowing through microchannels on the chip were discriminated by an image-recognition unit with a high-speed camera in real time at the rate of 200 event/s, e.g., microparticles 2.5 μm and 3.0 μm in diameter were differentiated with an error rate of less than 2%. Desired cells were separated automatically from other cells by electrophoretic or dielectrophoretic force one by one with a separation efficiency of 90%. Cells in suspension with fluorescent dye were separated using the same kind of microfluidic chip. Sample of 5 μL with 1 × 106 particle/mL was processed within 40 min. Separated cells could be cultured on the microfluidic chip without contamination. The whole operation of sample handling was automated using 3D micropipetting system. These results showed that the novel imaging flow cytometry system is practically applicable for biological research and clinical diagnostics.

  15. Identification of a murine erythroblast subpopulation enriched in enucleating events by multi-spectral imaging flow cytometry.

    Science.gov (United States)

    Konstantinidis, Diamantis G; Pushkaran, Suvarnamala; Giger, Katie; Manganaris, Stefanos; Zheng, Yi; Kalfa, Theodosia A

    2014-06-06

    Erythropoiesis in mammals concludes with the dramatic process of enucleation that results in reticulocyte formation. The mechanism of enucleation has not yet been fully elucidated. A common problem encountered when studying the localization of key proteins and structures within enucleating erythroblasts by microscopy is the difficulty to observe a sufficient number of cells undergoing enucleation. We have developed a novel analysis protocol using multiparameter high-speed cell imaging in flow (Multi-Spectral Imaging Flow Cytometry), a method that combines immunofluorescent microscopy with flow cytometry, in order to identify efficiently a significant number of enucleating events, that allows to obtain measurements and perform statistical analysis. We first describe here two in vitro erythropoiesis culture methods used in order to synchronize murine erythroblasts and increase the probability of capturing enucleation at the time of evaluation. Then, we describe in detail the staining of erythroblasts after fixation and permeabilization in order to study the localization of intracellular proteins or lipid rafts during enucleation by multi-spectral imaging flow cytometry. Along with size and DNA/Ter119 staining which are used to identify the orthochromatic erythroblasts, we utilize the parameters "aspect ratio" of a cell in the bright-field channel that aids in the recognition of elongated cells and "delta centroid XY Ter119/Draq5" that allows the identification of cellular events in which the center of Ter119 staining (nascent reticulocyte) is far apart from the center of Draq5 staining (nucleus undergoing extrusion), thus indicating a cell about to enucleate. The subset of the orthochromatic erythroblast population with high delta centroid and low aspect ratio is highly enriched in enucleating cells.

  16. Hyperspectral cytometry.

    Science.gov (United States)

    Grégori, Gérald; Rajwa, Bartek; Patsekin, Valery; Jones, James; Furuki, Motohiro; Yamamoto, Masanobu; Paul Robinson, J

    2014-01-01

    Hyperspectral cytometry is an emerging technology for single-cell analysis that combines ultrafast optical spectroscopy and flow cytometry. Spectral cytometry systems utilize diffraction gratings or prism-based monochromators to disperse fluorescence signals from multiple labels (organic dyes, nanoparticles, or fluorescent proteins) present in each analyzed bioparticle onto linear detector arrays such as multianode photomultipliers or charge-coupled device sensors. The resultant data, consisting of a series of characterizing every analyzed cell, are not compensated by employing the traditional cytometry approach, but rather are spectrally unmixed utilizing algorithms such as constrained Poisson regression or non-negative matrix factorization. Although implementations of spectral cytometry were envisioned as early as the 1980s, only recently has the development of highly sensitive photomultiplier tube arrays led to design and construction of functional prototypes and subsequently to introduction of commercially available systems. This chapter summarizes the historical efforts and work in the field of spectral cytometry performed at Purdue University Cytometry Laboratories and describes the technology developed by Sony Corporation that resulted in release of the first commercial spectral cytometry system-the Sony SP6800. A brief introduction to spectral data analysis is also provided, with emphasis on the differences between traditional polychromatic and spectral cytometry approaches.

  17. Imaging flow cytometry assays for quantifying pigment grade titanium dioxide particle internalization and interactions with immune cells in whole blood.

    Science.gov (United States)

    Hewitt, Rachel E; Vis, Bradley; Pele, Laetitia C; Faria, Nuno; Powell, Jonathan J

    2017-10-01

    Pigment grade titanium dioxide is composed of sub-micron sized particles, including a nanofraction, and is widely utilized in food, cosmetic, pharmaceutical, and biomedical industries. Oral exposure to pigment grade titanium dioxide results in at least some material entering the circulation in humans, although subsequent interactions with blood immune cells are unknown. Pigment grade titanium dioxide is employed for its strong light scattering properties, and this work exploited that attribute to determine whether single cell-particle associations could be determined in immune cells of human whole blood at "real life" concentrations. In vitro assays, initially using isolated peripheral blood mononuclear cells, identified titanium dioxide associated with the surface of, and within, immune cells by darkfield reflectance in imaging flow cytometry. This was confirmed at the population level by side scatter measurements using conventional flow cytometry. Next, it was demonstrated that imaging flow cytometry could quantify titanium dioxide particle-bearing cells, within the immune cell populations of fresh whole blood, down to titanium dioxide levels of 10 parts per billion, which is in the range anticipated for human blood following titanium dioxide ingestion. Moreover, surface association and internal localization of titanium dioxide particles could be discriminated in the assays. Overall, results showed that in addition to the anticipated activity of blood monocytes internalizing titanium dioxide particles, neutrophil internalization and cell membrane adhesion also occurred, the latter for both phagocytic and nonphagocytic cell types. What happens in vivo and whether this contributes to activation of one or more of these different cells types in blood merits further attention. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

  18. Semi-automatized segmentation method using image-based flow cytometry to study sperm physiology: the case of capacitation-induced tyrosine phosphorylation.

    Science.gov (United States)

    Matamoros-Volante, Arturo; Moreno-Irusta, Ayelen; Torres-Rodriguez, Paulina; Giojalas, Laura; Gervasi, María G; Visconti, Pablo E; Treviño, Claudia L

    2018-02-01

    Is image-based flow cytometry a useful tool to study intracellular events in human sperm such as protein tyrosine phosphorylation or signaling processes? Image-based flow cytometry is a powerful tool to study intracellular events in a relevant number of sperm cells, which enables a robust statistical analysis providing spatial resolution in terms of the specific subcellular localization of the labeling. Sperm capacitation is required for fertilization. During this process, spermatozoa undergo numerous physiological changes, via activation of different signaling pathways, which are not completely understood. Classical approaches for studying sperm physiology include conventional microscopy, flow cytometry and Western blotting. These techniques present disadvantages for obtaining detailed subcellular information of signaling pathways in a relevant number of cells. This work describes a new semi-automatized analysis using image-based flow cytometry which enables the study, at the subcellular and population levels, of different sperm parameters associated with signaling. The increase in protein tyrosine phosphorylation during capacitation is presented as an example. Sperm cells were isolated from seminal plasma by the swim-up technique. We evaluated the intensity and distribution of protein tyrosine phosphorylation in sperm incubated in non-capacitation and capacitation-supporting media for 1 and 18 h under different experimental conditions. We used an antibody against FER kinase and pharmacological inhibitors in an attempt to identify the kinases involved in protein tyrosine phosphorylation during human sperm capacitation. Semen samples from normospermic donors were obtained by masturbation after 2-3 days of sexual abstinence. We used the innovative technique image-based flow cytometry and image analysis tools to segment individual images of spermatozoa. We evaluated and quantified the regions of sperm where protein tyrosine phosphorylation takes place at the

  19. Label-free cell-cycle analysis by high-throughput quantitative phase time-stretch imaging flow cytometry

    Science.gov (United States)

    Mok, Aaron T. Y.; Lee, Kelvin C. M.; Wong, Kenneth K. Y.; Tsia, Kevin K.

    2018-02-01

    Biophysical properties of cells could complement and correlate biochemical markers to characterize a multitude of cellular states. Changes in cell size, dry mass and subcellular morphology, for instance, are relevant to cell-cycle progression which is prevalently evaluated by DNA-targeted fluorescence measurements. Quantitative-phase microscopy (QPM) is among the effective biophysical phenotyping tools that can quantify cell sizes and sub-cellular dry mass density distribution of single cells at high spatial resolution. However, limited camera frame rate and thus imaging throughput makes QPM incompatible with high-throughput flow cytometry - a gold standard in multiparametric cell-based assay. Here we present a high-throughput approach for label-free analysis of cell cycle based on quantitative-phase time-stretch imaging flow cytometry at a throughput of > 10,000 cells/s. Our time-stretch QPM system enables sub-cellular resolution even at high speed, allowing us to extract a multitude (at least 24) of single-cell biophysical phenotypes (from both amplitude and phase images). Those phenotypes can be combined to track cell-cycle progression based on a t-distributed stochastic neighbor embedding (t-SNE) algorithm. Using multivariate analysis of variance (MANOVA) discriminant analysis, cell-cycle phases can also be predicted label-free with high accuracy at >90% in G1 and G2 phase, and >80% in S phase. We anticipate that high throughput label-free cell cycle characterization could open new approaches for large-scale single-cell analysis, bringing new mechanistic insights into complex biological processes including diseases pathogenesis.

  20. Selective grazing by adults and larvae of the zebra mussel (Dreissena polymorpha): application of flow cytometry to natural seston

    NARCIS (Netherlands)

    Dionisio Pires, L.M.; Jonker, R.R.; Donk, E.van; Laanbroek, H.J.

    2004-01-01

    1. Selective grazing of adults and larvae of the zebra mussel (Dreissena polymorpha) on phytoplankton and detritus from both laboratory cultures and natural seston was quantified using flow cytometry. 2. Mean clearance rate of adult zebra mussels was higher on a mixture of the green

  1. Selective grazing by adults and larvae of the zebra mussel (Dreissena polymorpha): application of flow cytometry to natural seston

    NARCIS (Netherlands)

    Pires, L.M.D.; Jonker, R.M.; Van Donk, E.; Laanbroek, H.J.

    2004-01-01

    1. Selective grazing of adults and larvae of the zebra mussel (Dreissena polymorpha) on phytoplankton and detritus from both laboratory cultures and natural seston was quantified using flow cytometry. 2. Mean clearance rate of adult zebra mussels was higher on a mixture of the green alga Scenedesmus

  2. An imaging flow cytometry method to assess ricin trafficking in A549 human lung epithelial cells.

    Science.gov (United States)

    Jenner, Dominic; Chong, Damien; Walker, Nicola; Green, A Christopher

    2018-02-01

    The endocytosis and trafficking of ricin in mammalian cells is an important area of research for those producing ricin anti-toxins and other ricin therapeutics. Ricin trafficking is usually observed by fluorescence microscopy techniques. This gives good resolution and leads to a detailed understanding of the internal movement of ricin within cells. However, microscopy techniques are often hampered by complex analysis and quantification techniques, and the inability to look at ricin trafficking in large populations of cells. In these studies we have directly labelled ricin and assessed if its trafficking can be observed using Imaging Flow Cytometry (IFC) both to the cytoplasmic region of cells and specifically to the Golgi apparatus. Using IDEAS® data analysis software the specific fluorescence location of the ricin within the cells was analysed. Then, using cytoplasmic masking techniques to quantify the number of cells with endocytosed cytoplasmic ricin or cells with Golgi-associated ricin, kinetic endocytosis curves were generated. Here we present, to the authors' knowledge, the first example of using imaging flow cytometry for evaluating the subcellular transport of protein cargo, using the trafficking of ricin toxin in lung cells as a model. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  3. CytometryML with DICOM and FCS

    Science.gov (United States)

    Leif, Robert C.

    2018-02-01

    Abstract: Flow Cytometry Standard, FCS, and Digital Imaging and Communications in Medicine standard, DICOM, are based on extensive, superb domain knowledge, However, they are isolated systems, do not take advantage of data structures, require special programs to read and write the data, lack the capability to interoperate or work with other standards and FCS lacks many of the datatypes necessary for clinical laboratory data. The large overlap between imaging and flow cytometry provides strong evidence that both modalities should be covered by the same standard. Method: The XML Schema Definition Language, XSD 1.1 was used to translate FCS and/or DICOM objects. A MIFlowCyt file was tested with published values. Results: Previously, a significant part of an XML standard based upon a combination of FCS and DICOM has been implemented and validated with MIFlowCyt data. Strongly typed translations of FCS keywords have been constructed in XML. These keywords contain links to their DICOM and FCS equivalents.

  4. Simultaneous cathodoluminescence and electron microscopy cytometry of cellular vesicles labeled with fluorescent nanodiamonds

    Science.gov (United States)

    Nagarajan, Sounderya; Pioche-Durieu, Catherine; Tizei, Luiz H. G.; Fang, Chia-Yi; Bertrand, Jean-Rémi; Le Cam, Eric; Chang, Huan-Cheng; Treussart, François; Kociak, Mathieu

    2016-06-01

    Light and Transmission Electron Microscopies (LM and TEM) hold potential in bioimaging owing to the advantages of fast imaging of multiple cells with LM and ultrastructure resolution offered by TEM. Integrated or correlated LM and TEM are the current approaches to combine the advantages of both techniques. Here we propose an alternative in which the electron beam of a scanning TEM (STEM) is used to excite concomitantly the luminescence of nanoparticle labels (a process known as cathodoluminescence, CL), and image the cell ultrastructure. This CL-STEM imaging allows obtaining luminescence spectra and imaging ultrastructure simultaneously. We present a proof of principle experiment, showing the potential of this technique in image cytometry of cell vesicular components. To label the vesicles we used fluorescent diamond nanocrystals (nanodiamonds, NDs) of size ~150 nm coated with different cationic polymers, known to trigger different internalization pathways. Each polymer was associated with a type of ND with a different emission spectrum. With CL-STEM, for each individual vesicle, we were able to measure (i) their size with nanometric resolution, (ii) their content in different ND labels, and realize intracellular component cytometry. In contrast to the recently reported organelle flow cytometry technique that requires cell sonication, CL-STEM-based image cytometry preserves the cell integrity and provides a much higher resolution in size. Although this novel approach is still limited by a low throughput, the automatization of data acquisition and image analysis, combined with improved intracellular targeting, should facilitate applications in cell biology at the subcellular level.Light and Transmission Electron Microscopies (LM and TEM) hold potential in bioimaging owing to the advantages of fast imaging of multiple cells with LM and ultrastructure resolution offered by TEM. Integrated or correlated LM and TEM are the current approaches to combine the advantages of

  5. Dictionary-enhanced imaging cytometry

    Science.gov (United States)

    Orth, Antony; Schaak, Diane; Schonbrun, Ethan

    2017-02-01

    State-of-the-art high-throughput microscopes are now capable of recording image data at a phenomenal rate, imaging entire microscope slides in minutes. In this paper we investigate how a large image set can be used to perform automated cell classification and denoising. To this end, we acquire an image library consisting of over one quarter-million white blood cell (WBC) nuclei together with CD15/CD16 protein expression for each cell. We show that the WBC nucleus images alone can be used to replicate CD expression-based gating, even in the presence of significant imaging noise. We also demonstrate that accurate estimates of white blood cell images can be recovered from extremely noisy images by comparing with a reference dictionary. This has implications for dose-limited imaging when samples belong to a highly restricted class such as a well-studied cell type. Furthermore, large image libraries may endow microscopes with capabilities beyond their hardware specifications in terms of sensitivity and resolution. We call for researchers to crowd source large image libraries of common cell lines to explore this possibility.

  6. Morphological observation and analysis using automated image cytometry for the comparison of trypan blue and fluorescence-based viability detection method.

    Science.gov (United States)

    Chan, Leo Li-Ying; Kuksin, Dmitry; Laverty, Daniel J; Saldi, Stephanie; Qiu, Jean

    2015-05-01

    The ability to accurately determine cell viability is essential to performing a well-controlled biological experiment. Typical experiments range from standard cell culturing to advanced cell-based assays that may require cell viability measurement for downstream experiments. The traditional cell viability measurement method has been the trypan blue (TB) exclusion assay. However, since the introduction of fluorescence-based dyes for cell viability measurement using flow or image-based cytometry systems, there have been numerous publications comparing the two detection methods. Although previous studies have shown discrepancies between TB exclusion and fluorescence-based viability measurements, image-based morphological analysis was not performed in order to examine the viability discrepancies. In this work, we compared TB exclusion and fluorescence-based viability detection methods using image cytometry to observe morphological changes due to the effect of TB on dead cells. Imaging results showed that as the viability of a naturally-dying Jurkat cell sample decreased below 70 %, many TB-stained cells began to exhibit non-uniform morphological characteristics. Dead cells with these characteristics may be difficult to count under light microscopy, thus generating an artificially higher viability measurement compared to fluorescence-based method. These morphological observations can potentially explain the differences in viability measurement between the two methods.

  7. CytometryML: a data standard which has been designed to interface with other standards

    Science.gov (United States)

    Leif, Robert C.

    2007-02-01

    Because of the differences in the requirements, needs, and past histories including existing standards of the creating organizations, a single encompassing cytology-pathology standard will not, in the near future, replace the multiple existing or under development standards. Except for DICOM and FCS, these standardization efforts are all based on XML. CytometryML is a collection of XML schemas, which are based on the Digital Imaging and Communications in Medicine (DICOM) and Flow Cytometry Standard (FCS) datatypes. The CytometryML schemas contain attributes that link them to the DICOM standard and FCS. Interoperability with DICOM has been facilitated by, wherever reasonable, limiting the difference between CytometryML and the previous standards to syntax. In order to permit the Resource Description Framework, RDF, to reference the CytometryML datatypes, id attributes have been added to many CytometryML elements. The Laboratory Digital Imaging Project (LDIP) Data Exchange Specification and the Flowcyt standards development effort employ RDF syntax. Documentation from DICOM has been reused in CytometryML. The unity of analytical cytology was demonstrated by deriving a microscope type and a flow cytometer type from a generic cytometry instrument type. The feasibility of incorporating the Flowcyt gating schemas into CytometryML has been demonstrated. CytometryML is being extended to include many of the new DICOM Working Group 26 datatypes, which describe patients, specimens, and analytes. In situations where multiple standards are being created, interoperability can be facilitated by employing datatypes based on a common set of semantics and building in links to standards that employ different syntax.

  8. Simultaneous cathodoluminescence and electron microscopy cytometry of cellular vesicles labeled with fluorescent nanodiamonds.

    Science.gov (United States)

    Nagarajan, Sounderya; Pioche-Durieu, Catherine; Tizei, Luiz H G; Fang, Chia-Yi; Bertrand, Jean-Rémi; Le Cam, Eric; Chang, Huan-Cheng; Treussart, François; Kociak, Mathieu

    2016-06-02

    Light and Transmission Electron Microscopies (LM and TEM) hold potential in bioimaging owing to the advantages of fast imaging of multiple cells with LM and ultrastructure resolution offered by TEM. Integrated or correlated LM and TEM are the current approaches to combine the advantages of both techniques. Here we propose an alternative in which the electron beam of a scanning TEM (STEM) is used to excite concomitantly the luminescence of nanoparticle labels (a process known as cathodoluminescence, CL), and image the cell ultrastructure. This CL-STEM imaging allows obtaining luminescence spectra and imaging ultrastructure simultaneously. We present a proof of principle experiment, showing the potential of this technique in image cytometry of cell vesicular components. To label the vesicles we used fluorescent diamond nanocrystals (nanodiamonds, NDs) of size ≈150 nm coated with different cationic polymers, known to trigger different internalization pathways. Each polymer was associated with a type of ND with a different emission spectrum. With CL-STEM, for each individual vesicle, we were able to measure (i) their size with nanometric resolution, (ii) their content in different ND labels, and realize intracellular component cytometry. In contrast to the recently reported organelle flow cytometry technique that requires cell sonication, CL-STEM-based image cytometry preserves the cell integrity and provides a much higher resolution in size. Although this novel approach is still limited by a low throughput, the automatization of data acquisition and image analysis, combined with improved intracellular targeting, should facilitate applications in cell biology at the subcellular level.

  9. Role of Brush Biopsy and DNA Cytometry for Prevention, Diagnosis, Therapy, and Followup Care of Oral Cancer

    Directory of Open Access Journals (Sweden)

    Alfred Böcking

    2011-01-01

    Full Text Available Late diagnosis resulting in late treatment and locoregional failure after surgery are the main causes of death in patients with oral squamous cell carcinomas (SCCs. Actually, exfoliative cytology is increasingly used for early detection of oral cancer and has been the subject of intense research over the last five years. Significant advances have been made both in relation to screening and evaluation of precursor lesions. As this noninvasive procedure is well tolerated by patients, more lesions may be screened and thus more oral cancers may be found in early, curable stages. Moreover, the additional use of DNA image cytometry is a reasonable tool for the assessment of the resection margins of SCC. DNA image cytometry could help to find the appropriate treatment option for the patients. Finally, diagnostic DNA image cytometry is an accurate method and has internationally been standardized. In conclusion, DNA image cytometry has increasing impact on the prevention, diagnostic, and therapeutical considerations in head and neck SCC.

  10. Practical flow cytometry

    National Research Council Canada - National Science Library

    Shapiro, Howard M

    2003-01-01

    ... ... Conflict: Resolution ... 1.3 Problem Number One: Finding The Cell(s) ... Flow Cytometry: Quick on the Trigger ... The Main Event ... The Pulse Quickens, the Plot Thickens ... 1.4 Flow Cytometry: ...

  11. CMOS based image cytometry for detection of phytoplankton in ballast water.

    Science.gov (United States)

    Pérez, J M; Jofre, M; Martínez, P; Yáñez, M A; Catalan, V; Parker, A; Veldhuis, M; Pruneri, V

    2017-02-01

    We introduce an image cytometer (I-CYT) for the analysis of phytoplankton in fresh and marine water environments. A linear quantification of cell numbers was observed covering several orders of magnitude using cultures of Tetraselmis and Nannochloropsis measured by autofluorescence in a laboratory environment. We assessed the functionality of the system outside the laboratory by phytoplankton quantification of samples taken from a marine water environment (Dutch Wadden Sea, The Netherlands) and a fresh water environment (Lake Ijssel, The Netherlands). The I-CYT was also employed to study the effects of two ballast water treatment systems (BWTS), based on chlorine electrolysis and UV sterilization, with the analysis including the vitality of the phytoplankton. For comparative study and benchmarking of the I-CYT, a standard flow cytometer was used. Our results prove a limit of detection (LOD) of 10 cells/ml with an accuracy between 0.7 and 0.5 log, and a correlation of 88.29% in quantification and 96.21% in vitality, with respect to the flow cytometry results.

  12. Application of image flow cytometry for the characterization of red blood cell morphology

    Science.gov (United States)

    Pinto, Ruben N.; Sebastian, Joseph A.; Parsons, Michael; Chang, Tim C.; Acker, Jason P.; Kolios, Michael C.

    2017-02-01

    Red blood cells (RBCs) stored in hypothermic environments for the purpose of transfusion have been documented to undergo structural and functional changes over time. One sign of the so-called RBC storage lesion is irreversible damage to the cell membrane. Consequently, RBCs undergo a morphological transformation from regular, deformable biconcave discocytes to rigid spheroechinocytes. The spherically shaped RBCs lack the deformability to efficiently enter microvasculature, thereby reducing the capacity of RBCs to oxygenate tissue. Blood banks currently rely on microscope techniques that include fixing, staining and cell counting in order to morphologically characterize RBC samples; these methods are labor intensive and highly subjective. This study presents a novel, high-throughput RBC morphology characterization technique using image flow cytometry (IFC). An image segmentation template was developed to process 100,000 images acquired from the IFC system and output the relative spheroechinocyte percentage. The technique was applied on samples extracted from two blood bags to monitor the morphological changes of the RBCs during in vitro hypothermic storage. The study found that, for a given sample of RBCs, the IFC method was twice as fast in data acquisition, and analyzed 250-350 times more RBCs than the conventional method. Over the lifespan of the blood bags, the mean spheroechinocyte population increased by 37%. Future work will focus on expanding the template to segregate RBC images into more subpopulations for the validation of the IFC method against conventional techniques; the expanded template will aid in establishing quantitative links between spheroechinocyte increase and other RBC storage lesion characteristics.

  13. Opto-fluidics based microscopy and flow cytometry on a cell phone for blood analysis.

    Science.gov (United States)

    Zhu, Hongying; Ozcan, Aydogan

    2015-01-01

    Blood analysis is one of the most important clinical tests for medical diagnosis. Flow cytometry and optical microscopy are widely used techniques to perform blood analysis and therefore cost-effective translation of these technologies to resource limited settings is critical for various global health as well as telemedicine applications. In this chapter, we review our recent progress on the integration of imaging flow cytometry and fluorescent microscopy on a cell phone using compact, light-weight and cost-effective opto-fluidic attachments integrated onto the camera module of a smartphone. In our cell-phone based opto-fluidic imaging cytometry design, fluorescently labeled cells are delivered into the imaging area using a disposable micro-fluidic chip that is positioned above the existing camera unit of the cell phone. Battery powered light-emitting diodes (LEDs) are butt-coupled to the sides of this micro-fluidic chip without any lenses, which effectively acts as a multimode slab waveguide, where the excitation light is guided to excite the fluorescent targets within the micro-fluidic chip. Since the excitation light propagates perpendicular to the detection path, an inexpensive plastic absorption filter is able to reject most of the scattered light and create a decent dark-field background for fluorescent imaging. With this excitation geometry, the cell-phone camera can record fluorescent movies of the particles/cells as they are flowing through the microchannel. The digital frames of these fluorescent movies are then rapidly processed to quantify the count and the density of the labeled particles/cells within the solution under test. With a similar opto-fluidic design, we have recently demonstrated imaging and automated counting of stationary blood cells (e.g., labeled white blood cells or unlabeled red blood cells) loaded within a disposable cell counting chamber. We tested the performance of this cell-phone based imaging cytometry and blood analysis platform

  14. Quantitative image cytometry measurements of lipids, DNA, CD45 and cytokeratin for circulating tumor cell identification in a model system

    Science.gov (United States)

    Futia, Gregory L.; Qamar, Lubna; Behbakht, Kian; Gibson, Emily A.

    2016-04-01

    Circulating tumor cell (CTC) identification has applications in both early detection and monitoring of solid cancers. The rarity of CTCs, expected at ~1-50 CTCs per million nucleated blood cells (WBCs), requires identifying methods based on biomarkers with high sensitivity and specificity for accurate identification. Discovery of biomarkers with ever higher sensitivity and specificity to CTCs is always desirable to potentially find more CTCs in cancer patients thus increasing their clinical utility. Here, we investigate quantitative image cytometry measurements of lipids with the biomarker panel of DNA, Cytokeratin (CK), and CD45 commonly used to identify CTCs. We engineered a device for labeling suspended cell samples with fluorescent antibodies and dyes. We used it to prepare samples for 4 channel confocal laser scanning microscopy. The total data acquired at high resolution from one sample is ~ 1.3 GB. We developed software to perform the automated segmentation of these images into regions of interest (ROIs) containing individual cells. We quantified image features of total signal, spatial second moment, spatial frequency second moment, and their product for each ROI. We performed measurements on pure WBCs, cancer cell line MCF7 and mixed samples. Multivariable regressions and feature selection were used to determine combination features that are more sensitive and specific than any individual feature separately. We also demonstrate that computation of spatial characteristics provides higher sensitivity and specificity than intensity alone. Statistical models allowed quantification of the required sensitivity and specificity for detecting small levels of CTCs in a human blood sample.

  15. High-resolution cytometry represents the main technology used in the laboratory of molecular cytology and cytometry

    Czech Academy of Sciences Publication Activity Database

    Kozubek, Stanislav; Bártová, Eva; Lukášová, Emilie; Falk, Martin; Ondřej, Vladan; Kozubek, Michal; Kroupová, Jana; Matula, Pe.; Matula, Pa.

    2006-01-01

    Roč. 39, č. 5 (2006), s. 341-341 ISSN 0960-7722. [Cytomics Emerging from Cytometry 16th Annual Meeting of the german Society for cytometry. 18.10.2006-21.10.2006, Leipzig] Institutional research plan: CEZ:AV0Z50040507 Keywords : high-resolution cytometry * cytogenetics * epigenetics Subject RIV: BO - Biophysics

  16. Selective-imaging camera

    Science.gov (United States)

    Szu, Harold; Hsu, Charles; Landa, Joseph; Cha, Jae H.; Krapels, Keith A.

    2015-05-01

    How can we design cameras that image selectively in Full Electro-Magnetic (FEM) spectra? Without selective imaging, we cannot use, for example, ordinary tourist cameras to see through fire, smoke, or other obscurants contributing to creating a Visually Degraded Environment (VDE). This paper addresses a possible new design of selective-imaging cameras at firmware level. The design is consistent with physics of the irreversible thermodynamics of Boltzmann's molecular entropy. It enables imaging in appropriate FEM spectra for sensing through the VDE, and displaying in color spectra for Human Visual System (HVS). We sense within the spectra the largest entropy value of obscurants such as fire, smoke, etc. Then we apply a smart firmware implementation of Blind Sources Separation (BSS) to separate all entropy sources associated with specific Kelvin temperatures. Finally, we recompose the scene using specific RGB colors constrained by the HVS, by up/down shifting Planck spectra at each pixel and time.

  17. Prognostic Impact of DNA-Image-Cytometry in Neuroendocrine (Carcinoid Tumours

    Directory of Open Access Journals (Sweden)

    H. Raatz

    2004-01-01

    Full Text Available Establishing prognosis proves particularly difficult with neuroendocrine tumours (NETs as a benign looking histology can be associated with a malignant behaviour. In order to identify prognostic factors we examined 44 gastrointestinal and pulmonary, paraffin‐embedded NETs histologically and immunohistochemically. DNA‐image‐cytometry was used to examine 40 of these. We found that poor differentiation (corresponding to a Soga and Tazawa type D and infiltrative growth correlated with a poorer prognosis. Moreover, parameters determined by diagnostic DNA cytometry like the 5c‐exceeding rate, the 2c‐deviation index, DNA‐grade of malignancy, DNA‐entropy and the type of DNA histogram were found to be of prognostic relevance. Morphometric parameters like the form factor and the mean nuclear area were relevant for survival, tumour recurrence and metastasis. However, in the multivariate analysis the only independent risk factor was the histological differentiation. The 5c‐exceeding rate is a good objective risk factor, which can be used particularly in cases in which only a fine needle biopsie is available. Direct comparison of the histology and the 5c‐exceeding rate in the multivariate analysis suggests that the 5c‐exceeding rate taken as sole prognostic factor might be of higher prognostic relevance than the histology but larger studies are needed to confirm this.

  18. Direct concentration and viability measurement of yeast in corn mash using a novel imaging cytometry method.

    Science.gov (United States)

    Chan, Leo L; Lyettefi, Emily J; Pirani, Alnoor; Smith, Tim; Qiu, Jean; Lin, Bo

    2011-08-01

    Worldwide awareness of fossil-fuel depletion and global warming has been increasing over the last 30 years. Numerous countries, including the USA and Brazil, have introduced large-scale industrial fermentation facilities for bioethanol, biobutanol, or biodiesel production. Most of these biofuel facilities perform fermentation using standard baker's yeasts that ferment sugar present in corn mash, sugar cane, or other glucose media. In research and development in the biofuel industry, selection of yeast strains (for higher ethanol tolerance) and fermentation conditions (yeast concentration, temperature, pH, nutrients, etc.) can be studied to optimize fermentation performance. Yeast viability measurement is needed to identify higher ethanol-tolerant yeast strains, which may prolong the fermentation cycle and increase biofuel output. In addition, yeast concentration may be optimized to improve fermentation performance. Therefore, it is important to develop a simple method for concentration and viability measurement of fermenting yeast. In this work, we demonstrate an imaging cytometry method for concentration and viability measurements of yeast in corn mash directly from operating fermenters. It employs an automated cell counter, a dilution buffer, and staining solution from Nexcelom Bioscience to perform enumeration. The proposed method enables specific fluorescence detection of viable and nonviable yeasts, which can generate precise results for concentration and viability of yeast in corn mash. This method can provide an essential tool for research and development in the biofuel industry and may be incorporated into manufacturing to monitor yeast concentration and viability efficiently during the fermentation process.

  19. A method for the interpretation of flow cytometry data using genetic algorithms

    Directory of Open Access Journals (Sweden)

    Cesar Angeletti

    2018-01-01

    Full Text Available Background: Flow cytometry analysis is the method of choice for the differential diagnosis of hematologic disorders. It is typically performed by a trained hematopathologist through visual examination of bidimensional plots, making the analysis time-consuming and sometimes too subjective. Here, a pilot study applying genetic algorithms to flow cytometry data from normal and acute myeloid leukemia subjects is described. Subjects and Methods: Initially, Flow Cytometry Standard files from 316 normal and 43 acute myeloid leukemia subjects were transformed into multidimensional FITS image metafiles. Training was performed through introduction of FITS metafiles from 4 normal and 4 acute myeloid leukemia in the artificial intelligence system. Results: Two mathematical algorithms termed 018330 and 025886 were generated. When tested against a cohort of 312 normal and 39 acute myeloid leukemia subjects, both algorithms combined showed high discriminatory power with a receiver operating characteristic (ROC curve of 0.912. Conclusions: The present results suggest that machine learning systems hold a great promise in the interpretation of hematological flow cytometry data.

  20. An automated quantitative DNA image cytometry system detects abnormal cells in cervical cytology with high sensitivity.

    Science.gov (United States)

    Wong, O G; Ho, M W; Tsun, O K; Ng, A K; Tsui, E Y; Chow, J N; Ip, P P; Cheung, A N

    2018-03-26

    To evaluate the performance of an automated DNA-image-cytometry system as a tool to detect cervical carcinoma. Of 384 liquid-based cervical cytology samples with available biopsy follow-up were analyzed by both the Imager System and a high-risk HPV test (Cobas). The sensitivity and specificity of Imager System for detecting biopsy proven high-grade squamous intraepithelial lesion (HSIL, cervical intraepithelial neoplasia [CIN]2-3) and carcinoma were 89.58% and 56.25%, respectively, compared to 97.22% and 23.33% of HPV test but additional HPV 16/18 genotyping increased the specificity to 69.58%. The sensitivity and specificity of the Imager System for predicting HSIL+ (CIN2-3+) lesions among atypical squamous cells of undetermined significance samples were 80.00% and 70.53%, respectively, compared to 100% and 11.58% of HPV test whilst the HPV 16/18 genotyping increased the specificity to 77.89%. Among atypical squamous cells-cannot exclude HSIL, the sensitivity and specificity of Imager System for predicting HSIL+ (CIN2-3+) lesions upon follow up were 82.86% and 33.33%%, respectively, compared to 97.14% and 4.76% of HPV test and the HPV 16/18 genotyping increased the specificity to 19.05%. Among low-grade squamous intraepithelial lesion cases, the sensitivity and specificity of the Imager System for predicting HSIL+ (CIN2-3+) lesions were 66.67% and 35.71%%, respectively, compared to 66.67% and 29.76% of HPV test while HPV 16/18 genotyping increased the specificity to 79.76%. The overall results of imager and high-risk HPV test agreed in 69.43% (268) of all samples. The automated imager system and HPV 16/18 genotyping can enhance the specificity of detecting HSIL+ (CIN2-3+) lesions. © 2018 John Wiley & Sons Ltd.

  1. SCENERY: a web application for (causal) network reconstruction from cytometry data

    KAUST Repository

    Papoutsoglou, Georgios

    2017-05-08

    Flow and mass cytometry technologies can probe proteins as biological markers in thousands of individual cells simultaneously, providing unprecedented opportunities for reconstructing networks of protein interactions through machine learning algorithms. The network reconstruction (NR) problem has been well-studied by the machine learning community. However, the potentials of available methods remain largely unknown to the cytometry community, mainly due to their intrinsic complexity and the lack of comprehensive, powerful and easy-to-use NR software implementations specific for cytometry data. To bridge this gap, we present Single CEll NEtwork Reconstruction sYstem (SCENERY), a web server featuring several standard and advanced cytometry data analysis methods coupled with NR algorithms in a user-friendly, on-line environment. In SCENERY, users may upload their data and set their own study design. The server offers several data analysis options categorized into three classes of methods: data (pre)processing, statistical analysis and NR. The server also provides interactive visualization and download of results as ready-to-publish images or multimedia reports. Its core is modular and based on the widely-used and robust R platform allowing power users to extend its functionalities by submitting their own NR methods. SCENERY is available at scenery.csd.uoc.gr or http://mensxmachina.org/en/software/.

  2. Mercury effects on Thalassiosira weissflogii: Applications of two-photon excitation chlorophyll fluorescence lifetime imaging and flow cytometry

    International Nuclear Information System (INIS)

    Wu Yun; Zeng Yan; Qu, Jianan Y.; Wang Wenxiong

    2012-01-01

    The toxic effects of inorganic mercury [Hg(II)] and methylmercury (MeHg) on the photosynthesis and population growth in a marine diatom Thalassiosira weissflogii were investigated using two methods: two-photon excitation fluorescence lifetime imaging (FLIM) and flow cytometry (FCM). For photosynthesis, Hg(II) exposure increased the average chlorophyll fluorescence lifetime, whereas such increment was not found under MeHg stress. This may be caused by the inhibitory effect of Hg(II) instead of MeHg on the electron transport chain. For population growth, modeled specific growth rate data showed that the reduction in population growth by Hg(II) mainly resulted from an increased number of injured cells, while the live cells divided at the normal rates. However, MeHg inhibitory effects on population growth were contributed by the reduced division rates of all cells. Furthermore, the cell images and the FCM data reflected the morphological changes of diatom cells under Hg(II)/MeHg exposure vividly and quantitatively. Our results demonstrated that the toxigenicity mechanisms between Hg(II) and MeHg were different in the algal cells.

  3. Absence of DNA double-strand breaks in human peripheral blood mononuclear cells after 3 Tesla magnetic resonance imaging assessed by γH2AX flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Fasshauer, Martin; Staab, Wieland; Sohns, Jan M.; Ritter, Christian; Lotz, Joachim [Goettingen Heart Center, Department of Diagnostic and Interventional Radiology, University Medical Center Goettingen (Germany); German Centre for Cardiovascular Research (DZHK), Goettingen (Germany); Kruewel, Thomas; Stahnke, Vera C. [Goettingen Heart Center, Department of Diagnostic and Interventional Radiology, University Medical Center Goettingen (Germany); Zapf, Antonia [University Medical Center Goettingen, Department of Medical Statistics, Goettingen (Germany); Rave-Fraenk, Margret [University Medical Center Goettingen, Department of Radiotherapy and Radiooncology, Goettingen (Germany); Steinmetz, Michael [German Centre for Cardiovascular Research (DZHK), Goettingen (Germany); Goettingen Heart Center, Department of Pediatric Cardiology and Intensive Care Medicine, University Medical Center Goettingen (Germany); Unterberg-Buchwald, Christina [Goettingen Heart Center, Department of Diagnostic and Interventional Radiology, University Medical Center Goettingen (Germany); German Centre for Cardiovascular Research (DZHK), Goettingen (Germany); Goettingen Heart Center, Department of Cardiology and Pneumology, University Medical Center Goettingen (Germany); Schuster, Andreas [German Centre for Cardiovascular Research (DZHK), Goettingen (Germany); Goettingen Heart Center, Department of Cardiology and Pneumology, University Medical Center Goettingen (Germany)

    2018-03-15

    Magnetic resonance imaging (MRI) is regarded as a non-harming and non-invasive imaging modality with high tissue contrast and almost no side effects. Compared to other cross-sectional imaging modalities, MRI does not use ionising radiation. Recently, however, strong magnetic fields as applied in clinical MRI scanners have been suspected to induce DNA double-strand breaks in human lymphocytes. In this study we investigated the impact of 3-T cardiac MRI examinations on the induction of DNA double-strand breaks in peripheral mononuclear cells by γH2AX staining and flow cytometry analysis. The study cohort consisted of 73 healthy non-smoking volunteers with 36 volunteers undergoing CMRI and 37 controls without intervention. Differences between the two cohorts were analysed by a mixed linear model with repeated measures. Both cohorts showed a significant increase in the γH2AX signal from baseline to post-procedure of 6.7 % (SD 7.18 %) and 7.8 % (SD 6.61 %), respectively. However, the difference between the two groups was not significant. Based on our study, γH2AX flow cytometry shows no evidence that 3-T MRI examinations as used in cardiac scans impair DNA integrity in peripheral mononuclear cells. (orig.)

  4. Evaluation of Tumor Heterogeneity of Prostate Carcinoma by Flow- and Image DNA Cytometry and Histopathological Grading

    Directory of Open Access Journals (Sweden)

    Naining Wang

    2000-01-01

    Full Text Available Background. Heterogeneity of prostate carcinoma is one of the reasons for pretreatment underestimation of tumor aggressiveness. We studied tumor heterogeneity and the probability of finding the highest tumor grade and DNA aneuploidy with relation to the number of biopsies. Material and methods. Specimens simulating core biopsies from five randomly selected tumor areas from each of 16 Böcking’s grade II and 23 grade III prostate carcinomas were analyzed for tumor grade and DNA ploidy by flow‐ and fluorescence image cytometry (FCM, FICM. Cell cycle composition was measured by FCM. Results. By determination of ploidy and cell cycle composition, morphologically defined tumors can further be subdivided. Heterogeneity of tumor grade and DNA ploidy (FCM was 54% and 50%. Coexistence of diploid tumor cells in aneuploid specimens represents another form of tumor heterogeneity. The proportion of diploid tumor cells decreased significantly with tumor grade and with increase in the fraction of proliferating cell of the aneuploid tumor part. The probability of estimating the highest tumor grade or aneuploidy increased from 40% for one biopsy to 95% for 5 biopsies studied. By combining the tumor grade with DNA ploidy, the probability of detecting a highly aggressive tumor increased from 40% to 70% and 90% for one and two biopsies, respectively. Conclusion. Specimens of the size of core biopsies can be used for evaluation of DNA ploidy and cell cycle composition. Underestimation of aggressiveness of prostate carcinoma due to tumor heterogeneity is minimized by simultaneous study of the tumor grade and DNA ploidy more than by increasing the number of biopsies. The biological significance of coexistent diploid tumor cell in aneuploid lesions remains to be evaluated.

  5. Feasibility study of stain-free classification of cell apoptosis based on diffraction imaging flow cytometry and supervised machine learning techniques.

    Science.gov (United States)

    Feng, Jingwen; Feng, Tong; Yang, Chengwen; Wang, Wei; Sa, Yu; Feng, Yuanming

    2018-06-01

    This study was to explore the feasibility of prediction and classification of cells in different stages of apoptosis with a stain-free method based on diffraction images and supervised machine learning. Apoptosis was induced in human chronic myelogenous leukemia K562 cells by cis-platinum (DDP). A newly developed technique of polarization diffraction imaging flow cytometry (p-DIFC) was performed to acquire diffraction images of the cells in three different statuses (viable, early apoptotic and late apoptotic/necrotic) after cell separation through fluorescence activated cell sorting with Annexin V-PE and SYTOX® Green double staining. The texture features of the diffraction images were extracted with in-house software based on the Gray-level co-occurrence matrix algorithm to generate datasets for cell classification with supervised machine learning method. Therefore, this new method has been verified in hydrogen peroxide induced apoptosis model of HL-60. Results show that accuracy of higher than 90% was achieved respectively in independent test datasets from each cell type based on logistic regression with ridge estimators, which indicated that p-DIFC system has a great potential in predicting and classifying cells in different stages of apoptosis.

  6. Phaedra, a protocol-driven system for analysis and validation of high-content imaging and flow cytometry.

    Science.gov (United States)

    Cornelissen, Frans; Cik, Miroslav; Gustin, Emmanuel

    2012-04-01

    High-content screening has brought new dimensions to cellular assays by generating rich data sets that characterize cell populations in great detail and detect subtle phenotypes. To derive relevant, reliable conclusions from these complex data, it is crucial to have informatics tools supporting quality control, data reduction, and data mining. These tools must reconcile the complexity of advanced analysis methods with the user-friendliness demanded by the user community. After review of existing applications, we realized the possibility of adding innovative new analysis options. Phaedra was developed to support workflows for drug screening and target discovery, interact with several laboratory information management systems, and process data generated by a range of techniques including high-content imaging, multicolor flow cytometry, and traditional high-throughput screening assays. The application is modular and flexible, with an interface that can be tuned to specific user roles. It offers user-friendly data visualization and reduction tools for HCS but also integrates Matlab for custom image analysis and the Konstanz Information Miner (KNIME) framework for data mining. Phaedra features efficient JPEG2000 compression and full drill-down functionality from dose-response curves down to individual cells, with exclusion and annotation options, cell classification, statistical quality controls, and reporting.

  7. An introduction to mass cytometry: fundamentals and applications.

    Science.gov (United States)

    Tanner, Scott D; Baranov, Vladimir I; Ornatsky, Olga I; Bandura, Dmitry R; George, Thaddeus C

    2013-05-01

    Mass cytometry addresses the analytical challenges of polychromatic flow cytometry by using metal atoms as tags rather than fluorophores and atomic mass spectrometry as the detector rather than photon optics. The many available enriched stable isotopes of the transition elements can provide up to 100 distinguishable reporting tags, which can be measured simultaneously because of the essential independence of detection provided by the mass spectrometer. We discuss the adaptation of traditional inductively coupled plasma mass spectrometry to cytometry applications. We focus on the generation of cytometry-compatible data and on approaches to unsupervised multivariate clustering analysis. Finally, we provide a high-level review of some recent benchmark reports that highlight the potential for massively multi-parameter mass cytometry.

  8. One-dimensional acoustic standing waves in rectangular channels for flow cytometry.

    Science.gov (United States)

    Austin Suthanthiraraj, Pearlson P; Piyasena, Menake E; Woods, Travis A; Naivar, Mark A; Lόpez, Gabriel P; Graves, Steven W

    2012-07-01

    Flow cytometry has become a powerful analytical tool for applications ranging from blood diagnostics to high throughput screening of molecular assemblies on microsphere arrays. However, instrument size, expense, throughput, and consumable use limit its use in resource poor areas of the world, as a component in environmental monitoring, and for detection of very rare cell populations. For these reasons, new technologies to improve the size and cost-to-performance ratio of flow cytometry are required. One such technology is the use of acoustic standing waves that efficiently concentrate cells and particles to the center of flow channels for analysis. The simplest form of this method uses one-dimensional acoustic standing waves to focus particles in rectangular channels. We have developed one-dimensional acoustic focusing flow channels that can be fabricated in simple capillary devices or easily microfabricated using photolithography and deep reactive ion etching. Image and video analysis demonstrates that these channels precisely focus single flowing streams of particles and cells for traditional flow cytometry analysis. Additionally, use of standing waves with increasing harmonics and in parallel microfabricated channels is shown to effectively create many parallel focused streams. Furthermore, we present the fabrication of an inexpensive optical platform for flow cytometry in rectangular channels and use of the system to provide precise analysis. The simplicity and low-cost of the acoustic focusing devices developed here promise to be effective for flow cytometers that have reduced size, cost, and consumable use. Finally, the straightforward path to parallel flow streams using one-dimensional multinode acoustic focusing, indicates that simple acoustic focusing in rectangular channels may also have a prominent role in high-throughput flow cytometry. Copyright © 2012 Elsevier Inc. All rights reserved.

  9. Flow Cytometry Section

    Data.gov (United States)

    Federal Laboratory Consortium — The primary goal of the Flow Cytometry Section is to provide the services of state-of-the-art multi-parameter cellular analysis and cell sorting for researchers and...

  10. Detecting fetomaternal hemorrhage by flow cytometry

    DEFF Research Database (Denmark)

    Dziegiel, Morten Hanefeld; Nielsen, Leif Kofoed; Berkowicz, Adela

    2006-01-01

    The aim of this review is to summarize the most recent developments in the area of detection of fetomaternal hemorrhage by flow cytometry.......The aim of this review is to summarize the most recent developments in the area of detection of fetomaternal hemorrhage by flow cytometry....

  11. Quantification of green fluorescent protein by in vivo imaging, PCR, and flow cytometry: comparison of transgenic strains and relevance for fetal cell microchimerism.

    Science.gov (United States)

    Fujiki, Yutaka; Tao, Kai; Bianchi, Diana W; Giel-Moloney, Maryann; Leiter, Andrew B; Johnson, Kirby L

    2008-02-01

    Animal models are increasingly being used for the assessment of fetal cell microchimerism in maternal tissue. We wished to determine the optimal transgenic mouse strain and analytic technique to facilitate the detection of rare transgenic microchimeric fetal cells amongst a large number of maternal wild-type cells. We evaluated two strains of mice transgenic for the enhanced green fluorescent protein (EGFP): a commercially available, commonly used strain (C57BL/6-Tg(ACTB-EGFP)10sb/J) (CAG) and a newly created strain (ROSA26-EGFP) using three different techniques: in vivo and ex vivo fluorescent imaging (for whole body and dissected organs, respectively), PCR amplification of gfp, and flow cytometry (FCM). By fluorescent imaging, organs from CAG mice were 10-fold brighter than organs from ROSA26-EGFP mice (P characteristics that make it useful under specific experimental circumstances. The CAG mouse model is preferable when experiments require brighter cells, whereas ROSA26-EGFP is more appropriate when uniform or ubiquitous expression is more important than brightness. Investigators must carefully select the transgenic strain most suited to the experimental design to obtain the most consistent and reproducible data. In vivo imaging allows for phenotypic evaluation of whole animals and intact organs; however, we did not evaluate its utility for the detection of rare, fetal microchimeric cells in the maternal organs. Finally, while PCR amplification of a paternally inherited transgene does allow for the quantitative determination of rare microchimeric cells, FCM allows for both quantitative and qualitative evaluations of fetal cells at very high sensitivity in a plethora of maternal organs. (c) 2008 International Society for Analytical Cytology

  12. Comparative exploration of multidimensional flow cytometry software: a model approach evaluating T cell polyfunctional behavior.

    Science.gov (United States)

    Spear, Timothy T; Nishimura, Michael I; Simms, Patricia E

    2017-08-01

    Advancement in flow cytometry reagents and instrumentation has allowed for simultaneous analysis of large numbers of lineage/functional immune cell markers. Highly complex datasets generated by polychromatic flow cytometry require proper analytical software to answer investigators' questions. A problem among many investigators and flow cytometry Shared Resource Laboratories (SRLs), including our own, is a lack of access to a flow cytometry-knowledgeable bioinformatics team, making it difficult to learn and choose appropriate analysis tool(s). Here, we comparatively assess various multidimensional flow cytometry software packages for their ability to answer a specific biologic question and provide graphical representation output suitable for publication, as well as their ease of use and cost. We assessed polyfunctional potential of TCR-transduced T cells, serving as a model evaluation, using multidimensional flow cytometry to analyze 6 intracellular cytokines and degranulation on a per-cell basis. Analysis of 7 parameters resulted in 128 possible combinations of positivity/negativity, far too complex for basic flow cytometry software to analyze fully. Various software packages were used, analysis methods used in each described, and representative output displayed. Of the tools investigated, automated classification of cellular expression by nonlinear stochastic embedding (ACCENSE) and coupled analysis in Pestle/simplified presentation of incredibly complex evaluations (SPICE) provided the most user-friendly manipulations and readable output, evaluating effects of altered antigen-specific stimulation on T cell polyfunctionality. This detailed approach may serve as a model for other investigators/SRLs in selecting the most appropriate software to analyze complex flow cytometry datasets. Further development and awareness of available tools will help guide proper data analysis to answer difficult biologic questions arising from incredibly complex datasets. © Society

  13. Semiautomated analysis of embryoscope images: Using localized variance of image intensity to detect embryo developmental stages.

    Science.gov (United States)

    Mölder, Anna; Drury, Sarah; Costen, Nicholas; Hartshorne, Geraldine M; Czanner, Silvester

    2015-02-01

    Embryo selection in in vitro fertilization (IVF) treatment has traditionally been done manually using microscopy at intermittent time points during embryo development. Novel technique has made it possible to monitor embryos using time lapse for long periods of time and together with the reduced cost of data storage, this has opened the door to long-term time-lapse monitoring, and large amounts of image material is now routinely gathered. However, the analysis is still to a large extent performed manually, and images are mostly used as qualitative reference. To make full use of the increased amount of microscopic image material, (semi)automated computer-aided tools are needed. An additional benefit of automation is the establishment of standardization tools for embryo selection and transfer, making decisions more transparent and less subjective. Another is the possibility to gather and analyze data in a high-throughput manner, gathering data from multiple clinics and increasing our knowledge of early human embryo development. In this study, the extraction of data to automatically select and track spatio-temporal events and features from sets of embryo images has been achieved using localized variance based on the distribution of image grey scale levels. A retrospective cohort study was performed using time-lapse imaging data derived from 39 human embryos from seven couples, covering the time from fertilization up to 6.3 days. The profile of localized variance has been used to characterize syngamy, mitotic division and stages of cleavage, compaction, and blastocoel formation. Prior to analysis, focal plane and embryo location were automatically detected, limiting precomputational user interaction to a calibration step and usable for automatic detection of region of interest (ROI) regardless of the method of analysis. The results were validated against the opinion of clinical experts. © 2015 International Society for Advancement of Cytometry. © 2015 International

  14. Automated scoring of lymphocyte micronuclei by the MetaSystems Metafer image cytometry system and its application in studies of human mutagen sensitivity and biodosimetry of genotoxin exposure

    Czech Academy of Sciences Publication Activity Database

    Rössnerová, Andrea; Špátová, Milada; Schunck, CH.; Šrám, Radim

    2011-01-01

    Roč. 26, č. 1 (2011), s. 169-175 ISSN 0267-8357 R&D Projects: GA MŽP(CZ) SP/1B3/8/08; GA AV ČR 1QS500390506 Institutional research plan: CEZ:AV0Z50390512 Keywords : automated micronucleus assay * environmental exposure * Metasystems Metafer image cytometry system Subject RIV: DN - Health Impact of the Environment Quality Impact factor: 3.183, year: 2011

  15. Application of an Image Cytometry Protocol for Cellular and Mitochondrial Phenotyping on Fibroblasts from Patients with Inherited Disorders

    DEFF Research Database (Denmark)

    Fernandez-Guerra, Paula; Lund, Martin; Corydon, T J

    2015-01-01

    Cellular phenotyping of human dermal fibroblasts (HDFs) from patients with inherited diseases provides invaluable information for diagnosis, disease aetiology, prognosis and assessing of treatment options. Here we present a cell phenotyping protocol using image cytometry that combines measurements...... on a parallel one. We analysed HDFs from healthy individuals after treatment with various concentrations of hydrogen peroxide (H2O2) for different intervals, to mimic the physiological effects of oxidative stress. Our results show that cell number, viability, TRS and MMP decreased, while MSL increased both...... in a time- and concentration-dependent manner. To assess the use of our protocol for analysis of HDFs from patients with inherited diseases, we analysed HDFs from two patients with very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency (VLCADD), one with a severe clinical phenotype and one with a mild...

  16. Teaching Phagocytosis Using Flow Cytometry

    Directory of Open Access Journals (Sweden)

    John Boothby

    2009-12-01

    Full Text Available Investigative microbiology on protists in a basic teaching laboratory environment is limited by student skill level, ease of microbial culture and manipulation, instrumentation, and time. The flow cytometer is gaining use as a mainstream instrument in research and clinical laboratories, but has had minimal application in teaching laboratories. Although the cost of a flow cytometer is currently prohibitive for many microbiology teaching environments and the number of trained instructors and teaching materials is limited, in many ways the flow cytometer is an ideal instrument for teaching basic microbiology. We report here on a laboratory module to study phagocytosis in Tetrahymena sp. using flow cytometry in a basic microbiology teaching laboratory. Students and instructors found the flow cytometry data analysis program, Paint-A-GatePRO-TM, to be very intuitive and easy to learn within a short period of time. Assessment of student learning about Tetrahymena sp., phagocytosis, flow cytometry, and investigative microbiology using an inquiry-based format demonstrated an overall positive response from students.

  17. Immuno flow cytometry in marine phytoplankton research

    NARCIS (Netherlands)

    Peperzak, L; Vrieling, EG; Sandee, B; Rutten, T

    The developments in the combination of flow cytometry and immunology as a tool to identify, count and examine marine phytoplankton cells are reviewed. The concepts of immunology and now cytometry are described. A distinction is made between quantitative and qualitative immunofluorescence.

  18. Stress response assessment of Lactobacillus sakei strains selected as potential autochthonous starter cultures by flow cytometry and nucleic acid double-staining analyses.

    Science.gov (United States)

    Bonomo, M G; Milella, L; Martelli, G; Salzano, G

    2013-09-01

    The aim of this study was to apply the flow cytometry to Lactobacillus sakei strains, selected as potential autochthonous starters, to investigate dynamics and physiological heterogeneity of microbial behaviour under different stress conditions. A simultaneous nucleic acid double-staining assay was applied to discriminate cell populations in different physiological states after exposure to heat (50 and 55°C) and acid (pH 2·5 and 3·0) stresses. Alive cells with intact membranes, damaged cells still alive but with injured membranes, so with even a recovery ability, and dead cells with a permanent membrane damage were differentiated with a significant increase in damaged cells after stronger stress treatments. The existence and characteristics of subpopulations displaying heterogeneity in particular conditions are highly relevant, because specific subpopulations may show improved survival, changes and dynamics under stress conditions. This assay has potential for physiological research on lactic acid bacteria and for application in the food industry. The assessment of intermediate physiological states in Lb. sakei strains with recovery possibility could be an important criterion for application of potential starter cultures. Application of flow cytometry and characterization of sorted subpopulations may contribute to further understanding of diversity and heterogeneity in physiology of bacterial populations. © 2013 The Society for Applied Microbiology.

  19. Confocal Microscopy and Flow Cytometry System Performance: Assessment of QA Parameters that affect data Quanitification

    Science.gov (United States)

    Flow and image cytometers can provide useful quantitative fluorescence data. We have devised QA tests to be used on both a flow cytometer and a confocal microscope to assure that the data is accurate, reproducible and precise. Flow Cytometry: We have provided two simple perform...

  20. Immense random colocalization, revealed by automated high content image cytometry, seriously questions FISH as gold standard for detecting EML4-ALK fusion.

    Science.gov (United States)

    Smuk, Gábor; Tornóczky, Tamás; Pajor, László; Chudoba, Ilse; Kajtár, Béla; Sárosi, Veronika; Pajor, Gábor

    2018-05-19

    EML4-ALK gene fusion (inv2(p21p23)) of non-small cell lung cancer (NSCLC) predisposes to tyrosine kinase inhibitor treatment. One of the gold standard diagnostics is the dual color (DC) break-apart (BA) FISH technique, however, the unusual closeness of the involved genes has been suggested to raise likelihood of random co-localization (RCL) of signals. Although this is suspected to decrease sensitivity (often to as low as 40-70%), the exact level and effect of RCL has not been revealed thus far. Signal distances were analyzed to the 0.1 µm precision in more than 25,000 nuclei, via automated high content-image cytometry. Negative and positive controls were created using conventional DC BA-, and inv2(p21p23) mimicking probe-sets, respectively. Average distance between red and green signals was 9.72 pixels (px) (±5.14px) and 3.28px (±2.44px), in positives and negatives, respectively; overlap in distribution being 41%. Specificity and sensitivity of correctly determining ALK status was 97% and 29%, respectively. When investigating inv2(p21p23) with DC BA FISH, specificity is high, but seven out of ten aberrant nuclei are inevitably falsely classified as negative, due to the extreme level of RCL. Together with genetic heterogeneity and dilution effect of non-tumor cells in NSCLC, this immense analytical false negativity is the primary cause behind the often described low diagnostic sensitivity. These results convincingly suggest that if FISH is to remain a gold standard for detecting the therapy relevant inv(2), either a modified evaluation protocol, or a more reliable probe-design should be considered than the current DC BA one. © 2018 International Society for Advancement of Cytometry. © 2018 International Society for Advancement of Cytometry.

  1. ggCyto: Next Generation Open-Source Visualization Software for Cytometry.

    Science.gov (United States)

    Van, Phu; Jiang, Wenxin; Gottardo, Raphael; Finak, Greg

    2018-06-01

    Open source software for computational cytometry has gained in popularity over the past few years. Efforts such as FlowCAP, the Lyoplate and Euroflow projects have highlighted the importance of efforts to standardize both experimental and computational aspects of cytometry data analysis. The R/BioConductor platform hosts the largest collection of open source cytometry software covering all aspects of data analysis and providing infrastructure to represent and analyze cytometry data with all relevant experimental, gating, and cell population annotations enabling fully reproducible data analysis. Data visualization frameworks to support this infrastructure have lagged behind. ggCyto is a new open-source BioConductor software package for cytometry data visualization built on ggplot2 that enables ggplot-like functionality with the core BioConductor flow cytometry data structures. Amongst its features are the ability to transform data and axes on-the-fly using cytometry-specific transformations, plot faceting by experimental meta-data variables, and partial matching of channel, marker and cell populations names to the contents of the BioConductor cytometry data structures. We demonstrate the salient features of the package using publicly available cytometry data with complete reproducible examples in a supplementary material vignette. https://bioconductor.org/packages/devel/bioc/html/ggcyto.html. gfinak@fredhutch.org. Supplementary data are available at Bioinformatics online and at http://rglab.org/ggcyto/.

  2. Measurement and Characterization of Apoptosis by Flow Cytometry.

    Science.gov (United States)

    Telford, William; Tamul, Karen; Bradford, Jolene

    2016-07-01

    Apoptosis is an important mechanism in cell biology, playing a critical regulatory role in virtually every organ system. It has been particularly well characterized in the immune system, with roles ranging from immature immune cell development and selection to down-regulation of the mature immune response. Apoptosis is also the primary mechanism of action of anti-cancer drugs. Flow cytometry has been the method of choice for analyzing apoptosis in suspension cells for more than 25 years. Numerous assays have been devised to measure both the earliest and latest steps in the apoptotic process, from the earliest signal-transduction events to the late morphological changes in cell shape and granularity, proteolysis, and chromatin condensation. These assays are particularly powerful when combined into multicolor assays determining several apoptotic characteristics simultaneously. The multiparametric nature of flow cytometry makes this technology particularly suited to measuring apoptosis. In this unit, we will describe the four main techniques for analyzing caspase activity in apoptotic cells, combined with annexin V and cell permeability analysis. These relatively simple multiparametric assays are powerful techniques for assessing cell death. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.

  3. Merging Mixture Components for Cell Population Identification in Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Greg Finak

    2009-01-01

    Full Text Available We present a framework for the identification of cell subpopulations in flow cytometry data based on merging mixture components using the flowClust methodology. We show that the cluster merging algorithm under our framework improves model fit and provides a better estimate of the number of distinct cell subpopulations than either Gaussian mixture models or flowClust, especially for complicated flow cytometry data distributions. Our framework allows the automated selection of the number of distinct cell subpopulations and we are able to identify cases where the algorithm fails, thus making it suitable for application in a high throughput FCM analysis pipeline. Furthermore, we demonstrate a method for summarizing complex merged cell subpopulations in a simple manner that integrates with the existing flowClust framework and enables downstream data analysis. We demonstrate the performance of our framework on simulated and real FCM data. The software is available in the flowMerge package through the Bioconductor project.

  4. Applications of flow cytometry in food microbiology

    International Nuclear Information System (INIS)

    Serrano Valerin, Pamela

    2014-01-01

    A compilation of data about cytometry and its applications is performed to analyze the impact on food microbiology. The technique of flow cytometry is described and the use in various fields of microbiology is analyzed. Flow cytometry future could be implemented in many clinical laboratories and food, considering the cost / benefit test to be done, because at the moment it has a high cost. The existence of new fluorochromes and monoclonal antibodies enable that many intracellular and extracellular cell parameters are detected in the future. The technique can be developed in the country in few years considering that the technique has improved the sensitivity and specificity of many tests [es

  5. On a selection method of imaging condition in scintigraphy

    International Nuclear Information System (INIS)

    Ikeda, Hozumi; Kishimoto, Kenji; Shimonishi, Yoshihiro; Ohmura, Masahiro; Kosakai, Kazuhisa; Ochi, Hironobu

    1992-01-01

    Selection of imaging condition in scintigraphy was evaluated using analytic hierarchy process. First, a method of the selection was led by determining at the points of image quantity and imaging time. Influence of image quality was thought to depend on changes of system resolution, count density, image size, and image density. Also influence of imaging time was thought to depend on changes of system sensitivity and data acquisition time. Phantom study was done for paired comparison of these selection factors, and relations of sample data and the factors, that is Rollo phantom images were taken by changing count density, image size, and image density. Image quality was shown by calculating the score of visual evaluation that done by comparing of a pair of images in clearer cold lesion on the scintigrams. Imaging time was shown by relative values for changes of count density. However, system resolution and system sensitivity were constant in this study. Next, using these values analytic hierarchy process was adapted for this selection of imaging conditions. We conclude that this selection of imaging conditions can be analyzed quantitatively using analytic hierarchy process and this analysis develops theoretical consideration of imaging technique. (author)

  6. Flow cytometry protocols

    National Research Council Canada - National Science Library

    Jaroszeski, Mark J; Heller, Richard

    1998-01-01

    ... are individually analyzed, and it is typical for flow cytometers to quantitatively process thousands of individual particles in a matter of seconds. This a powerful analytic feat particularly if one relates it to the time required to examine several thousand individual cells using a microscope. This leaves little doubt regarding why the field of flow cytometry has...

  7. Internalisation of polymeric nanosensors in mesenchymal stem cells: analysis by flow cytometry and confocal microscopy.

    Science.gov (United States)

    Coupland, Paul G; Fisher, Karen A; Jones, D Rhodri E; Aylott, Jonathan W

    2008-09-10

    The aim of this study was to demonstrate that flow cytometry and confocal microscopy could be applied in a complementary manner to analyse the internalisation of polymeric nanosensors in mesenchymal stem cells (MSC). The two techniques are able to provide en masse data analysis of nanosensors from large cell populations and detailed images of intracellular nanosensor localisation, respectively. The polyacrylamide nanosensors used in this investigation had been modified to contain free amine groups which were subsequently conjugated to Tat peptide, which acted as a delivery vector for nanosensor internalisation. Flow cytometry was used to confirm the health of MSC culture and assess the impact of nanosensor internalisation. MSC were characterised using fluorescently tagged CD cell surface markers that were also used to show that nanosensor internalisation did not negatively impact on MSC culture. Additionally it was shown that flow cytometry can be used to measure fluorophores located both on the cell surface and internalised within the cell. Complementary data was obtained using confocal microscopy to confirm nanosensor internalisation within MSC.

  8. The role of flow cytometry in the study of cell growth in the rat anterior pituitary gland

    Directory of Open Access Journals (Sweden)

    M Vitale

    2009-12-01

    Full Text Available Flow cytometry is a suitable technique for studying in vivo and in vitro the cell cycle kinetics of different animal and human tissues, both in normal and tumoral conditions. The rat anterior pituitary gland is a model to investigate cell growth and replication of differentiated, neuroendocrine cells, and we report current evidence on its cell cycle kinetics as well as on the role played by flow cytometry in this type of study. The proliferation potential of normal anterior pituitary cells is related to a number of different conditions, including heterogeneity of cell types, age and sex of donors, and circadian influences. In addition, the trend of cell proliferation in both in vivo and in vitro studies is similar, suggesting that cultured anterior pituitary elements may, at least in parts, retain growth features analogous to those of the intact gland. Sorting of selective cell types and analysis of the relation between proliferating anterior pituitary cells and the light-dark cycle have shown that flow cytometry may be useful to investigate the replication process of the gland. By using a combination of flow cytometry, light microscopic immunocytochemistry and morphometry, we have reported a peculiar trend of proliferation in prima- ry monolayer cultures of rat anterior pituitary gland, characterized by a non-linear reduction in their proliferation rate with advancing age, primarily dependent on a reduced transition of cells from the G0/G1- to the early S-phase pool. These studies indicate that flow cytometry offers insights into cell cycle check points of anterior pituitary cells, and suggest that it might be applied to the study of growth of selective pituitary elements, both in normal and tumoral conditions.

  9. Rapid detection of microbial contamination in grape juice by flow cytometry

    Directory of Open Access Journals (Sweden)

    Marielle Bouix

    1999-03-01

    Full Text Available This study presents an application of flow cytometry to evaluate rapidly the viable micro-organisms in grape juice. In this method, viable cells are firstly specitically labelled with a fluorescent reagent. The sample is then injected into the flow cytometer where the labelled micro-organisms are individually illuminated by a laser beam. The emission of fluorescence is measured. The system counts the number of fluorescent events and prints out a histogram of the fluorescence intensity which is characteristic of the micro-organism being analysed. In laboratory conditions, preliminary trials have been undertaken with an artificially inoculated grape juice with pure yeast and bacteria cultures. This method succeeded in counting simultaneously yeasts and bacteria within 15 minutes, with a high degree of sensitivity, 5.103 yeasts perml and 5.104 bacteria per ml. This technique can also be applied to the detection of mould contamination and the test has been done with Botrytis spores. The method makes direct cell counts possible and is capable of analysing 30 samples per hour. It can be automatised and easily used in industrial laboratory. During the last harvest, more than a thousand of must samples were controled using this technique. The results let to determine the yeast contamination level of a grape juice tank even before unloading. The results obtained by flow cytometry were compared to the plate count reference method. The correlation between cytometry and count by plate culture was 99 p. cent for the threshold of 5.1 04 yeasts/ml which seemed to point out a high contamination. By using this flow cytometry method during the harvest period, the results were supplied in real time. This allowed a rapid selection of the musts, depending upon the scale of their contamination and improved the quality of the wine by corrective actions.

  10. An active, collaborative approach to learning skills in flow cytometry.

    Science.gov (United States)

    Fuller, Kathryn; Linden, Matthew D; Lee-Pullen, Tracey; Fragall, Clayton; Erber, Wendy N; Röhrig, Kimberley J

    2016-06-01

    Advances in science education research have the potential to improve the way students learn to perform scientific interpretations and understand science concepts. We developed active, collaborative activities to teach skills in manipulating flow cytometry data using FlowJo software. Undergraduate students were given compensated clinical flow cytometry listmode output (FCS) files and asked to design a gating strategy to diagnose patients with different hematological malignancies on the basis of their immunophenotype. A separate cohort of research trainees was given uncompensated data files on which they performed their own compensation, calculated the antibody staining index, designed a sequential gating strategy, and quantified rare immune cell subsets. Student engagement, confidence, and perceptions of flow cytometry were assessed using a survey. Competency against the learning outcomes was assessed by asking students to undertake tasks that required understanding of flow cytometry dot plot data and gating sequences. The active, collaborative approach allowed students to achieve learning outcomes not previously possible with traditional teaching formats, for example, having students design their own gating strategy, without forgoing essential outcomes such as the interpretation of dot plots. In undergraduate students, favorable perceptions of flow cytometry as a field and as a potential career choice were correlated with student confidence but not the ability to perform flow cytometry data analysis. We demonstrate that this new pedagogical approach to teaching flow cytometry is beneficial for student understanding and interpretation of complex concepts. It should be considered as a useful new method for incorporating complex data analysis tasks such as flow cytometry into curricula. Copyright © 2016 The American Physiological Society.

  11. Microfluidic Impedance Flow Cytometry Enabling High-Throughput Single-Cell Electrical Property Characterization

    Science.gov (United States)

    Chen, Jian; Xue, Chengcheng; Zhao, Yang; Chen, Deyong; Wu, Min-Hsien; Wang, Junbo

    2015-01-01

    This article reviews recent developments in microfluidic impedance flow cytometry for high-throughput electrical property characterization of single cells. Four major perspectives of microfluidic impedance flow cytometry for single-cell characterization are included in this review: (1) early developments of microfluidic impedance flow cytometry for single-cell electrical property characterization; (2) microfluidic impedance flow cytometry with enhanced sensitivity; (3) microfluidic impedance and optical flow cytometry for single-cell analysis and (4) integrated point of care system based on microfluidic impedance flow cytometry. We examine the advantages and limitations of each technique and discuss future research opportunities from the perspectives of both technical innovation and clinical applications. PMID:25938973

  12. Choosing face: The curse of self in profile image selection.

    Science.gov (United States)

    White, David; Sutherland, Clare A M; Burton, Amy L

    2017-01-01

    People draw automatic social inferences from photos of unfamiliar faces and these first impressions are associated with important real-world outcomes. Here we examine the effect of selecting online profile images on first impressions. We model the process of profile image selection by asking participants to indicate the likelihood that images of their own face ("self-selection") and of an unfamiliar face ("other-selection") would be used as profile images on key social networking sites. Across two large Internet-based studies (n = 610), in line with predictions, image selections accentuated favorable social impressions and these impressions were aligned to the social context of the networking sites. However, contrary to predictions based on people's general expertise in self-presentation, other-selected images conferred more favorable impressions than self-selected images. We conclude that people make suboptimal choices when selecting their own profile pictures, such that self-perception places important limits on facial first impressions formed by others. These results underscore the dynamic nature of person perception in real-world contexts.

  13. Report of the results of the International Clinical Cytometry Society and American Society for Clinical Pathology workload survey of clinical flow cytometry laboratories.

    Science.gov (United States)

    Wolniak, Kristy; Goolsby, Charles; Choi, Sarah; Ali, Asma; Serdy, Nina; Stetler-Stevenson, Maryalice

    2017-11-01

    Thorough review of current workload, staffing, and testing practices in clinical laboratories allows for optimization of laboratory efficiency and quality. This information is largely missing with regard to clinical flow cytometry laboratories. The purpose of this survey is to provide comprehensive, current, and accurate data on testing practices and laboratory staffing in clinical laboratories performing flow cytometric studies. Survey data was collected from flow cytometry laboratories through the ASCP website. Data was collected on the workload during a 1-year time period of full-time and part-time technical and professional (M.D./D.O./Ph.D. or equivalent) flow cytometry employees. Workload was examined as number of specimens and tubes per full time equivalent (FTE) technical and professional staff. Test complexity, test result interpretation, and reporting practices were also evaluated. There were 205 respondent laboratories affiliated predominantly with academic and health system institutions. Overall, 1,132 FTE employees were reported with 29% professional FTE employees and 71% technical. Fifty-one percent of the testing performed was considered high complexity and 49% was low complexity. The average number of tubes per FTE technologist was 1,194 per year and the average number of specimens per FTE professional was 1,659 per year. The flow cytometry reports were predominantly written by pathologists (57%) and were typically written as a separate report (58%). This survey evaluates the overall status of the current practice of clinical flow cytometry and provides a comprehensive dataset as a framework to help laboratory departments, directors, and managers make appropriate, cost-effective staffing decisions. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

  14. Preparing a Minimum Information about a Flow Cytometry Experiment (MIFlowCyt) compliant manuscript using the International Society for Advancement of Cytometry (ISAC) FCS file repository (FlowRepository.org).

    Science.gov (United States)

    Spidlen, Josef; Breuer, Karin; Brinkman, Ryan

    2012-07-01

    FlowRepository.org is a Web-based flow cytometry data repository provided by the International Society for Advancement of Cytometry (ISAC). It supports storage, annotation, analysis, and sharing of flow cytometry datasets. A fundamental tenet of scientific research is that published results should be open to independent validation and refutation. With FlowRepository, researchers can annotate their datasets in compliance with the Minimum Information about a Flow Cytometry Experiment (MIFlowCyt) standard, thus greatly facilitating third-party interpretation of their data. In this unit, we will mainly focus on the deposition, sharing, and annotation of flow cytometry data.

  15. Imaging- and Flow Cytometry-based Analysis of Cell Position and the Cell Cycle in 3D Melanoma Spheroids

    Science.gov (United States)

    Beaumont, Kimberley A.; Anfosso, Andrea; Ahmed, Farzana

    2015-01-01

    Three-dimensional (3D) tumor spheroids are utilized in cancer research as a more accurate model of the in vivo tumor microenvironment, compared to traditional two-dimensional (2D) cell culture. The spheroid model is able to mimic the effects of cell-cell interaction, hypoxia and nutrient deprivation, and drug penetration. One characteristic of this model is the development of a necrotic core, surrounded by a ring of G1 arrested cells, with proliferating cells on the outer layers of the spheroid. Of interest in the cancer field is how different regions of the spheroid respond to drug therapies as well as genetic or environmental manipulation. We describe here the use of the fluorescence ubiquitination cell cycle indicator (FUCCI) system along with cytometry and image analysis using commercial software to characterize the cell cycle status of cells with respect to their position inside melanoma spheroids. These methods may be used to track changes in cell cycle status, gene/protein expression or cell viability in different sub-regions of tumor spheroids over time and under different conditions. PMID:26779761

  16. Functional characterization of neotropical snakes peripheral blood leukocytes subsets: Linking flow cytometry cell features, microscopy images and serum corticosterone levels.

    Science.gov (United States)

    de Carvalho, Marcelo Pires Nogueira; Queiroz-Hazarbassanov, Nicolle Gilda Teixeira; de Oliveira Massoco, Cristina; Sant'Anna, Sávio Stefanini; Lourenço, Mariana Mathias; Levin, Gabriel; Sogayar, Mari Cleide; Grego, Kathleen Fernandes; Catão-Dias, José Luiz

    2017-09-01

    Reptiles are the unique ectothermic amniotes, providing the key link between ectothermic anamniotes fish and amphibians, and endothermic birds and mammals; becoming an important group to study with the aim of providing significant knowledge into the evolutionary history of vertebrate immunity. Classification systems for reptiles' leukocytes have been described by their appearance rather than function, being still inconsistent. With the advent of modern techniques and the establishment of analytical protocols for snakes' blood by flow cytometry, we bring a qualitative and quantitative assessment of innate activities presented by snakes' peripheral blood leukocytes, thereby linking flow cytometric features with fluorescent and light microscopy images. Moreover, since corticosterone is an important immunomodulator in reptiles, hormone levels of all blood samples were measured. We provide novel and additional information which should contribute to better understanding of the development of the immune system of reptiles and vertebrates. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Effectiveness of pulse-shape criteria for the selection of dicentric chromosomes by slit-scan flow cytometry and sorting

    NARCIS (Netherlands)

    Rens, W.; van Oven, C. H.; Stap, J.; Aten, J. A.

    1993-01-01

    A method was developed to detect dicentric chromosomes by slit-scan flow cytometry. The two centromeres of dicentric chromosomes are represented by the two dips in the trimodal fluorescence profile. A trimodal profile can, however, also be generated by aggregates of chromosomes. We tested the

  18. Flow Cytometry Enables Multiplexed Measurements of Genetically Encoded Intramolecular FRET Sensors Suitable for Screening.

    Science.gov (United States)

    Doucette, Jaimee; Zhao, Ziyan; Geyer, Rory J; Barra, Melanie M; Balunas, Marcy J; Zweifach, Adam

    2016-07-01

    Genetically encoded sensors based on intramolecular FRET between CFP and YFP are used extensively in cell biology research. Flow cytometry has been shown to offer a means to measure CFP-YFP FRET; we suspected it would provide a unique way to conduct multiplexed measurements from cells expressing different FRET sensors, which is difficult to do with microscopy, and that this could be used for screening. We confirmed that flow cytometry accurately measures FRET signals using cells transiently transfected with an ERK activity reporter, comparing responses measured with imaging and cytometry. We created polyclonal long-term transfectant lines, each expressing a different intramolecular FRET sensor, and devised a way to bar-code four distinct populations of cells. We demonstrated the feasibility of multiplexed measurements and determined that robust multiplexed measurements can be conducted in plate format. To validate the suitability of the method for screening, we measured responses from a plate of bacterial extracts that in unrelated experiments we had determined contained the protein kinase C (PKC)-activating compound teleocidin A-1. The multiplexed assay correctly identifying the teleocidin A-1-containing well. We propose that multiplexed cytometric FRET measurements will be useful for analyzing cellular function and for screening compound collections. © 2016 Society for Laboratory Automation and Screening.

  19. Basic strategies for valid cytometry using image analysis

    NARCIS (Netherlands)

    Jonker, A.; Geerts, W. J.; Chieco, P.; Moorman, A. F.; Lamers, W. H.; van Noorden, C. J.

    1997-01-01

    The present review provides a starting point for setting up an image analysis system for quantitative densitometry and absorbance or fluorescence measurements in cell preparations, tissue sections or gels. Guidelines for instrumental settings that are essential for the valid application of image

  20. Visual selection and maintenance of the cell lines with high plant regeneration ability and low ploidy level in Dianthus acicularis by monitoring with flow cytometry analysis.

    Science.gov (United States)

    Shiba, Tomonori; Mii, Masahiro

    2005-12-01

    Efficient plant regeneration system from cell suspension cultures was established in D. acicularis (2n=90) by monitoring ploidy level and visual selection of the cultures. The ploidy level of the cell cultures closely related to the shoot regeneration ability. The cell lines comprising original ploidy levels (2C+4C cells corresponding to DNA contents of G1 and G2 cells of diploid plant, respectively) showed high regeneration ability, whereas those containing the cells with 8C or higher DNA C-values showed low or no regeneration ability. The highly regenerable cell lines thus selected consisted of compact cell clumps with yellowish color and relatively moderate growth, suggesting that it is possible to select visually the highly regenerable cell lines with the original ploidy level. All the regenerated plantlets from the highly regenerable cell cultures exhibited normal phenotypes and no variations in ploidy level were observed by flow cytometry (FCM) analysis.

  1. Highly multiparametric analysis by mass cytometry.

    Science.gov (United States)

    Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Nitz, Mark; Winnik, Mitchell A; Tanner, Scott

    2010-09-30

    This review paper describes a new technology, mass cytometry, that addresses applications typically run by flow cytometer analyzers, but extends the capability to highly multiparametric analysis. The detection technology is based on atomic mass spectrometry. It offers quantitation, specificity and dynamic range of mass spectrometry in a format that is familiar to flow cytometry practitioners. The mass cytometer does not require compensation, allowing the application of statistical techniques; this has been impossible given the constraints of fluorescence noise with traditional cytometry instruments. Instead of "colors" the mass cytometer "reads" the stable isotope tags attached to antibodies using metal-chelating labeling reagents. Because there are many available stable isotopes, and the mass spectrometer provides exquisite resolution between detection channels, many parameters can be measured as easily as one. For example, in a single tube the technique allows for the ready detection and characterization of the major cell subsets in blood or bone marrow. Here we describe mass cytometric immunophenotyping of human leukemia cell lines and leukemia patient samples, differential cell analysis of normal peripheral and umbilical cord blood; intracellular protein identification and metal-encoded bead arrays. Copyright © 2010 Elsevier B.V. All rights reserved.

  2. TEM Imaging of Mass-selected Polymer Molecules

    International Nuclear Information System (INIS)

    Nasibulin, Albert G.; Kauppinen, Esko I.; Thomson, Bruce A.; Fernandez de la Mora, J.

    2002-01-01

    Polyethylene glycol (PEG) molecules with masses below 1300 amu are electrosprayed (ES) from solution, mobility-selected at high resolution in a differential mobility analyzer (DMA), collected on a grid and imaged by transmission electron microscopy (ES-DMA-TEM). The DMA resolves individual n-mers, and selects only one out of the many present in the original sample. Ion identity is established from parallel mass spectra (ES-MS). The images reveal spherical particles 1.46 nm in diameter, in good agreement with the known ion mass and bulk density. The DMA-selection technique opens new paths for the study of very small particles

  3. Flow Cytometry Technician | Center for Cancer Research

    Science.gov (United States)

    PROGRAM DESCRIPTION The Basic Science Program (BSP) pursues independent, multidisciplinary research in basic and applied molecular biology, immunology, retrovirology, cancer biology, and human genetics. Research efforts and support are an integral part of the Center for Cancer Research (CCR) at the Frederick National Laboratory for Cancer Research (FNLCR). KEY ROLES/RESPONSIBILITIES The Flow Cytometry Core (Flow Core) of the Cancer and Inflammation Program (CIP) is a service core which supports the research efforts of the CCR by providing expertise in the field of flow cytometry (using analyzers and sorters) with the goal of gaining a more thorough understanding of the biology of cancer and cancer cells. The Flow Core provides service to 12-15 CIP laboratories and more than 22 non-CIP laboratories. Flow core staff provide technical advice on the experimental design of applications, which include immunological phenotyping, cell function assays, and cell cycle analysis. Work is performed per customer requirements, and no independent research is involved. The Flow Cytometry Technician will be responsible for: Monitor performance of and maintain high dimensional flow cytometer analyzers and cell sorters Operate high dimensional flow cytometer analyzers and cell sorters Monitoring lab supply levels and order lab supplies, perform various record keeping responsibilities Assist in the training of scientific end users on the use of flow cytometry in their research, as well as how to operate and troubleshoot the bench-top analyzer instruments Experience with sterile technique and tissue culture

  4. FuGEFlow: data model and markup language for flow cytometry

    Directory of Open Access Journals (Sweden)

    Manion Frank J

    2009-06-01

    Full Text Available Abstract Background Flow cytometry technology is widely used in both health care and research. The rapid expansion of flow cytometry applications has outpaced the development of data storage and analysis tools. Collaborative efforts being taken to eliminate this gap include building common vocabularies and ontologies, designing generic data models, and defining data exchange formats. The Minimum Information about a Flow Cytometry Experiment (MIFlowCyt standard was recently adopted by the International Society for Advancement of Cytometry. This standard guides researchers on the information that should be included in peer reviewed publications, but it is insufficient for data exchange and integration between computational systems. The Functional Genomics Experiment (FuGE formalizes common aspects of comprehensive and high throughput experiments across different biological technologies. We have extended FuGE object model to accommodate flow cytometry data and metadata. Methods We used the MagicDraw modelling tool to design a UML model (Flow-OM according to the FuGE extension guidelines and the AndroMDA toolkit to transform the model to a markup language (Flow-ML. We mapped each MIFlowCyt term to either an existing FuGE class or to a new FuGEFlow class. The development environment was validated by comparing the official FuGE XSD to the schema we generated from the FuGE object model using our configuration. After the Flow-OM model was completed, the final version of the Flow-ML was generated and validated against an example MIFlowCyt compliant experiment description. Results The extension of FuGE for flow cytometry has resulted in a generic FuGE-compliant data model (FuGEFlow, which accommodates and links together all information required by MIFlowCyt. The FuGEFlow model can be used to build software and databases using FuGE software toolkits to facilitate automated exchange and manipulation of potentially large flow cytometry experimental data sets

  5. FuGEFlow: data model and markup language for flow cytometry.

    Science.gov (United States)

    Qian, Yu; Tchuvatkina, Olga; Spidlen, Josef; Wilkinson, Peter; Gasparetto, Maura; Jones, Andrew R; Manion, Frank J; Scheuermann, Richard H; Sekaly, Rafick-Pierre; Brinkman, Ryan R

    2009-06-16

    Flow cytometry technology is widely used in both health care and research. The rapid expansion of flow cytometry applications has outpaced the development of data storage and analysis tools. Collaborative efforts being taken to eliminate this gap include building common vocabularies and ontologies, designing generic data models, and defining data exchange formats. The Minimum Information about a Flow Cytometry Experiment (MIFlowCyt) standard was recently adopted by the International Society for Advancement of Cytometry. This standard guides researchers on the information that should be included in peer reviewed publications, but it is insufficient for data exchange and integration between computational systems. The Functional Genomics Experiment (FuGE) formalizes common aspects of comprehensive and high throughput experiments across different biological technologies. We have extended FuGE object model to accommodate flow cytometry data and metadata. We used the MagicDraw modelling tool to design a UML model (Flow-OM) according to the FuGE extension guidelines and the AndroMDA toolkit to transform the model to a markup language (Flow-ML). We mapped each MIFlowCyt term to either an existing FuGE class or to a new FuGEFlow class. The development environment was validated by comparing the official FuGE XSD to the schema we generated from the FuGE object model using our configuration. After the Flow-OM model was completed, the final version of the Flow-ML was generated and validated against an example MIFlowCyt compliant experiment description. The extension of FuGE for flow cytometry has resulted in a generic FuGE-compliant data model (FuGEFlow), which accommodates and links together all information required by MIFlowCyt. The FuGEFlow model can be used to build software and databases using FuGE software toolkits to facilitate automated exchange and manipulation of potentially large flow cytometry experimental data sets. Additional project documentation, including

  6. Features Selection for Skin Micro-Image Symptomatic Recognition

    Institute of Scientific and Technical Information of China (English)

    HUYue-li; CAOJia-lin; ZHAOQian; FENGXu

    2004-01-01

    Automatic recognition of skin micro-image symptom is important in skin diagnosis and treatment. Feature selection is to improve the classification performance of skin micro-image symptom.This paper proposes a hybrid approach based on the support vector machine (SVM) technique and genetic algorithm (GA) to select an optimum feature subset from the feature group extracted from the skin micro-images. An adaptive GA is introduced for maintaining the convergence rate. With the proposed method, the average cross validation accuracy is increased from 88.25% using all features to 96.92% using only selected features provided by a classifier for classification of 5 classes of skin symptoms. The experimental results are satisfactory.

  7. Features Selection for Skin Micro-Image Symptomatic Recognition

    Institute of Scientific and Technical Information of China (English)

    HU Yue-li; CAO Jia-lin; ZHAO Qian; FENG Xu

    2004-01-01

    Automatic recognition of skin micro-image symptom is important in skin diagnosis and treatment. Feature selection is to improve the classification performance of skin micro-image symptom.This paper proposes a hybrid approach based on the support vector machine (SVM) technique and genetic algorithm (GA) to select an optimum feature subset from the feature group extracted from the skin micro-images. An adaptive GA is introduced for maintaining the convergence rate. With the proposed method, the average cross validation accuracy is increased from 88.25% using all features to 96.92 % using only selected features provided by a classifier for classification of 5 classes of skin symptoms. The experimental results are satisfactory.

  8. Hyperspectral small animal fluorescence imaging: spectral selection imaging

    Science.gov (United States)

    Leavesley, Silas; Jiang, Yanan; Patsekin, Valery; Hall, Heidi; Vizard, Douglas; Robinson, J. Paul

    2008-02-01

    Molecular imaging is a rapidly growing area of research, fueled by needs in pharmaceutical drug-development for methods for high-throughput screening, pre-clinical and clinical screening for visualizing tumor growth and drug targeting, and a growing number of applications in the molecular biology fields. Small animal fluorescence imaging employs fluorescent probes to target molecular events in vivo, with a large number of molecular targeting probes readily available. The ease at which new targeting compounds can be developed, the short acquisition times, and the low cost (compared to microCT, MRI, or PET) makes fluorescence imaging attractive. However, small animal fluorescence imaging suffers from high optical scattering, absorption, and autofluorescence. Much of these problems can be overcome through multispectral imaging techniques, which collect images at different fluorescence emission wavelengths, followed by analysis, classification, and spectral deconvolution methods to isolate signals from fluorescence emission. We present an alternative to the current method, using hyperspectral excitation scanning (spectral selection imaging), a technique that allows excitation at any wavelength in the visible and near-infrared wavelength range. In many cases, excitation imaging may be more effective at identifying specific fluorescence signals because of the higher complexity of the fluorophore excitation spectrum. Because the excitation is filtered and not the emission, the resolution limit and image shift imposed by acousto-optic tunable filters have no effect on imager performance. We will discuss design of the imager, optimizing the imager for use in small animal fluorescence imaging, and application of spectral analysis and classification methods for identifying specific fluorescence signals.

  9. Interfacing 3D magnetic twisting cytometry with confocal fluorescence microscopy to image force responses in living cells.

    Science.gov (United States)

    Zhang, Yuejin; Wei, Fuxiang; Poh, Yeh-Chuin; Jia, Qiong; Chen, Junjian; Chen, Junwei; Luo, Junyu; Yao, Wenting; Zhou, Wenwen; Huang, Wei; Yang, Fang; Zhang, Yao; Wang, Ning

    2017-07-01

    Cells and tissues can undergo a variety of biological and structural changes in response to mechanical forces. Only a few existing techniques are available for quantification of structural changes at high resolution in response to forces applied along different directions. 3D-magnetic twisting cytometry (3D-MTC) is a technique for applying local mechanical stresses to living cells. Here we describe a protocol for interfacing 3D-MTC with confocal fluorescence microscopy. In 3D-MTC, ferromagnetic beads are bound to the cell surface via surface receptors, followed by their magnetization in any desired direction. A magnetic twisting field in a different direction is then applied to generate rotational shear stresses in any desired direction. This protocol describes how to combine magnetic-field-induced mechanical stimulation with confocal fluorescence microscopy and provides an optional extension for super-resolution imaging using stimulated emission depletion (STED) nanoscopy. This technology allows for rapid real-time acquisition of a living cell's mechanical responses to forces via specific receptors and for quantifying structural and biochemical changes in the same cell using confocal fluorescence microscopy or STED. The integrated 3D-MTC-microscopy platform takes ∼20 d to construct, and the experimental procedures require ∼4 d when carried out by a life sciences graduate student.

  10. Image selection in static telepathology through the Internet.

    Science.gov (United States)

    Della Mea, V; Cataldi, P; Boi, S; Finato, N; Della Palma, P; Beltrami, C A

    1998-01-01

    A telepathology study was carried out to examine the differences occurring when the images were selected by an experienced pathologist, a junior pathologist and a first-year resident. One hundred and fifty-five consecutive frozen-section pathology cases were collected and sent for consultation to a remote experienced pathologist using multimedia email. Local diagnoses (as reported in the files of the Institute, not from the image selector) and remote diagnoses (based on the images) were compared with those performed on paraffin-embedded sections. Acquisition time and number of selected images were recorded for each case and used to compare the different behaviour of the three local pathologists. Of the 155 cases sent by telepathology, four were considered insufficient for a diagnosis by the remote pathologist and thus the diagnosis was postponed. In the remaining 151 cases, the overall diagnostic agreement between remote and definitive diagnosis was 96.7%. The results indicate that in the routine diagnostic work of a frozen-section service, an inexperienced pathologist can select images which are sufficiently informative for a remote diagnosis, in a sufficiently short time.

  11. Spaceflight Flow Cytometry: Design Challenges and Applications

    Science.gov (United States)

    Pappas, Dimitri; Kao, Shih-Hsin; Jeevarajan, Antony S.

    2004-01-01

    Future space exploration missions will require analytical technology capable of providing both autonomous medical care to the crew and investigative capabilities to researchers. While several promising candidate technologies exist for further development, flow cytometry is an attractive technology as it offers both crew health and a wide array of biochemistry and immunology assays. While flow cytometry has been widely used for cellular analysis in both clinical and research settings, the requirements for proper operation in spaceflight impose constraints on any instrument designs. The challenges of designing a spaceflight-ready flow cytometer are discussed, as well as some preliminary results using a prototype system.

  12. Flow cytometry measurements of human chromosome kinetochore labeling

    International Nuclear Information System (INIS)

    Fantes, J.A.; Green, D.K.; Malloy, P.; Sumner, A.T.

    1989-01-01

    A method for the preparation and measurement of immunofluorescent human chromosome centromeres in suspension is described using CREST antibodies, which bind to the centromeric region of chromosomes. Fluorescein isothiocyanate (FITC)-conjugated antihuman antibodies provide the fluorescent label. Labeled chromosomes are examined on microscope slides and by flow cytometry. In both cases a dye which binds to DNA is added to provide identification of the chromosome groups. Sera from different CREST patients vary in their ability to bind to chromosome arms in addition to the centromeric region. Flow cytometry and microfluorimetry measurements have shown that with a given CREST serum the differences in kinetochore fluorescence between chromosomes are only minor. Flow cytometry experiments to relate the number of dicentric chromosomes, induced by in vitro radiation of peripheral blood cells to the slightly increased number of chromosomes with above-average kinetochore fluorescence did not produce decisive radiation dosimetry results

  13. Revelation of Different Nanoparticle-Uptake Behavior in Two Standard Cell Lines NIH/3T3 and A549 by Flow Cytometry and Time-Lapse Imaging

    Directory of Open Access Journals (Sweden)

    André Jochums

    2017-07-01

    Full Text Available The uptake of nanomaterials into different cell types is a central pharmacological issue for the determination of nanotoxicity as well as for the development of drug delivery strategies. Most responses of the cells depend on their intracellular interactions with nanoparticles (NPs. Uptake behavior can be precisely investigated in vitro, with sensitive high throughput methods such as flow cytometry. In this study, we investigated two different standard cell lines, human lung carcinoma (A549 and mouse fibroblast (NIH/3T3 cells, regarding their uptake behavior of titanium dioxide NPs. Cells were incubated with different concentrations of TiO2 NPs and samples were taken at certain time points to compare the uptake kinetics of both cell lines. Samples were analyzed with the help of flow cytometry by studying changes in the side and forward scattering signal. To additionally enable a detection via fluorescence, NPs were labeled with the fluorescent dye fluorescein isothiocyanate (FITC and propidium iodide (PI. We found that NIH/3T3 cells take up the studied NPs more efficiently than A549 cells. These findings were supported by time-lapse microscopic imaging of the cells incubated with TiO2 NPs. Our results confirm that the uptake behavior of individual cell types has to be considered before interpreting any results of nanomaterial studies.

  14. Characterization of glycosylphosphatidylinositol biosynthesis defects by clinical features, flow cytometry, and automated image analysis

    DEFF Research Database (Denmark)

    Knaus, Alexej; Pantel, Jean Tori; Pendziwiat, Manuela

    2018-01-01

    , the increasing number of individuals with a GPIBD shows that hyperphosphatasia is a variable feature that is not ideal for a clinical classification. METHODS: We studied the discriminatory power of multiple GPI-linked substrates that were assessed by flow cytometry in blood cells and fibroblasts of 39 and 14...... those with PIGA mutations. Although the impairment of GPI-linked substrates is supposed to play the key role in the pathophysiology of GPIBDs, we could not observe gene-specific profiles for flow cytometric markers or a correlation between their cell surface levels and the severity of the phenotype...

  15. Single-Particle Discrimination of Retroviruses from Extracellular Vesicles by Nanoscale Flow Cytometry.

    Science.gov (United States)

    Tang, Vera A; Renner, Tyler M; Fritzsche, Anna K; Burger, Dylan; Langlois, Marc-André

    2017-12-19

    Retroviruses and small EVs overlap in size, buoyant densities, refractive indices and share many cell-derived surface markers making them virtually indistinguishable by standard biochemical methods. This poses a significant challenge when purifying retroviruses for downstream analyses or for phenotypic characterization studies of markers on individual virions given that EVs are a major contaminant of retroviral preparations. Nanoscale flow cytometry (NFC), also called flow virometry, is an adaptation of flow cytometry technology for the analysis of individual nanoparticles such as extracellular vesicles (EVs) and retroviruses. In this study we systematically optimized NFC parameters for the detection of retroviral particles in the range of 115-130 nm, including viral production, sample labeling, laser power and voltage settings. By using the retroviral envelope glycoprotein as a selection marker, and evaluating a number of fluorescent dyes and labeling methods, we demonstrate that it is possible to confidently distinguish retroviruses from small EVs by NFC. Our findings make it now possible to individually phenotype genetically modified retroviral particles that express a fluorescent envelope glycoprotein without removing EV contaminants from the sample.

  16. Discriminative Projection Selection Based Face Image Hashing

    Science.gov (United States)

    Karabat, Cagatay; Erdogan, Hakan

    Face image hashing is an emerging method used in biometric verification systems. In this paper, we propose a novel face image hashing method based on a new technique called discriminative projection selection. We apply the Fisher criterion for selecting the rows of a random projection matrix in a user-dependent fashion. Moreover, another contribution of this paper is to employ a bimodal Gaussian mixture model at the quantization step. Our simulation results on three different databases demonstrate that the proposed method has superior performance in comparison to previously proposed random projection based methods.

  17. Measurement of lipid accumulation in Chlorella vulgaris via flow cytometry and liquid-state ¹H NMR spectroscopy for development of an NMR-traceable flow cytometry protocol.

    Directory of Open Access Journals (Sweden)

    Michael S Bono

    Full Text Available In this study, we cultured Chlorella vulgaris cells with a range of lipid contents, induced via nitrogen starvation, and characterized them via flow cytometry, with BODIPY 505/515 as a fluorescent lipid label, and liquid-state 1H NMR spectroscopy. In doing so, we demonstrate the utility of calibrating flow cytometric measurements of algal lipid content using triacylglyceride (TAG, also known as triacylglycerol or triglyceride content per cell as measured via quantitative 1H NMR. Ensemble-averaged fluorescence of BODIPY-labeled cells was highly correlated with average TAG content per cell measured by bulk NMR, with a linear regression yielding a linear fit with r2 = 0.9974. This correlation compares favorably to previous calibrations of flow cytometry protocols to lipid content measured via extraction, and calibration by NMR avoids the time and complexity that is generally required for lipid quantitation via extraction. Flow cytometry calibrated to a direct measurement of TAG content can be used to investigate the distribution of lipid contents for cells within a culture. Our flow cytometry measurements showed that Chlorella vulgaris cells subjected to nitrogen limitation exhibited higher mean lipid content but a wider distribution of lipid content that overlapped the relatively narrow distribution of lipid content for replete cells, suggesting that nitrogen limitation induces lipid accumulation in only a subset of cells. Calibration of flow cytometry protocols using direct in situ measurement of TAG content via NMR will facilitate rapid development of more precise flow cytometry protocols, enabling investigation of algal lipid accumulation for development of more productive algal biofuel feedstocks and cultivation protocols.

  18. Measurement of Lipid Accumulation in Chlorella vulgaris via Flow Cytometry and Liquid-State ¹H NMR Spectroscopy for Development of an NMR-Traceable Flow Cytometry Protocol

    Science.gov (United States)

    Bono Jr., Michael S.; Garcia, Ravi D.; Sri-Jayantha, Dylan V.; Ahner, Beth A.; Kirby, Brian J.

    2015-01-01

    In this study, we cultured Chlorella vulgaris cells with a range of lipid contents, induced via nitrogen starvation, and characterized them via flow cytometry, with BODIPY 505/515 as a fluorescent lipid label, and liquid-state 1H NMR spectroscopy. In doing so, we demonstrate the utility of calibrating flow cytometric measurements of algal lipid content using triacylglyceride (TAG, also known as triacylglycerol or triglyceride) content per cell as measured via quantitative 1H NMR. Ensemble-averaged fluorescence of BODIPY-labeled cells was highly correlated with average TAG content per cell measured by bulk NMR, with a linear regression yielding a linear fit with r 2 = 0.9974. This correlation compares favorably to previous calibrations of flow cytometry protocols to lipid content measured via extraction, and calibration by NMR avoids the time and complexity that is generally required for lipid quantitation via extraction. Flow cytometry calibrated to a direct measurement of TAG content can be used to investigate the distribution of lipid contents for cells within a culture. Our flow cytometry measurements showed that Chlorella vulgaris cells subjected to nitrogen limitation exhibited higher mean lipid content but a wider distribution of lipid content that overlapped the relatively narrow distribution of lipid content for replete cells, suggesting that nitrogen limitation induces lipid accumulation in only a subset of cells. Calibration of flow cytometry protocols using direct in situ measurement of TAG content via NMR will facilitate rapid development of more precise flow cytometry protocols, enabling investigation of algal lipid accumulation for development of more productive algal biofuel feedstocks and cultivation protocols. PMID:26267664

  19. Measurement of lipid accumulation in Chlorella vulgaris via flow cytometry and liquid-state ¹H NMR spectroscopy for development of an NMR-traceable flow cytometry protocol.

    Science.gov (United States)

    Bono, Michael S; Garcia, Ravi D; Sri-Jayantha, Dylan V; Ahner, Beth A; Kirby, Brian J

    2015-01-01

    In this study, we cultured Chlorella vulgaris cells with a range of lipid contents, induced via nitrogen starvation, and characterized them via flow cytometry, with BODIPY 505/515 as a fluorescent lipid label, and liquid-state 1H NMR spectroscopy. In doing so, we demonstrate the utility of calibrating flow cytometric measurements of algal lipid content using triacylglyceride (TAG, also known as triacylglycerol or triglyceride) content per cell as measured via quantitative 1H NMR. Ensemble-averaged fluorescence of BODIPY-labeled cells was highly correlated with average TAG content per cell measured by bulk NMR, with a linear regression yielding a linear fit with r2 = 0.9974. This correlation compares favorably to previous calibrations of flow cytometry protocols to lipid content measured via extraction, and calibration by NMR avoids the time and complexity that is generally required for lipid quantitation via extraction. Flow cytometry calibrated to a direct measurement of TAG content can be used to investigate the distribution of lipid contents for cells within a culture. Our flow cytometry measurements showed that Chlorella vulgaris cells subjected to nitrogen limitation exhibited higher mean lipid content but a wider distribution of lipid content that overlapped the relatively narrow distribution of lipid content for replete cells, suggesting that nitrogen limitation induces lipid accumulation in only a subset of cells. Calibration of flow cytometry protocols using direct in situ measurement of TAG content via NMR will facilitate rapid development of more precise flow cytometry protocols, enabling investigation of algal lipid accumulation for development of more productive algal biofuel feedstocks and cultivation protocols.

  20. Content-based image retrieval: Color-selection exploited

    NARCIS (Netherlands)

    Broek, E.L. van den; Vuurpijl, L.G.; Kisters, P. M. F.; Schmid, J.C.M. von; Moens, M.F.; Busser, R. de; Hiemstra, D.; Kraaij, W.

    2002-01-01

    This research presents a new color selection interface that facilitates query-by-color in Content-Based Image Retrieval (CBIR). Existing CBIR color selection interfaces, are being judged as non-intuitive and difficult to use. Our interface copes with these problems of usability. It is based on 11

  1. Content-Based Image Retrieval: Color-selection exploited

    NARCIS (Netherlands)

    Moens, Marie-Francine; van den Broek, Egon; Vuurpijl, L.G.; de Brusser, Rik; Kisters, P.M.F.; Hiemstra, Djoerd; Kraaij, Wessel; von Schmid, J.C.M.

    2002-01-01

    This research presents a new color selection interface that facilitates query-by-color in Content-Based Image Retrieval (CBIR). Existing CBIR color selection interfaces, are being judged as non-intuitive and difficult to use. Our interface copes with these problems of usability. It is based on 11

  2. Studying apoptotic cell death by flow cytometry

    International Nuclear Information System (INIS)

    Ormerod, Michael G.

    1998-01-01

    Full text: Programmed cell death (PCD) is of fundamental importance in the normal development of an animal and also in tumour biology and radiation biology. During PCD a sequence of changes occurs in cells giving rise to an apoptotic cascade of events. The main elements of this cascade are rapidly being elucidated. Flow cytometry has been used to follow many of these changes. It also has been used to quantify the number of apoptotic cells in a culture and, more recently, in clinical samples. In this review, the properties of apoptotic cells and the main feature of apoptotic cascade will be described. How flow cytometry can be used to follow changes during the apoptotic cascade will be discussed

  3. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry

    Science.gov (United States)

    Majeed, Muhammed; Majeed, Shaheen; Nagabhushanam, Kalyanam; Punnapuzha, Ardra; Philip, Sheena

    2018-01-01

    Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC) retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore®) is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM) was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856. PMID:29474436

  4. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry.

    Directory of Open Access Journals (Sweden)

    Muhammed Majeed

    Full Text Available Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore® is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856.

  5. Rapid assessment of viable but non-culturable Bacillus coagulans MTCC 5856 in commercial formulations using Flow cytometry.

    Science.gov (United States)

    Majeed, Muhammed; Majeed, Shaheen; Nagabhushanam, Kalyanam; Punnapuzha, Ardra; Philip, Sheena; Mundkur, Lakshmi

    2018-01-01

    Accurate enumeration of bacterial count in probiotic formulation is imperative to ensure that the product adheres to regulatory standards and citation in consumer product label. Standard methods like plate count, can enumerate only replicating bacterial population under selected culture conditions. Viable but non culturable bacteria (VBNC) retain characteristics of living cells and can regain cultivability by a process known as resuscitation. This is a protective mechanism adapted by bacteria to evade stressful environmental conditions. B. coagulans MTCC 5856(LactoSpore®) is a probiotic endospore which can survive for decades in hostile environments without dividing. In the present study, we explored the use of flow cytometry to enumerate the viable count of B. coagulans MTCC 5856 under acidic and alkaline conditions, high temperature and in commercial formulations like compressed tablets and capsules. Flow cytometry (FCM) was comparable to plate count method when the spores were counted at physiological conditions. We show that VBNC state is induced in B. coagulans MTCC 5856by high temperature and acidic pH. The cells get resuscitated under physiological conditions and FCM was sensitive to detect the VBNC spores. Flow cytometry showed excellent ability to assess the viable spore count in commercial probiotic formulations of B. coagulans MTCC 5856. The results establish Flow cytometry as a reliable method to count viable bacteria in commercial probiotic preparations. Sporulation as well as existence as VBNC could contribute to the extreme stability of B. coagulans MTCC 5856.

  6. Technical advances in flow cytometry-based diagnosis and monitoring of paroxysmal nocturnal hemoglobinuria

    Science.gov (United States)

    Correia, Rodolfo Patussi; Bento, Laiz Cameirão; Bortolucci, Ana Carolina Apelle; Alexandre, Anderson Marega; Vaz, Andressa da Costa; Schimidell, Daniela; Pedro, Eduardo de Carvalho; Perin, Fabricio Simões; Nozawa, Sonia Tsukasa; Mendes, Cláudio Ernesto Albers; Barroso, Rodrigo de Souza; Bacal, Nydia Strachman

    2016-01-01

    ABSTRACT Objective: To discuss the implementation of technical advances in laboratory diagnosis and monitoring of paroxysmal nocturnal hemoglobinuria for validation of high-sensitivity flow cytometry protocols. Methods: A retrospective study based on analysis of laboratory data from 745 patient samples submitted to flow cytometry for diagnosis and/or monitoring of paroxysmal nocturnal hemoglobinuria. Results: Implementation of technical advances reduced test costs and improved flow cytometry resolution for paroxysmal nocturnal hemoglobinuria clone detection. Conclusion: High-sensitivity flow cytometry allowed more sensitive determination of paroxysmal nocturnal hemoglobinuria clone type and size, particularly in samples with small clones. PMID:27759825

  7. A Fully Automated High-Throughput Flow Cytometry Screening System Enabling Phenotypic Drug Discovery.

    Science.gov (United States)

    Joslin, John; Gilligan, James; Anderson, Paul; Garcia, Catherine; Sharif, Orzala; Hampton, Janice; Cohen, Steven; King, Miranda; Zhou, Bin; Jiang, Shumei; Trussell, Christopher; Dunn, Robert; Fathman, John W; Snead, Jennifer L; Boitano, Anthony E; Nguyen, Tommy; Conner, Michael; Cooke, Mike; Harris, Jennifer; Ainscow, Ed; Zhou, Yingyao; Shaw, Chris; Sipes, Dan; Mainquist, James; Lesley, Scott

    2018-05-01

    The goal of high-throughput screening is to enable screening of compound libraries in an automated manner to identify quality starting points for optimization. This often involves screening a large diversity of compounds in an assay that preserves a connection to the disease pathology. Phenotypic screening is a powerful tool for drug identification, in that assays can be run without prior understanding of the target and with primary cells that closely mimic the therapeutic setting. Advanced automation and high-content imaging have enabled many complex assays, but these are still relatively slow and low throughput. To address this limitation, we have developed an automated workflow that is dedicated to processing complex phenotypic assays for flow cytometry. The system can achieve a throughput of 50,000 wells per day, resulting in a fully automated platform that enables robust phenotypic drug discovery. Over the past 5 years, this screening system has been used for a variety of drug discovery programs, across many disease areas, with many molecules advancing quickly into preclinical development and into the clinic. This report will highlight a diversity of approaches that automated flow cytometry has enabled for phenotypic drug discovery.

  8. Optical encryption with selective computational ghost imaging

    International Nuclear Information System (INIS)

    Zafari, Mohammad; Kheradmand, Reza; Ahmadi-Kandjani, Sohrab

    2014-01-01

    Selective computational ghost imaging (SCGI) is a technique which enables the reconstruction of an N-pixel image from N measurements or less. In this paper we propose an optical encryption method based on SCGI and experimentally demonstrate that this method has much higher security under eavesdropping and unauthorized accesses compared with previous reported methods. (paper)

  9. Cytobank: providing an analytics platform for community cytometry data analysis and collaboration.

    Science.gov (United States)

    Chen, Tiffany J; Kotecha, Nikesh

    2014-01-01

    Cytometry is used extensively in clinical and laboratory settings to diagnose and track cell subsets in blood and tissue. High-throughput, single-cell approaches leveraging cytometry are developed and applied in the computational and systems biology communities by researchers, who seek to improve the diagnosis of human diseases, map the structures of cell signaling networks, and identify new cell types. Data analysis and management present a bottleneck in the flow of knowledge from bench to clinic. Multi-parameter flow and mass cytometry enable identification of signaling profiles of patient cell samples. Currently, this process is manual, requiring hours of work to summarize multi-dimensional data and translate these data for input into other analysis programs. In addition, the increase in the number and size of collaborative cytometry studies as well as the computational complexity of analytical tools require the ability to assemble sufficient and appropriately configured computing capacity on demand. There is a critical need for platforms that can be used by both clinical and basic researchers who routinely rely on cytometry. Recent advances provide a unique opportunity to facilitate collaboration and analysis and management of cytometry data. Specifically, advances in cloud computing and virtualization are enabling efficient use of large computing resources for analysis and backup. An example is Cytobank, a platform that allows researchers to annotate, analyze, and share results along with the underlying single-cell data.

  10. Misty Mountain clustering: application to fast unsupervised flow cytometry gating

    Directory of Open Access Journals (Sweden)

    Sealfon Stuart C

    2010-10-01

    Full Text Available Abstract Background There are many important clustering questions in computational biology for which no satisfactory method exists. Automated clustering algorithms, when applied to large, multidimensional datasets, such as flow cytometry data, prove unsatisfactory in terms of speed, problems with local minima or cluster shape bias. Model-based approaches are restricted by the assumptions of the fitting functions. Furthermore, model based clustering requires serial clustering for all cluster numbers within a user defined interval. The final cluster number is then selected by various criteria. These supervised serial clustering methods are time consuming and frequently different criteria result in different optimal cluster numbers. Various unsupervised heuristic approaches that have been developed such as affinity propagation are too expensive to be applied to datasets on the order of 106 points that are often generated by high throughput experiments. Results To circumvent these limitations, we developed a new, unsupervised density contour clustering algorithm, called Misty Mountain, that is based on percolation theory and that efficiently analyzes large data sets. The approach can be envisioned as a progressive top-down removal of clouds covering a data histogram relief map to identify clusters by the appearance of statistically distinct peaks and ridges. This is a parallel clustering method that finds every cluster after analyzing only once the cross sections of the histogram. The overall run time for the composite steps of the algorithm increases linearly by the number of data points. The clustering of 106 data points in 2D data space takes place within about 15 seconds on a standard laptop PC. Comparison of the performance of this algorithm with other state of the art automated flow cytometry gating methods indicate that Misty Mountain provides substantial improvements in both run time and in the accuracy of cluster assignment. Conclusions

  11. Design Guidelines for a Content-Based Image Retrieval Color-Selection Interface

    NARCIS (Netherlands)

    Eggen, Berry; van den Broek, Egon; van der Veer, Gerrit C.; Kisters, Peter M.F.; Willems, Rob; Vuurpijl, Louis G.

    2004-01-01

    In Content-Based Image Retrieval (CBIR) two query-methods exist: query-by-example and query-by-memory. The user either selects an example image or selects image features retrieved from memory (such as color, texture, spatial attributes, and shape) to define his query. Hitherto, research on CBIR

  12. Clinical cytometry and progress in HLA antibody detection.

    Science.gov (United States)

    Bray, Robert A; Tarsitani, Christine; Gebel, Howard M; Lee, Jar-How

    2011-01-01

    For most solid organ and selected stem cell transplants, antibodies against mismatched HLA antigens can lead to early and late graft failure. In recognition of the clinical significance of these antibodies, HLA antibody identification is one of the most critical functions of histocompatibility laboratories. Early methods employed cumbersome and insensitive complement-dependent cytotoxicity assays with a visual read-out. A little over 20 years ago flow cytometry entered the realm of antibody detection with the introduction of the flow cytometric crossmatch. Cytometry's increased sensitivity and objectivity quickly earned it popularity as a preferred crossmatch method especially for sensitized recipients. Although a sensitive method, the flow crossmatch was criticized as being "too sensitive" as false positive reactions were a know drawback. In part, the shortcomings of the flow crossmatch were due to the lack of corresponding sensitive and specific HLA antibody screening assays. However, in the mid 1990s, solid phase assays, capable of utilizing standard flow cytometers, were developed. These assays used microparticles coated with purified HLA molecules. Hence, the era of solid-phase, microparticle technology for HLA antibody detection was born permitting the sensitive and specific detection of HLA antibody. It was now possible to provide better correlation between HLA antibody detection and the flow cytometric crossmatch. This flow-based technology was soon followed by adaptation to the Luminex platform permitting a mutltiplexed approach for the identification and characterization of HLA antibodies. It is hoped that these technologies will ultimately lead to the identification of parameters that best correlate with and/or predict transplant outcomes. Copyright © 2011 Elsevier Inc. All rights reserved.

  13. Robust tie points selection for InSAR image coregistration

    Science.gov (United States)

    Skanderi, Takieddine; Chabira, Boulerbah; Afifa, Belkacem; Belhadj Aissa, Aichouche

    2013-10-01

    Image coregistration is an important step in SAR interferometry which is a well known method for DEM generation and surface displacement monitoring. A practical and widely used automatic coregistration algorithm is based on selecting a number of tie points in the master image and looking for the correspondence of each point in the slave image using correlation technique. The characteristics of these points, their number and their distribution have a great impact on the reliability of the estimated transformation. In this work, we present a method for automatic selection of suitable tie points that are well distributed over the common area without decreasing the desired tie points' number. First we select candidate points using Harris operator. Then from these points we select tie points depending on their cornerness measure (the highest first). Once a tie point is selected, its correspondence is searched for in the slave image, if the similarity measure maximum is less than a given threshold or it is at the border of the search window, this point is discarded and we proceed to the next Harris point, else, the cornerness of the remaining candidates Harris points are multiplied by a spatially radially increasing function centered at the selected point to disadvantage the points in a neighborhood of a radius determined from the size of the common area and the desired number of points. This is repeated until the desired number of points is selected. Results of an ERS1/2 tandem pair are presented and discussed.

  14. The effect of an optimized imaging flow cytometry analysis template on sample throughput in the reduced culture cytokinesis-block micronucleus assay

    International Nuclear Information System (INIS)

    Rodrigues, M.A.; Beaton-Green, L.A.; Wilkins, R.C.; Probst, C.E.

    2016-01-01

    In cases of overexposure to ionizing radiation, the cytokinesis-block micronucleus (CBMN) assay can be performed in order to estimate the dose of radiation to an exposed individual. However, in the event of a large-scale radiation accident with many potentially exposed casualties, the assay must be able to generate accurate dose estimates to within ±0.5 Gy as quickly as possible. The assay has been adapted to, validated and optimized on the ImageStream"X imaging flow cyto-meter. The ease of running this automated version of the CBMN assay allowed investigation into the accuracy of dose estimates after reducing the volume of whole blood cultured to 200 μl and reducing the culture time to 48 h. The data analysis template used to identify binucleated lymphocyte cells (BNCs) and micronuclei (MN) has since been optimized to improve the sensitivity and specificity of BNC and MN detection. This paper presents a re-analysis of existing data using this optimized analysis template to demonstrate that dose estimations from blinded samples can be obtained to the same level of accuracy in a shorter data collection time. Here, we show that dose estimates from blinded samples were obtained to within ±0.5 Gy of the delivered dose when data collection time was reduced by 30 min at standard culture conditions and by 15 min at reduced culture conditions. Reducing data collection time while retaining the same level of accuracy in our imaging flow cytometry-based version of the CBMN assay results in higher throughput and further increases the relevancy of the CBMN assay as a radiation bio-dosimeter. (authors)

  15. Flow cytometry, fluorescent probes, and flashing bacteria

    NARCIS (Netherlands)

    Bunthof, C.J.

    2002-01-01


    Key words: fluorescent probes, flow cytometry, CSLM, viability, survival, microbial physiology, lactic acid bacteria, Lactococcus lactis , Lactobacillus plantarum , cheese, milk,

  16. Design and Application of Sensors for Chemical Cytometry.

    Science.gov (United States)

    Vickerman, Brianna M; Anttila, Matthew M; Petersen, Brae V; Allbritton, Nancy L; Lawrence, David S

    2018-02-08

    The bulk cell population response to a stimulus, be it a growth factor or a cytotoxic agent, neglects the cell-to-cell variability that can serve as a friend or as a foe in human biology. Biochemical variations among closely related cells furnish the basis for the adaptability of the immune system but also act as the root cause of resistance to chemotherapy by tumors. Consequently, the ability to probe for the presence of key biochemical variables at the single-cell level is now recognized to be of significant biological and biomedical impact. Chemical cytometry has emerged as an ultrasensitive single-cell platform with the flexibility to measure an array of cellular components, ranging from metabolite concentrations to enzyme activities. We briefly review the various chemical cytometry strategies, including recent advances in reporter design, probe and metabolite separation, and detection instrumentation. We also describe strategies for improving intracellular delivery, biochemical specificity, metabolic stability, and detection sensitivity of probes. Recent applications of these strategies to small molecules, lipids, proteins, and other analytes are discussed. Finally, we assess the current scope and limitations of chemical cytometry and discuss areas for future development to meet the needs of single-cell research.

  17. Subnuclear foci quantification using high-throughput 3D image cytometry

    Science.gov (United States)

    Wadduwage, Dushan N.; Parrish, Marcus; Choi, Heejin; Engelward, Bevin P.; Matsudaira, Paul; So, Peter T. C.

    2015-07-01

    Ionising radiation causes various types of DNA damages including double strand breaks (DSBs). DSBs are often recognized by DNA repair protein ATM which forms gamma-H2AX foci at the site of the DSBs that can be visualized using immunohistochemistry. However most of such experiments are of low throughput in terms of imaging and image analysis techniques. Most of the studies still use manual counting or classification. Hence they are limited to counting a low number of foci per cell (5 foci per nucleus) as the quantification process is extremely labour intensive. Therefore we have developed a high throughput instrumentation and computational pipeline specialized for gamma-H2AX foci quantification. A population of cells with highly clustered foci inside nuclei were imaged, in 3D with submicron resolution, using an in-house developed high throughput image cytometer. Imaging speeds as high as 800 cells/second in 3D were achieved by using HiLo wide-field depth resolved imaging and a remote z-scanning technique. Then the number of foci per cell nucleus were quantified using a 3D extended maxima transform based algorithm. Our results suggests that while most of the other 2D imaging and manual quantification studies can count only up to about 5 foci per nucleus our method is capable of counting more than 100. Moreover we show that 3D analysis is significantly superior compared to the 2D techniques.

  18. Selections from 2017: Image Processing with AstroImageJ

    Science.gov (United States)

    Kohler, Susanna

    2017-12-01

    Editors note:In these last two weeks of 2017, well be looking at a few selections that we havent yet discussed on AAS Nova from among the most-downloaded paperspublished in AAS journals this year. The usual posting schedule will resume in January.AstroImageJ: Image Processing and Photometric Extraction for Ultra-Precise Astronomical Light CurvesPublished January2017The AIJ image display. A wide range of astronomy specific image display options and image analysis tools are available from the menus, quick access icons, and interactive histogram. [Collins et al. 2017]Main takeaway:AstroImageJ is a new integrated software package presented in a publication led byKaren Collins(Vanderbilt University,Fisk University, andUniversity of Louisville). Itenables new users even at the level of undergraduate student, high school student, or amateur astronomer to quickly start processing, modeling, and plotting astronomical image data.Why its interesting:Science doesnt just happen the momenta telescope captures a picture of a distantobject. Instead, astronomical images must firstbe carefully processed to clean up thedata, and this data must then be systematically analyzed to learn about the objects within it. AstroImageJ as a GUI-driven, easily installed, public-domain tool is a uniquelyaccessible tool for thisprocessing and analysis, allowing even non-specialist users to explore and visualizeastronomical data.Some features ofAstroImageJ:(as reported by Astrobites)Image calibration:generate master flat, dark, and bias framesImage arithmetic:combineimages viasubtraction, addition, division, multiplication, etc.Stack editing:easily perform operations on a series of imagesImage stabilization and image alignment featuresPrecise coordinate converters:calculate Heliocentric and Barycentric Julian DatesWCS coordinates:determine precisely where atelescope was pointed for an image by PlateSolving using Astronomy.netMacro and plugin support:write your own macrosMulti-aperture photometry

  19. Some selected quantitative methods of thermal image analysis in Matlab.

    Science.gov (United States)

    Koprowski, Robert

    2016-05-01

    The paper presents a new algorithm based on some selected automatic quantitative methods for analysing thermal images. It shows the practical implementation of these image analysis methods in Matlab. It enables to perform fully automated and reproducible measurements of selected parameters in thermal images. The paper also shows two examples of the use of the proposed image analysis methods for the area of ​​the skin of a human foot and face. The full source code of the developed application is also provided as an attachment. The main window of the program during dynamic analysis of the foot thermal image. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. State-selective imaging of cold atoms

    NARCIS (Netherlands)

    Sheludko, D.V.; Bell, S.C.; Anderson, R.; Hofmann, C.S.; Vredenbregt, E.J.D.; Scholten, R.E.

    2008-01-01

    Atomic coherence phenomena are usually investigated using single beam techniques without spatial resolution. Here we demonstrate state-selective imaging of cold 85Rb atoms in a three-level ladder system, where the atomic refractive index is sensitive to the quantum coherence state of the atoms. We

  1. Organizing the Cellular and Molecular Heterogeneity in High-Grade Serous Ovarian Cancer by Mass Cytometry

    Science.gov (United States)

    2014-10-01

    Bendall SC, Sung P, Nolan GP, Arvin AM. Single-cell mass cytometry analysis of human tonsil T cell remodeling by varicella zoster virus. Cell Rep...Perspectives on Flow Cytometry 2013, September 20, 2013, Mass Cytometry and Cell Cycle, Mexico City, Mexico (by Web Conference) Nolan: Nuclear

  2. A dual-color flow cytometry protocol for the simultaneous detection of Vibrio parahaemolyticus and Salmonella typhimurium using aptamer conjugated quantum dots as labels

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Nuo; Wu, Shijia [State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi 214122 (China); Yu, Ye [Zhangjiagang Entry-Exit Inspection and Quarantine Bureau, Zhangjiangang 215600 (China); Ma, Xiaoyuan; Xia, Yu; Chen, Xiujuan; Huang, Yukun [State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi 214122 (China); Wang, Zhouping, E-mail: wangzp@jiangnan.edu.cn [State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi 214122 (China)

    2013-12-04

    Graphical abstract: -- Highlights: •Two bacteria were simultaneously detected using QD-apt as labels by flow cytometry. •QD-apt were used for recognition and fluorescence detection of two bacteria. •The method was applied successfully for bacteria detection in real samples. -- Abstract: A sensitive, specific method for the collection and detection of pathogenic bacteria was demonstrated using quantum dots (QDs) as a fluorescence marker coupled with aptamers as the molecular recognition element by flow cytometry. The aptamer sequences were selected using a bacterium-based SELEX strategy in our laboratory for Vibrio parahaemolyticus and Salmonella typhimurium that, when applied in this method, allows for the specific recognition of the bacteria from complex mixtures including shrimp samples. Aptamer-modified QDs (QD-apt) were employed to selectively capture and simultaneously detect the target bacteria with high sensitivity using the fluorescence of the labeled QDs. The signal intensity is amplified due to the high photostability of QDs nanoparticles, resulting in improved sensitivity over methods using individual dye-labeled probes. This proposed method is promising for the sensitive detection of other pathogenic bacteria in food stuff if suitable aptamers are chosen. The method may also provide another potential platform for the application of aptamer-conjugated QDs in flow cytometry.

  3. A dual-color flow cytometry protocol for the simultaneous detection of Vibrio parahaemolyticus and Salmonella typhimurium using aptamer conjugated quantum dots as labels

    International Nuclear Information System (INIS)

    Duan, Nuo; Wu, Shijia; Yu, Ye; Ma, Xiaoyuan; Xia, Yu; Chen, Xiujuan; Huang, Yukun; Wang, Zhouping

    2013-01-01

    Graphical abstract: -- Highlights: •Two bacteria were simultaneously detected using QD-apt as labels by flow cytometry. •QD-apt were used for recognition and fluorescence detection of two bacteria. •The method was applied successfully for bacteria detection in real samples. -- Abstract: A sensitive, specific method for the collection and detection of pathogenic bacteria was demonstrated using quantum dots (QDs) as a fluorescence marker coupled with aptamers as the molecular recognition element by flow cytometry. The aptamer sequences were selected using a bacterium-based SELEX strategy in our laboratory for Vibrio parahaemolyticus and Salmonella typhimurium that, when applied in this method, allows for the specific recognition of the bacteria from complex mixtures including shrimp samples. Aptamer-modified QDs (QD-apt) were employed to selectively capture and simultaneously detect the target bacteria with high sensitivity using the fluorescence of the labeled QDs. The signal intensity is amplified due to the high photostability of QDs nanoparticles, resulting in improved sensitivity over methods using individual dye-labeled probes. This proposed method is promising for the sensitive detection of other pathogenic bacteria in food stuff if suitable aptamers are chosen. The method may also provide another potential platform for the application of aptamer-conjugated QDs in flow cytometry

  4. Flow cytometry approach for studying the interaction between ...

    African Journals Online (AJOL)

    Flow cytometry approach for studying the interaction between Bacillus mojavensis and Alternaria alternata. Asma Milet, Noreddine Kacem Chaouche, Laid Dehimat, Asma Ait Kaki, Mounira Kara Ali, Philippe Thonart ...

  5. Automatic anatomically selective image enhancement in digital chest radiography

    International Nuclear Information System (INIS)

    Sezan, M.I.; Minerbo, G.N.; Schaetzing, R.

    1989-01-01

    The authors develop a technique for automatic anatomically selective enhancement of digital chest radiographs. Anatomically selective enhancement is motivated by the desire to simultaneously meet the different enhancement requirements of the lung field and the mediastinum. A recent peak detection algorithm and a set of rules are applied to the image histogram to determine automatically a gray-level threshold between the lung field and mediastinum. The gray-level threshold facilitates anatomically selective gray-scale modification and/or unsharp masking. Further, in an attempt to suppress possible white-band or black-band artifacts due to unsharp masking at sharp edges, local-contrast adaptivity is incorporated into anatomically selective unsharp masking by designing an anatomy-sensitive emphasis parameter which varies asymmetrically with positive and negative values of the local image contrast

  6. Imaging B lymphocytes in autoimmune inflammatory diseases

    International Nuclear Information System (INIS)

    Iodice, V.; Lauri, C.; Capriotti, G.; Lagana', B.; Germano, V.; D’Amelio, R.; Picchianti Diamanti, A.

    2014-01-01

    B cells arise from stem cells precursor and develop through a tightly regulated and selective process that lead to the generation of different B cell populations such as transitional, mature, memory and plasma cells. These B cell subsets can be identified using flow cytometry by the expression of specific surface antigens. The growing knowledge of the pivotal role played by B cells in the development and progression of autoimmune diseases combined with the advances in monoclonal antibody technology, led in the last years to the generation of different biological agents targeting B cells. In this context, nuclear medicine can offer the possibility to use a panel of biologic radiopharmaceuticals for molecular imaging of inflammatory diseases. Radiopharmaceuticals bind to their targets with high affinity and specificity and have an excellent imaging diagnostic potential for the evaluation of disease activity, selection and monitoring of immune therapies. Several molecules have been radiolabelled for the imaging of T lymphocytes whereas, by now, the anti CD20 rituximab is the only biological therapy targeting B cells that demonstrated to be efficiently radiolabelled and used to detect inflammation in autoimmune patients

  7. Online Feature Selection for Classifying Emphysema in HRCT Images

    Directory of Open Access Journals (Sweden)

    M. Prasad

    2008-06-01

    Full Text Available Feature subset selection, applied as a pre- processing step to machine learning, is valuable in dimensionality reduction, eliminating irrelevant data and improving classifier performance. In the classic formulation of the feature selection problem, it is assumed that all the features are available at the beginning. However, in many real world problems, there are scenarios where not all features are present initially and must be integrated as they become available. In such scenarios, online feature selection provides an efficient way to sort through a large space of features. It is in this context that we introduce online feature selection for the classification of emphysema, a smoking related disease that appears as low attenuation regions in High Resolution Computer Tomography (HRCT images. The technique was successfully evaluated on 61 HRCT scans and compared with different online feature selection approaches, including hill climbing, best first search, grafting, and correlation-based feature selection. The results were also compared against ldensity maskr, a standard approach used for emphysema detection in medical image analysis.

  8. Immune response to mycobacterial infection: lessons from flow cytometry.

    Science.gov (United States)

    Rovina, Nikoletta; Panagiotou, Marios; Pontikis, Konstantinos; Kyriakopoulou, Magdalini; Koulouris, Nikolaos G; Koutsoukou, Antonia

    2013-01-01

    Detecting and treating active and latent tuberculosis are pivotal elements for effective infection control; yet, due to their significant inherent limitations, the diagnostic means for these two stages of tuberculosis (TB) to date remain suboptimal. This paper reviews the current diagnostic tools for mycobacterial infection and focuses on the application of flow cytometry as a promising method for rapid and reliable diagnosis of mycobacterial infection as well as discrimination between active and latent TB: it summarizes diagnostic biomarkers distinguishing the two states of infection and also features of the distinct immune response against Mycobacterium tuberculosis (Mtb) at certain stages of infection as revealed by flow cytometry to date.

  9. Immune Response to Mycobacterial Infection: Lessons from Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Nikoletta Rovina

    2013-01-01

    Full Text Available Detecting and treating active and latent tuberculosis are pivotal elements for effective infection control; yet, due to their significant inherent limitations, the diagnostic means for these two stages of tuberculosis (TB to date remain suboptimal. This paper reviews the current diagnostic tools for mycobacterial infection and focuses on the application of flow cytometry as a promising method for rapid and reliable diagnosis of mycobacterial infection as well as discrimination between active and latent TB: it summarizes diagnostic biomarkers distinguishing the two states of infection and also features of the distinct immune response against Mycobacterium tuberculosis (Mtb at certain stages of infection as revealed by flow cytometry to date.

  10. Data File Standard for Flow Cytometry, version FCS 3.1.

    Science.gov (United States)

    Spidlen, Josef; Moore, Wayne; Parks, David; Goldberg, Michael; Bray, Chris; Bierre, Pierre; Gorombey, Peter; Hyun, Bill; Hubbard, Mark; Lange, Simon; Lefebvre, Ray; Leif, Robert; Novo, David; Ostruszka, Leo; Treister, Adam; Wood, James; Murphy, Robert F; Roederer, Mario; Sudar, Damir; Zigon, Robert; Brinkman, Ryan R

    2010-01-01

    The flow cytometry data file standard provides the specifications needed to completely describe flow cytometry data sets within the confines of the file containing the experimental data. In 1984, the first Flow Cytometry Standard format for data files was adopted as FCS 1.0. This standard was modified in 1990 as FCS 2.0 and again in 1997 as FCS 3.0. We report here on the next generation flow cytometry standard data file format. FCS 3.1 is a minor revision based on suggested improvements from the community. The unchanged goal of the standard is to provide a uniform file format that allows files created by one type of acquisition hardware and software to be analyzed by any other type.The FCS 3.1 standard retains the basic FCS file structure and most features of previous versions of the standard. Changes included in FCS 3.1 address potential ambiguities in the previous versions and provide a more robust standard. The major changes include simplified support for international characters and improved support for storing compensation. The major additions are support for preferred display scale, a standardized way of capturing the sample volume, information about originality of the data file, and support for plate and well identification in high throughput, plate based experiments. Please see the normative version of the FCS 3.1 specification in Supporting Information for this manuscript (or at http://www.isac-net.org/ in the Current standards section) for a complete list of changes.

  11. Mass Cytometry for Detection of Silver at the Bacterial Single Cell Level

    Directory of Open Access Journals (Sweden)

    Yuting Guo

    2017-07-01

    Full Text Available Background: Mass cytometry (Cytometry by Time of Flight, CyTOF allows single-cell characterization on the basis of specific metal-based cell markers. In addition, other metals in the mass range such as silver can be detected per cell. Bacteria are known to be sensible to silver and a protocol was developed to measure both the number of affected cells per population and the quantities of silver per cell.Methods: For mass cytometry ruthenium red was used as a marker for all cells of a population while parallel application of cisplatin discriminated live from dead cells. Silver quantities per cell and frequencies of silver containing cells in a population were measured by mass cytometry. In addition, live/dead subpopulations were analyzed by flow cytometry and distinguished by cell sorting based on ruthenium red and propidium iodide double staining. Verification of the cells’ silver load was performed on the bulk level by using ICP-MS in combination with cell sorting. The protocol was developed by conveying both, fast and non-growing Pseudomonas putida cells as test organisms.Results: A workflow for labeling bacteria in order to be analyzed by mass cytometry was developed. Three different parameters were tested: ruthenium red provided counts for all bacterial cells in a population while consecutively applied cisplatin marked the frequency of dead cells. Apparent population heterogeneity was detected by different frequencies of silver containing cells. Silver quantities per cell were also well measurable. Generally, AgNP-10 treatment caused higher frequencies of dead cells, higher frequencies of silver containing cells and higher per-cell silver quantities. Due to an assumed chemical equilibrium of free and bound silver ions live and dead cells were associated with silver in equal quantities and this preferably during exponential growth. With ICP-MS up to 1.5 fg silver per bacterial cell were detected.Conclusion: An effective mass cytometry

  12. Individual Cell Based Traits Obtained by Scanning Flow-Cytometry Show Selection by Biotic and Abiotic Environmental Factors during a Phytoplankton Spring Bloom

    Science.gov (United States)

    Pomati, Francesco; Kraft, Nathan J. B.; Posch, Thomas; Eugster, Bettina; Jokela, Jukka; Ibelings, Bas W.

    2013-01-01

    In ecology and evolution, the primary challenge in understanding the processes that shape biodiversity is to assess the relationship between the phenotypic traits of organisms and the environment. Here we tested for selection on physio-morphological traits measured by scanning flow-cytometry at the individual level in phytoplankton communities under a temporally changing biotic and abiotic environment. Our aim was to study how high-frequency temporal changes in the environment influence biodiversity dynamics in a natural community. We focused on a spring bloom in Lake Zurich (Switzerland), characterized by rapid changes in phytoplankton, water conditions, nutrients and grazing (mainly mediated by herbivore ciliates). We described bloom dynamics in terms of taxonomic and trait-based diversity and found that diversity dynamics of trait-based groups were more pronounced than those of identified phytoplankton taxa. We characterized the linkage between measured phytoplankton traits, abiotic environmental factors and abundance of the main grazers and observed weak but significant correlations between changing abiotic and biotic conditions and measured size-related and fluorescence-related traits. We tested for deviations in observed community-wide distributions of focal traits from random patterns and found evidence for both clustering and even spacing of traits, occurring sporadically over the time series. Patterns were consistent with environmental filtering and phenotypic divergence under herbivore pressure, respectively. Size-related traits showed significant even spacing during the peak of herbivore abundance, suggesting that morphology-related traits were under selection from grazing. Pigment distribution within cells and colonies appeared instead to be associated with acclimation to temperature and water chemistry. We found support for trade-offs among grazing resistance and environmental tolerance traits, as well as for substantial periods of dynamics in which

  13. Image selection as a service for cloud computing environments

    KAUST Repository

    Filepp, Robert

    2010-12-01

    Customers of Cloud Services are expected to choose specific machine images to instantiate in order to host their workloads. Unfortunately very little information is provided to the users to enable them to make intelligent choices. We believe that as the number of images proliferates it will become increasingly difficult for users to decide effectively. Cloud service providers often allow their customers to instantiate standard system images, to modify their instances, and to store images of these customized instances for public or private future use. Storing modified instances as images enables customers to avoid re-provisioning and re-configuration of required resources thereby reducing their future costs. However Cloud service providers generally do not expose details regarding the configurations of the images in a rigorous canonical fashion nor offer services that assist clients in the best target image selection to support client transformation objectives. Rather, they allow customers to enter a free-form description of an image based on client\\'s best effort. This means in order to find a "best fit" image to instantiate, a human user must review potentially thousands of image descriptions, reading each description to evaluate its suitability as a platform to host their source application. Furthermore, the actual content of the selected image may differ greatly from its description. Finally, even images that have been customized and retained for future use may need additional provisioning and customization to accommodate specific needs. In this paper we propose a service that accumulates image configuration details in a canonical fashion and a further service that employs an algorithm to order images per best fit /least cost in conformance to user-specified policies. These services collectively facilitate workload transformation into enterprise cloud environments.

  14. Angularly-selective transmission imaging in a scanning electron microscope.

    Science.gov (United States)

    Holm, Jason; Keller, Robert R

    2016-08-01

    This work presents recent advances in transmission scanning electron microscopy (t-SEM) imaging control capabilities. A modular aperture system and a cantilever-style sample holder that enable comprehensive angular selectivity of forward-scattered electrons are described. When combined with a commercially available solid-state transmission detector having only basic bright-field and dark-field imaging capabilities, the advances described here enable numerous transmission imaging modes. Several examples are provided that demonstrate how contrast arising from diffraction to mass-thickness can be obtained. Unanticipated image contrast at some imaging conditions is also observed and addressed. Published by Elsevier B.V.

  15. Analysis of the Budding Yeast Cell Cycle by Flow Cytometry.

    Science.gov (United States)

    Rosebrock, Adam P

    2017-01-03

    DNA synthesis is one of the landmark events in the cell cycle: G 1 cells have one copy of the genome, S phase cells are actively engaged in DNA synthesis, and G 2 cells have twice as much nuclear DNA as G 1 cells. Cellular DNA content can be measured by staining with a fluorescent dye followed by a flow-cytometric readout. This method provides a quantitative measurement of cell cycle position on a cell-by-cell basis at high speed. Using flow cytometry, tens of thousands of single-cell measurements can be generated in a few seconds. This protocol details staining of cells of the budding yeast Saccharomyces cerevisiae for flow cytometry using Sytox Green dye in a method that can be scaled widely-from one sample to many thousands and operating on inputs ranging from 1 million to more than 100 million cells. Flow cytometry is preferred over light microscopy or Coulter analyses for the analysis of the cell cycle as DNA content and cell cycle position are being directly measured. © 2017 Cold Spring Harbor Laboratory Press.

  16. Installation of a flow cytometry facility and some applications in radiobiology

    International Nuclear Information System (INIS)

    Walsh, M.; Kellington, J.P.

    1988-01-01

    Flow cytometry has enormous potential in many areas of experimental pathology. Details of the installation and commissioning of a flow cytometer at the Harwell Laboratory are described. Following an explanation of the principles of flow cytometry, several applications to specific problems in radiobiology are discussed. Also included are results of some preliminary studies with the Harwell flow cytometer on samples such as blood, bone marrow, macrophages and cell cultures, and a discussion of future applications. (author)

  17. Method of detaching adherent cells for flow cytometry

    KAUST Repository

    Kaur, Mandeep

    2015-12-24

    In one aspect, a method for detaching adherent cells can include adding a cell lifting solution to the media including a sample of adherent cells and incubating the sample of adherent cells with the cell lifting solution. No scraping or pipetting is needed to facilitate cell detachment. The method do not require inactivation of cell lifting solution and no washing of detaching cells is required to remove cell lifting solution. Detached cells can be stained with dye in the presence of cell lifting solution and are further analyzed using flow cytometer. The method has been tested using 6 different cell lines, 4 different assays, two different plate formats (96 and 384 well plates) and two different flow cytometry instruments. The method is simple to perform, less time consuming, with no cell loss and makes high throughput flow cytometry on adherent cells a reality.

  18. Flow cytometry for the evaluation of anti-plasmodial activity of drugs on Plasmodium falciparum gametocytes

    Directory of Open Access Journals (Sweden)

    Pipy Bernard

    2010-02-01

    Full Text Available Abstract Background The activity of promising anti-malarial drugs against Plasmodium gametocytes is hard to evaluate even in vitro. This is because visual examination of stained smears, which is commonly used, is not totally convenient. In the current study, flow cytometry has been used to study the effect of established anti-malarial drugs against sexual stages obtained from W2 strain of Plasmodium falciparum. Gametocytes were treated for 48 h with different drug concentrations and the gametocytaemia was then determined by flow cytometry and compared with visual estimation by microscopy. Results and conclusions Initially gametocytaemia was evaluated either using light microscopy or flow cytometry. A direct correlation (r2 = 0.9986 was obtained. Two distinct peaks were observed on cytometry histograms and were attributed to gametocyte populations. The activities of established anti-malarial compounds were then measured by flow cytometry and the results were equivalent to those obtained using light microscopy. Primaquine and artemisinin had IC50 of 17.6 μM and 1.0 μM, respectively. Gametocyte sex was apparently distinguishable by flow cytometry as evaluated after induction of exflagellation by xanthurenic acid. These data form the basis of further studies for developing new methods in drug discovery to decrease malaria transmission.

  19. Alternatives to current flow cytometry data analysis for clinical and research studies.

    Science.gov (United States)

    Gondhalekar, Carmen; Rajwa, Bartek; Patsekin, Valery; Ragheb, Kathy; Sturgis, Jennifer; Robinson, J Paul

    2018-02-01

    Flow cytometry has well-established methods for data analysis based on traditional data collection techniques. These techniques typically involved manual insertion of tube samples into an instrument that, historically, could only measure 1-3 colors. The field has since evolved to incorporate new technologies for faster and highly automated sample preparation and data collection. For example, the use of microwell plates on benchtop instruments is now a standard on virtually every new instrument, and so users can easily accumulate multiple data sets quickly. Further, because the user must carefully define the layout of the plate, this information is already defined when considering the analytical process, expanding the opportunities for automated analysis. Advances in multi-parametric data collection, as demonstrated by the development of hyperspectral flow-cytometry, 20-40 color polychromatic flow cytometry, and mass cytometry (CyTOF), are game-changing. As data and assay complexity increase, so too does the complexity of data analysis. Complex data analysis is already a challenge to traditional flow cytometry software. New methods for reviewing large and complex data sets can provide rapid insight into processes difficult to define without more advanced analytical tools. In settings such as clinical labs where rapid and accurate data analysis is a priority, rapid, efficient and intuitive software is needed. This paper outlines opportunities for analysis of complex data sets using examples of multiplexed bead-based assays, drug screens and cell cycle analysis - any of which could become integrated into the clinical environment. Copyright © 2017. Published by Elsevier Inc.

  20. Band Subset Selection for Hyperspectral Image Classification

    Directory of Open Access Journals (Sweden)

    Chunyan Yu

    2018-01-01

    Full Text Available This paper develops a new approach to band subset selection (BSS for hyperspectral image classification (HSIC which selects multiple bands simultaneously as a band subset, referred to as simultaneous multiple band selection (SMMBS, rather than one band at a time sequentially, referred to as sequential multiple band selection (SQMBS, as most traditional band selection methods do. In doing so, a criterion is particularly developed for BSS that can be used for HSIC. It is a linearly constrained minimum variance (LCMV derived from adaptive beamforming in array signal processing which can be used to model misclassification errors as the minimum variance. To avoid an exhaustive search for all possible band subsets, two numerical algorithms, referred to as sequential (SQ and successive (SC algorithms are also developed for LCMV-based SMMBS, called SQ LCMV-BSS and SC LCMV-BSS. Experimental results demonstrate that LCMV-based BSS has advantages over SQMBS.

  1. Flow cytometry as an improved method for the titration of Chlamydiaceae and other intracellular bacteria.

    Science.gov (United States)

    Käser, T; Pasternak, J A; Hamonic, G; Rieder, M; Lai, K; Delgado-Ortega, M; Gerdts, V; Meurens, F

    2016-05-01

    Chlamydiaceae is a family of intracellular bacteria causing a range of diverse pathological outcomes. The most devastating human diseases are ocular infections with C. trachomatis leading to blindness and genital infections causing pelvic inflammatory disease with long-term sequelae including infertility and chronic pelvic pain. In order to enable the comparison of experiments between laboratories investigating host-chlamydia interactions, the infectious titer has to be determined. Titer determination of chlamydia is most commonly performed via microscopy of host cells infected with a serial dilution of chlamydia. However, other methods including fluorescent ELISpot (Fluorospot) and DNA Chip Scanning Technology have also been proposed to enumerate chlamydia-infected cells. For viruses, flow cytometry has been suggested as a superior alternative to standard titration methods. In this study we compared the use of flow cytometry with microscopy and Fluorospot for the titration of C. suis as a representative of other intracellular bacteria. Titer determination via Fluorospot was unreliable, while titration via microscopy led to a linear read-out range of 16 - 64 dilutions and moderate reproducibility with acceptable standard deviations within and between investigators. In contrast, flow cytometry had a vast linear read-out range of 1,024 dilutions and the lowest standard deviations given a basic training in these methods. In addition, flow cytometry was faster and material costs were lower compared to microscopy. Flow cytometry offers a fast, cheap, precise, and reproducible alternative for the titration of intracellular bacteria like C. suis. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

  2. Wild immunology assessed by multidimensional mass cytometry.

    Science.gov (United States)

    Japp, Alberto Sada; Hoffmann, Kerstin; Schlickeiser, Stephan; Glauben, Rainer; Nikolaou, Christos; Maecker, Holden T; Braun, Julian; Matzmohr, Nadine; Sawitzki, Birgit; Siegmund, Britta; Radbruch, Andreas; Volk, Hans-Dieter; Frentsch, Marco; Kunkel, Desiree; Thiel, Andreas

    2017-01-01

    A great part of our knowledge on mammalian immunology has been established in laboratory settings. The use of inbred mouse strains enabled controlled studies of immune cell and molecule functions in defined settings. These studies were usually performed in specific-pathogen free (SPF) environments providing standardized conditions. In contrast, mammalians including humans living in their natural habitat are continuously facing pathogen encounters throughout their life. The influences of environmental conditions on the signatures of the immune system and on experimental outcomes are yet not well defined. Thus, the transferability of results obtained in current experimental systems to the physiological human situation has always been a matter of debate. Studies elucidating the diversity of "wild immunology" imprintings in detail and comparing it with those of "clean" lab mice are sparse. Here, we applied multidimensional mass cytometry to dissect phenotypic and functional differences between distinct groups of laboratory and pet shop mice as a source for "wild mice". For this purpose, we developed a 31-antibody panel for murine leukocyte subsets identification and a 35-antibody panel assessing various cytokines. Established murine leukocyte populations were easily identified and diverse immune signatures indicative of numerous pathogen encounters were classified particularly in pet shop mice and to a lesser extent in quarantine and non-SPF mice as compared to SPF mice. In addition, unsupervised analysis identified distinct clusters that associated strongly with the degree of pathogenic priming, including increased frequencies of activated NK cells and antigen-experienced B- and T-cell subsets. Our study unravels the complexity of immune signatures altered under physiological pathogen challenges and highlights the importance of carefully adapting laboratory settings for immunological studies in mice, including drug and therapy testing. © 2016 International Society

  3. Multi-frequency Defect Selective Imaging via Nonlinear Ultrasound

    Science.gov (United States)

    Solodov, Igor; Busse, Gerd

    The concept of defect-selective ultrasonic nonlinear imaging is based on visualization of strongly nonlinear inclusions in the form of localized cracked defects. For intense excitation, the ultrasonic response of defects is affected by mechanical constraint between their fragments that makes their vibrations extremely nonlinear. The cracked flaws, therefore, efficiently generate multiple new frequencies, which can be used as a nonlinear "tag" to detect and image them. In this paper, the methodologies of nonlinear scanning laser vibrometry (NSLV) and nonlinear air-coupled emission (NACE) are applied for nonlinear imaging of various defects in hi-tech and constructional materials. A broad database obtained demonstrates evident advantages of the nonlinear approach over its linear counterpart. The higher-order nonlinear frequencies provide increase in signal-to-noise ratio and enhance the contrast of imaging. Unlike conventional ultrasonic instruments, the nonlinear approach yields abundant multi-frequency information on defect location. The application of image recognition and processing algorithms is described and shown to improve reliability and quality of ultrasonic imaging.

  4. [Flow cytometry in datecting lymph node micrometastasis in colorectal cancer].

    Science.gov (United States)

    Sun, Q; Ding, Y; Zhang, J

    2001-01-25

    To study the methodology and significance of flow cytometry in detecting lymph node micrometastasis of colorectal cancer. One hundred sixty-two cellular suspensions were prepared with lymph nodes which were resected radically on 25 patients with colorectal cancer and in which no cancer cells were found by HE staining. Different concentrations of cultured Lovo colorectal cancer cells were added into the celular suspension prepared from lymph node tissue of persons without colorectal cancer in order to prepare a control model. Dual staining with CK/FTTC and PI was made to the sedimetns from those 2 kinds of suspension. Flow cytometry was used to detect cancer cells. An ideal correlation was obtained between the detection value and the theoretical value of cancer cells in the specimen suspensions and control models (r = 0.097 6) with a sensitivity rate of 10/10(5). Cancer cells were detected from 7 out of the 25 patients and 30 of the 162 cellular suspensions. The detection rate was correlated with the size and infiltrating depth of the cancer. Flow cytometry is a reliable, rapid, and quantitative method for detecting lymph node micrometastasis in colorectal cancer.

  5. Sorting catalytically active polymersome nanoreactors by flow cytometry

    NARCIS (Netherlands)

    Nallani, M.; Woestenenk, R.; de Hoog, H.P.M.; van Dongen, S.F.M.; Boezeman, J.; Cornelissen, J.J.L.M.; Nolte, R.J.M.; van Hest, J.C.M.

    2009-01-01

    A strategy that involves a versatile one-step preparation procedure of enzyme filled porous and stable polymeric catalytically active nanoreactors (polymersomes) by flow cytometry was reported. A 1:1 mixture of the polymerase dispersions was analyzed in a Coulter Epics Elite Flow Cytometer, while

  6. Automated training site selection for large-area remote-sensing image analysis

    Science.gov (United States)

    McCaffrey, Thomas M.; Franklin, Steven E.

    1993-11-01

    A computer program is presented to select training sites automatically from remotely sensed digital imagery. The basic ideas are to guide the image analyst through the process of selecting typical and representative areas for large-area image classifications by minimizing bias, and to provide an initial list of potential classes for which training sites are required to develop a classification scheme or to verify classification accuracy. Reducing subjectivity in training site selection is achieved by using a purely statistical selection of homogeneous sites which then can be compared to field knowledge, aerial photography, or other remote-sensing imagery and ancillary data to arrive at a final selection of sites to be used to train the classification decision rules. The selection of the homogeneous sites uses simple tests based on the coefficient of variance, the F-statistic, and the Student's i-statistic. Comparisons of site means are conducted with a linear growing list of previously located homogeneous pixels. The program supports a common pixel-interleaved digital image format and has been tested on aerial and satellite optical imagery. The program is coded efficiently in the C programming language and was developed under AIX-Unix on an IBM RISC 6000 24-bit color workstation.

  7. Surface profiling of normally responding and nonreleasing basophils by flow cytometry

    DEFF Research Database (Denmark)

    Kistrup, Kasper; Poulsen, Lars Kærgaard; Jensen, Bettina Margrethe

    a maximum release blood mononuclear cells were purified by density centrifugation and using flow cytometry, basophils, defined as FceRIa+CD3-CD14-CD19-CD56-,were analysed for surface expression of relevant markers. All samples were compensated and analysed in logicle display. All gates......c, C3aR, C5aR CCR3, FPR1, ST2, CRTH2 on anti-IgE respondsive and nonreleasing basophils by flow cytometry, thereby generating a surface profile of the two phenotypes. Methods Fresh buffy coat blood (

  8. Selectable six-element multicoil array for entire spine imaging

    International Nuclear Information System (INIS)

    Byrne, J.W.; Bluma-Walter, J.; Prorok, R.J.

    1990-01-01

    This article introduces a new multicoil array that can provide entire spine imaging in two acquisitions with no need to manually reposition either the coil or the patient. A selectable contoured multicoil array with six elements was used to obtain coverage of the entire spine. The first four elements were used for imaging the upper spine region (cervical/thoracic) during the first acquisition, and the last four elements were used for imaging the lower spine region (thoracic/lumbar) during the second acquisition. The overall coil length was approximately 75 cm

  9. Honey bee hemocyte profiling by flow cytometry.

    Science.gov (United States)

    Marringa, William J; Krueger, Michael J; Burritt, Nancy L; Burritt, James B

    2014-01-01

    Multiple stress factors in honey bees are causing loss of bee colonies worldwide. Several infectious agents of bees are believed to contribute to this problem. The mechanisms of honey bee immunity are not completely understood, in part due to limited information about the types and abundances of hemocytes that help bees resist disease. Our study utilized flow cytometry and microscopy to examine populations of hemolymph particulates in honey bees. We found bee hemolymph includes permeabilized cells, plasmatocytes, and acellular objects that resemble microparticles, listed in order of increasing abundance. The permeabilized cells and plasmatocytes showed unexpected differences with respect to properties of the plasma membrane and labeling with annexin V. Both permeabilized cells and plasmatocytes failed to show measurable mitochondrial membrane potential by flow cytometry using the JC-1 probe. Our results suggest hemolymph particulate populations are dynamic, revealing significant differences when comparing individual hive members, and when comparing colonies exposed to diverse conditions. Shifts in hemocyte populations in bees likely represent changing conditions or metabolic differences of colony members. A better understanding of hemocyte profiles may provide insight into physiological responses of honey bees to stress factors, some of which may be related to colony failure.

  10. A trial of selective imaging for the breast mass shadow by computed radiography

    International Nuclear Information System (INIS)

    Muramatsu, Yukio; Anan, Mitsuhiro; Tanaka, Takashi; Matsue, Hiroto; Yamada, Tatsuya

    1990-01-01

    CR has ability to make many kinds of images by several imaging processings. Especially, gradation processing is more important than frequency processing to make images in CR mammography. We developed new method to image breast masses selectively with new gradation processing and tried it for 18 patients over sixty years old with breast cancer. All of breast mass shadows were separated selectively from other parencymal shadow. So, we conclude that the auto-recognition of breast mass shadow can be possible in near future in CR system. (author)

  11. Screening of carcinoma metastasis by flow cytometry: A study of 238 cases.

    Science.gov (United States)

    Acosta, Maria; Pereira, José; Arroz, Maria

    2016-05-01

    Malignant epithelial cells may be detected in different specimens, by immunophenotyping using flow cytometry (FCM). CD326 (epithelial-specific antigen, clone Ber-Ep4) was used to identify epithelial cells, CD45 to discriminate between leucocytes (positive for this antigen) and non-hematological cells (negative for this antigen), and CD33 to identify monocytes/macrophages. This combination is particularly useful in effusions to characterize large cells and distinguish between monocyte/macrophages (CD45+ CD33+ CD326-), mesothelial cells (CD45 ± (dim) CD33 - CD326-) and epithelial cells (CD45 - CD33 - CD326 +). We evaluated the efficiency of flow cytometry to detect malignant epithelial cells in 238 fresh samples, including effusions, lymph node biopsies, fine needle aspirates, bone marrow aspirates, cerebrospinal fluid, among others. These are specimens expected to lack epithelial cells. FCM results were then compared to the results of smear and cell block morphology, as well as immunocytochemistry on paraffin wax embedded cell blocks, when available. Final diagnosis was the gold standard and a very good sensitivity (96.7%) and specificity (99.3%) were obtained. We concluded that the detection of CD326 positive cells using FCM is strongly indicative of the presence of carcinoma cells. © 2015 International Clinical Cytometry Society. © 2015 International Clinical Cytometry Society.

  12. The Means: Cytometry and Mass Spectrometry Converge in a Single Cell Deep Profiling Platform

    Science.gov (United States)

    Weis-Garcia, Frances; Bandura, Dmitry; Baranov, Vladimir; Ornatsky, Olga; Tanner, Scott

    2013-01-01

    Inductively Coupled Plasma Mass Spectrometry (ICP-MS) is a distinct flavor of mass spectrometry that has had little association with cell biology: it remains the state of the art for the determination of the atomic composition of materials. Unrelatedly, flow cytometry is the superior method for distinguishing the heterogeneity of cells through the determination of antigen signatures using tagged antibodies. Simply replacing fluorophore tags with stable isotopes of the heavy metals, and measuring these cell-by-cell with ICP-MS, dramatically increases the number of probes that can be simultaneously measured in cytometry and enables a transformative increase in the resolution of rare cell populations in complex biological samples. While this can be thought of as a novel incarnation of single-cell targeted proteomics, the metal-labeling reagents, ICP-MS of single cells, and accompanying informatics comprise a new field of technology termed Mass Cytometry. While the conception of mass cytometry is simple the embodiment to address the issues of multi-parameter flow cytometry has been far more challenging. There are many elements, and many more stable isotopes of those elements, that might be used as distinct reporter tags. Still, there are many approaches to conjugating metals to antibodies (or other affinity reagents) and work in this area along with developing new applications is ongoing. The mass resolution and linear (quantitative) dynamic range of ICP-MS allows those many stable isotopes to be measured simultaneously and without the spectral overlap issues that limit fluorescence assay. However, the adaptation of ICP-MS to allow high-speed simultaneous measurement with single cell distinction at high throughput required innovation of the cell introduction system, ion optics (sampling, transmission and beam-shaping), mass analysis, and signal handling and processing. An overview of “the nuts and bolts” of Mass Cytometry is presented.

  13. Methodology and application of flow cytometry for investigation of human malaria parasites.

    Science.gov (United States)

    Grimberg, Brian T

    2011-03-31

    Historically, examinations of the inhibition of malaria parasite growth/invasion, whether using drugs or antibodies, have relied on the use of microscopy or radioactive hypoxanthine uptake. These are considered gold standards for measuring the effectiveness of antimalarial treatments, however, these methods have well known shortcomings. With the advent of flow cytometry coupled with the use of fluorescent DNA stains allowed for increased speed, reproducibility, and qualitative estimates of the effectiveness of antibodies and drugs to limit malaria parasite growth which addresses the challenges of traditional techniques. Because materials and machines available to research facilities are so varied, different methods have been developed to investigate malaria parasites by flow cytometry. This review is intended to serve as a reference guide for advanced users and importantly, as a primer for new users, to support expanded use and improvements to malaria flow cytometry, particularly in endemic countries. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Effect of using different cover image quality to obtain robust selective embedding in steganography

    Science.gov (United States)

    Abdullah, Karwan Asaad; Al-Jawad, Naseer; Abdulla, Alan Anwer

    2014-05-01

    One of the common types of steganography is to conceal an image as a secret message in another image which normally called a cover image; the resulting image is called a stego image. The aim of this paper is to investigate the effect of using different cover image quality, and also analyse the use of different bit-plane in term of robustness against well-known active attacks such as gamma, statistical filters, and linear spatial filters. The secret messages are embedded in higher bit-plane, i.e. in other than Least Significant Bit (LSB), in order to resist active attacks. The embedding process is performed in three major steps: First, the embedding algorithm is selectively identifying useful areas (blocks) for embedding based on its lighting condition. Second, is to nominate the most useful blocks for embedding based on their entropy and average. Third, is to select the right bit-plane for embedding. This kind of block selection made the embedding process scatters the secret message(s) randomly around the cover image. Different tests have been performed for selecting a proper block size and this is related to the nature of the used cover image. Our proposed method suggests a suitable embedding bit-plane as well as the right blocks for the embedding. Experimental results demonstrate that different image quality used for the cover images will have an effect when the stego image is attacked by different active attacks. Although the secret messages are embedded in higher bit-plane, but they cannot be recognised visually within the stegos image.

  15. A flow cytometry-based method for a high-throughput analysis of drug-stabilized topoisomerase II cleavage complexes in human cells.

    Science.gov (United States)

    de Campos-Nebel, Marcelo; Palmitelli, Micaela; González-Cid, Marcela

    2016-09-01

    Topoisomerase II (Top2) is an important target for anticancer therapy. A variety of drugs that poison Top2, including several epipodophyllotoxins, anthracyclines, and anthracenediones, are widely used in the clinic for both hematologic and solid tumors. The poisoning of Top2 involves the formation of a reaction intermediate Top2-DNA, termed Top2 cleavage complex (Top2cc), which is persistent in the presence of the drug and involves a 5' end of DNA covalently bound to a tyrosine from the enzyme through a phosphodiester group. Drug-induced Top2cc leads to Top2 linked-DNA breaks which are the major responsible for their cytotoxicity. While biochemical detection is very laborious, quantification of drug-induced Top2cc by immunofluorescence-based microscopy techniques is time consuming and requires extensive image segmentation for the analysis of a small population of cells. Here, we developed a flow cytometry-based method for the analysis of drug-induced Top2cc. This method allows a rapid analysis of a high number of cells in their cell cycle phase context. Moreover, it can be applied to almost any human cell type, including clinical samples. The methodology is useful for a high-throughput analysis of drugs that poison Top2, allowing not just the discrimination of the Top2 isoform that is targeted but also to track its removal. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

  16. International Society for the Advancement of Cytometry cell sorter biosafety standards.

    Science.gov (United States)

    Holmes, Kevin L; Fontes, Benjamin; Hogarth, Philip; Konz, Richard; Monard, Simon; Pletcher, Charles H; Wadley, Robert B; Schmid, Ingrid; Perfetto, Stephen P

    2014-05-01

    Flow cytometric cell sorting of biological specimens has become prevalent in basic and clinical research laboratories. These specimens may contain known or unknown infectious agents, necessitating precautions to protect instrument operators and the environment from biohazards arising from the use of sorters. To this end the International Society of Analytical Cytology (ISAC) was proactive in establishing biosafety guidelines in 1997 (Schmid et al., Cytometry 1997;28:99-117) and subsequently published revised biosafety standards for cell sorting of unfixed samples in 2007 (Schmid et al., Cytometry Part A J Int Soc Anal Cytol 2007;71A:414-437). Since their publication, these documents have become recognized worldwide as the standard of practice and safety precautions for laboratories performing cell sorting experiments. However, the field of cytometry has progressed since 2007, and the document requires an update. The new Standards provides guidance: (1) for laboratory design for cell sorter laboratories; (2) for the creation of laboratory or instrument specific Standard Operating Procedures (SOP); and (3) on procedures for the safe operation of cell sorters, including personal protective equipment (PPE) and validation of aerosol containment. Published © 2014 Wiley Periodicals Inc.

  17. Analysis of repetitive DNA in chromosomes by flow cytometry

    NARCIS (Netherlands)

    Brind'Amour, Julie; Lansdorp, Peter M.

    We developed a flow cytometry method, chromosome flow fluorescence in situ hybridization (FISH), called CFF, to analyze repetitive DNA in chromosomes using FISH with directly labeled peptide nucleic acid (PNA) probes. We used CFF to measure the abundance of interstitial telomeric sequences in

  18. DIRECT FLOW-CYTOMETRY OF ANAEROBIC-BACTERIA IN HUMAN FECES

    NARCIS (Netherlands)

    VANDERWAAIJ, LA; MESANDER, G; LIMBURG, PC; VANDERWAAIJ, D

    1994-01-01

    We describe a flow cytometry method for analysis of noncultured anaerobic bacteria present in human fecal suspensions. Nonbacterial fecal compounds, bacterial fragments, and large aggregates could be discriminated from bacteria by staining with propidium iodide (PI) and setting a discriminator on PI

  19. Gating-ML: XML-based gating descriptions in flow cytometry.

    Science.gov (United States)

    Spidlen, Josef; Leif, Robert C; Moore, Wayne; Roederer, Mario; Brinkman, Ryan R

    2008-12-01

    The lack of software interoperability with respect to gating due to lack of a standardized mechanism for data exchange has traditionally been a bottleneck, preventing reproducibility of flow cytometry (FCM) data analysis and the usage of multiple analytical tools. To facilitate interoperability among FCM data analysis tools, members of the International Society for the Advancement of Cytometry (ISAC) Data Standards Task Force (DSTF) have developed an XML-based mechanism to formally describe gates (Gating-ML). Gating-ML, an open specification for encoding gating, data transformations and compensation, has been adopted by the ISAC DSTF as a Candidate Recommendation. Gating-ML can facilitate exchange of gating descriptions the same way that FCS facilitated for exchange of raw FCM data. Its adoption will open new collaborative opportunities as well as possibilities for advanced analyses and methods development. The ISAC DSTF is satisfied that the standard addresses the requirements for a gating exchange standard.

  20. Flow cytometry susceptibility testing for conventional antifungal drugs and Comparison with the NCCLS Broth Macrodilution Test

    Directory of Open Access Journals (Sweden)

    M.J. Najafzadeh

    2009-08-01

    Full Text Available Introduction: During the last decade, the incidence of fungal infection has been increased in many countries. Because of the advent of resistant to antifungal agents, determination of an efficient strategic plan for treatment of fungal disease is an important issue in clinical mycology. Many methods have been introduced and developed for determination of invitro susceptibility tests. During the recent years, flow cytometry has developed to solving the problem and many papers have documented the usefulness of this technique. Materials and methods: As the first step, the invitro susceptibility of standard PTCC (Persian Type of Culture Collection strain and some clinical isolates of Candida consisting of Candida albicans, C. dubliniensis, C. glabrata, C. kefyer and C. parapsilosis were evaluated by macrodilution broth method according to NCCLS (National Committee for Clinical Laboratory Standards guidelines and flow cytometry susceptibility test. Results:  The data indicated that macro dilution broth methods and flow cytometry have the same results in determination of MIC (Minimum Inhibitory Concentration for amphotericin B, clotrimazole, fluconazole, ketoconazole and miconazole in C. albicans PTCC 5027 as well as clinical Candida isolates, such as C.albicans, C.dubliniensis, C.glabrata C.kefyr, and C.parapsilosis. Discussion: Comparing the results obtained by macrodilution broth and flow cytometry methods revealed that flow cytometry was faster. It is suggested that flow cytometry susceptibility test can be used as a powerful tool for determination of MIC and administration of the best antifungal drug in treatment of patients with Candida infections.

  1. Content dependent selection of image enhancement parameters for mobile displays

    Science.gov (United States)

    Lee, Yoon-Gyoo; Kang, Yoo-Jin; Kim, Han-Eol; Kim, Ka-Hee; Kim, Choon-Woo

    2011-01-01

    Mobile devices such as cellular phones and portable multimedia player with capability of playing terrestrial digital multimedia broadcasting (T-DMB) contents have been introduced into consumer market. In this paper, content dependent image quality enhancement method for sharpness and colorfulness and noise reduction is presented to improve perceived image quality on mobile displays. Human visual experiments are performed to analyze viewers' preference. Relationship between the objective measures and the optimal values of image control parameters are modeled by simple lookup tables based on the results of human visual experiments. Content dependent values of image control parameters are determined based on the calculated measures and predetermined lookup tables. Experimental results indicate that dynamic selection of image control parameters yields better image quality.

  2. The effect of base image window level selection on the dimensional measurement accuracy of resultant three-dimensional image displays

    International Nuclear Information System (INIS)

    Kurmis, A.P.; Hearn, T.C.; Reynolds, K.J.

    2003-01-01

    Purpose: The aim of this study was to determine the effect of base image window level selection on direct linear measurement of knee structures displayed using new magnetic resonance (MR)-based three-dimensional reconstructed computer imaging techniques. Methods: A prospective comparative study was performed using a series of three-dimensional knee images, generated from conventional MR imaging (MRI) sections. Thirty distinct anatomical structural features were identified within the image series of which repeated measurements were compared at 10 different window grey scale levels. Results: Statistical analysis demonstrated an excellent raw correlation between measurements and suggested no significant difference between measurements made at each of the 10 window level settings (P>0.05). Conclusions: The findings of this study suggest that unlike conventional MR or CT applications, grey scale window level selection at the time of imaging does not significantly affect the visual quality of resultant three-dimensional reconstructed images and hence the accuracy of subsequent direct linear measurement. The diagnostic potential of clinical progression from routine two-dimensional to advanced three-dimensional reconstructed imaging techniques may therefore be less likely to be degraded by inappropriate MR technician image windowing during the capturing of image series

  3. Hyperspectral band selection and classification of Hyperion image of Bhitarkanika mangrove ecosystem, eastern India

    Science.gov (United States)

    Ashokkumar, L.; Shanmugam, S.

    2014-10-01

    Tropical mangrove forests along the coast evolve dynamically due to constant changes in the natural ecosystem and ecological cycle. Remote sensing has paved the way for periodic monitoring and conservation of such floristic resources, compared to labour intensive in-situ observations. With the laboratory quality image spectra obtained from hyperspectral image data, species level discrimination in habitats and ecosystems is attainable. One of the essential steps before classification of hyperspectral image data is band selection. It is important to eliminate the redundant bands to mitigate the problems of Hughes effect that are likely to affect further image analysis and classification accuracy. This paper presents a methodology for the selection of appropriate hyperspectral bands from the EO-1 Hyperion image for the identification and mapping of mangrove species and coastal landcover types in the Bhitarkanika coastal forest region, eastern India. Band selection procedure follows class based elimination procedure and the separability of the classes are tested in the band selection process. Individual bands are de-correlated and redundant bands are removed from the bandwise correlation matrix. The percent contribution of class variance in each band is analysed from the factors of PCA component ranking. Spectral bands are selected from the wavelength groups and statistically tested. Further, the band selection procedure is compared with similar techniques (Band Index and Mutual information) for validation. The number of bands in the Hyperion image was reduced from 196 to 88 by the Factor-based ranking approach. Classification was performed by Support Vector Machine approach. It is observed that the proposed Factor-based ranking approach performed well in discriminating the mangrove species and other landcover units compared to the other statistical approaches. The predominant mangrove species Heritiera fomes, Excoecaria agallocha and Cynometra ramiflora are spectral

  4. Report of the European Myeloma Network on multiparametric flow cytometry in multiple myeloma and related disorders

    DEFF Research Database (Denmark)

    Rawstron, Andy C; Orfao, Alberto; Beksac, Meral

    2008-01-01

    The European Myeloma Network (EMN) organized two flow cytometry workshops. The first aimed to identify specific indications for flow cytometry in patients with monoclonal gammopathies, and consensus technical approaches through a questionnaire-based review of current practice in participating...

  5. Flow cytometry sorting of nuclei enables the first global characterization of Paramecium germline DNA and transposable elements.

    Science.gov (United States)

    Guérin, Frédéric; Arnaiz, Olivier; Boggetto, Nicole; Denby Wilkes, Cyril; Meyer, Eric; Sperling, Linda; Duharcourt, Sandra

    2017-04-26

    DNA elimination is developmentally programmed in a wide variety of eukaryotes, including unicellular ciliates, and leads to the generation of distinct germline and somatic genomes. The ciliate Paramecium tetraurelia harbors two types of nuclei with different functions and genome structures. The transcriptionally inactive micronucleus contains the complete germline genome, while the somatic macronucleus contains a reduced genome streamlined for gene expression. During development of the somatic macronucleus, the germline genome undergoes massive and reproducible DNA elimination events. Availability of both the somatic and germline genomes is essential to examine the genome changes that occur during programmed DNA elimination and ultimately decipher the mechanisms underlying the specific removal of germline-limited sequences. We developed a novel experimental approach that uses flow cell imaging and flow cytometry to sort subpopulations of nuclei to high purity. We sorted vegetative micronuclei and macronuclei during development of P. tetraurelia. We validated the method by flow cell imaging and by high throughput DNA sequencing. Our work establishes the proof of principle that developing somatic macronuclei can be sorted from a complex biological sample to high purity based on their size, shape and DNA content. This method enabled us to sequence, for the first time, the germline DNA from pure micronuclei and to identify novel transposable elements. Sequencing the germline DNA confirms that the Pgm domesticated transposase is required for the excision of all ~45,000 Internal Eliminated Sequences. Comparison of the germline DNA and unrearranged DNA obtained from PGM-silenced cells reveals that the latter does not provide a faithful representation of the germline genome. We developed a flow cytometry-based method to purify P. tetraurelia nuclei to high purity and provided quality control with flow cell imaging and high throughput DNA sequencing. We identified 61

  6. Primary gamma ray selection in a hybrid timing/imaging Cherenkov array

    Directory of Open Access Journals (Sweden)

    Postnikov E.B.

    2017-01-01

    Full Text Available This work is a methodical study on hybrid reconstruction techniques for hybrid imaging/timing Cherenkov observations. This type of hybrid array is to be realized at the gamma-observatory TAIGA intended for very high energy gamma-ray astronomy (> 30 TeV. It aims at combining the cost-effective timing-array technique with imaging telescopes. Hybrid operation of both of these techniques can lead to a relatively cheap way of development of a large area array. The joint approach of gamma event selection was investigated on both types of simulated data: the image parameters from the telescopes, and the shower parameters reconstructed from the timing array. The optimal set of imaging parameters and shower parameters to be combined is revealed. The cosmic ray background suppression factor depending on distance and energy is calculated. The optimal selection technique leads to cosmic ray background suppression of about 2 orders of magnitude on distances up to 450 m for energies greater than 50 TeV.

  7. Rapid detection of aneuploidy in Musa using flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Roux, N.; Toloza, A.; Radecki, Z.; Zapata-Arias, F. J.; Doležel, Jaroslav

    2003-01-01

    Roč. 21, - (2003), s. 483-490 ISSN 0721-7714 R&D Projects: GA AV ČR IAA6038204 Institutional research plan: CEZ:AV0Z5038910 Keywords : banana * flow cytometry * nuclear DNA content Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.423, year: 2003

  8. Parametric images evaluation of selected phases of the heart cycle with PET

    International Nuclear Information System (INIS)

    Just, U.; Will, E.; Beuthien Baumann, B.; Bredow, J.

    2002-01-01

    The standard evaluation of dynamic heart acquisitions with PET uses image data not corrected for heart wall movement. The evaluation of parametric data sets (Patlak Plot) was investigated for gated studies of selected heart phases (diastolic, systolic) and compared to the standard evaluation. Parametric images of selected heart phases have improved resolution. The values for metabolic rate are different for a ''normal'' and gated evaluation, up to 50% more for the systole compared to the normal one. (orig.)

  9. Aptamer-fluorescent silica nanoparticles bioconjugates based dual-color flow cytometry for specific detection of Staphylococcus aureus.

    Science.gov (United States)

    He, Xiaoxiao; Li, Yuhong; He, Dinggen; Wang, Kemin; Shangguan, Jingfang; Shi, Hui

    2014-07-01

    This paper describes a sensitive and specific determination strategy for Staphylococcus aureus (S. aureus) detection using aptamer recognition and fluorescent silica nanoparticles (FSiNPs) label based dual-color flow cytometry assay (Aptamer/FSiNPs-DCFCM). In the protocol, an aptamer, having high affinity to S. aureus, was first covalently immobilized onto chloropropyl functionalized FSiNPs through a click chemistry approach to generate aptamer-nanoparticles bioconjugates (Aptamer/FSiNPs). Next, S. aureus was incubated with Aptamer/FSiNPs, and then stained with SYBR Green I (a special staining material for the duplex DNA). Upon target binding and nucleic acid staining with SYBR Green I, the S. aureus was determined using two-color flow cytometry. The method took advantage of the specificity of aptamer, signal amplification of FSiNPs label and decreased false positives of two-color flow cytometry assay. It was demonstrated that these Aptamer/FSiNPs could efficiently recognize and fluorescently label target S. aureus. Through multiparameter determination with flow cytometry, this assay allowed for detection of as low as 1.5 x 10(2) and 7.6 x 10(2) cells mL(-1) S. aureus in buffer and spiked milk, respectively, with higher sensitivity than the Aptamer/FITC based flow cytometry.

  10. Candidiasis and the impact of flow cytometry on antifungal drug discovery.

    Science.gov (United States)

    Ku, Tsun Sheng N; Bernardo, Stella; Walraven, Carla J; Lee, Samuel A

    2017-11-01

    Invasive candidiasis continues to be associated with significant morbidity and mortality as well as substantial health care costs nationally and globally. One of the contributing factors is the development of resistance to antifungal agents that are already in clinical use. Moreover, there are known treatment limitations with all of the available antifungal agents. Since traditional techniques in novel drug discovery are time consuming, high-throughput screening using flow cytometry presents as a potential tool to identify new antifungal agents that would be useful in the management of these patients. Areas covered: In this review, the authors discuss the use of automated high-throughput screening assays based upon flow cytometry to identify potential antifungals from a library comprised of a large number of bioactive compounds. They also review studies that employed the use of this research methodology that has identified compounds with antifungal activity. Expert opinion: High-throughput screening using flow cytometry has substantially decreased the processing time necessary for screening thousands of compounds, and has helped enhance our understanding of fungal pathogenesis. Indeed, the authors see this technology as a powerful tool to help scientists identify new antifungal agents that can be added to the clinician's arsenal in their fight against invasive candidiasis.

  11. flowClust: a Bioconductor package for automated gating of flow cytometry data

    Directory of Open Access Journals (Sweden)

    Lo Kenneth

    2009-05-01

    Full Text Available Abstract Background As a high-throughput technology that offers rapid quantification of multidimensional characteristics for millions of cells, flow cytometry (FCM is widely used in health research, medical diagnosis and treatment, and vaccine development. Nevertheless, there is an increasing concern about the lack of appropriate software tools to provide an automated analysis platform to parallelize the high-throughput data-generation platform. Currently, to a large extent, FCM data analysis relies on the manual selection of sequential regions in 2-D graphical projections to extract the cell populations of interest. This is a time-consuming task that ignores the high-dimensionality of FCM data. Results In view of the aforementioned issues, we have developed an R package called flowClust to automate FCM analysis. flowClust implements a robust model-based clustering approach based on multivariate t mixture models with the Box-Cox transformation. The package provides the functionality to identify cell populations whilst simultaneously handling the commonly encountered issues of outlier identification and data transformation. It offers various tools to summarize and visualize a wealth of features of the clustering results. In addition, to ensure its convenience of use, flowClust has been adapted for the current FCM data format, and integrated with existing Bioconductor packages dedicated to FCM analysis. Conclusion flowClust addresses the issue of a dearth of software that helps automate FCM analysis with a sound theoretical foundation. It tends to give reproducible results, and helps reduce the significant subjectivity and human time cost encountered in FCM analysis. The package contributes to the cytometry community by offering an efficient, automated analysis platform which facilitates the active, ongoing technological advancement.

  12. Identification of contact and respiratory sensitizers using flow cytometry

    International Nuclear Information System (INIS)

    Goutet, Michele; Pepin, Elsa; Langonne, Isabelle; Huguet, Nelly; Ban, Masarin

    2005-01-01

    Identification of the chemicals responsible for respiratory and contact allergies in the industrial area is an important occupational safety issue. This study was conducted in mice to determine whether flow cytometry is an appropriate method to analyze and differentiate the specific immune responses to the respiratory sensitizer trimellitic anhydride (TMA) and to the contact sensitizer dinitrochlorobenzene (DNCB) used at concentrations with comparable immunogenic potential. Mice were exposed twice on the flanks (days 0, 5) to 10% TMA or 1% DNCB and challenged three times on the ears (days 10, 11, 12) with 2.5% TMA or 0.25% DNCB. Flow cytometry analyses were conducted on draining lymph node cells harvested on days 13 and 18. Comparing TMA and DNCB immune responses on day 13, we found obvious differences that persisted for most of them on day 18. An increased proportion of IgE+ cells correlated to total serum IgE level and an enhancement of MHC II molecule expression were observed in the lymph node B lymphocytes from TMA-treated mice. The percentage of IL-4-producing CD4+ lymphocytes and the IL-4 receptor expression were clearly higher following TMA exposure. In contrast, higher proportions of IL-2-producing cells were detected in CD4+ and CD8+ cells from DNCB-treated mice. Both chemicals induced a significant increase in the percentage of IFN-γ-producing cells among CD8+ lymphocytes but to a greater proportion following TMA treatment. In conclusion, this study encourages the use of flow cytometry to discriminate between contact and respiratory sensitizers by identifying divergent expression of immune response parameters

  13. Optimizing transformations for automated, high throughput analysis of flow cytometry data.

    Science.gov (United States)

    Finak, Greg; Perez, Juan-Manuel; Weng, Andrew; Gottardo, Raphael

    2010-11-04

    In a high throughput setting, effective flow cytometry data analysis depends heavily on proper data preprocessing. While usual preprocessing steps of quality assessment, outlier removal, normalization, and gating have received considerable scrutiny from the community, the influence of data transformation on the output of high throughput analysis has been largely overlooked. Flow cytometry measurements can vary over several orders of magnitude, cell populations can have variances that depend on their mean fluorescence intensities, and may exhibit heavily-skewed distributions. Consequently, the choice of data transformation can influence the output of automated gating. An appropriate data transformation aids in data visualization and gating of cell populations across the range of data. Experience shows that the choice of transformation is data specific. Our goal here is to compare the performance of different transformations applied to flow cytometry data in the context of automated gating in a high throughput, fully automated setting. We examine the most common transformations used in flow cytometry, including the generalized hyperbolic arcsine, biexponential, linlog, and generalized Box-Cox, all within the BioConductor flowCore framework that is widely used in high throughput, automated flow cytometry data analysis. All of these transformations have adjustable parameters whose effects upon the data are non-intuitive for most users. By making some modelling assumptions about the transformed data, we develop maximum likelihood criteria to optimize parameter choice for these different transformations. We compare the performance of parameter-optimized and default-parameter (in flowCore) data transformations on real and simulated data by measuring the variation in the locations of cell populations across samples, discovered via automated gating in both the scatter and fluorescence channels. We find that parameter-optimized transformations improve visualization, reduce

  14. Optimizing transformations for automated, high throughput analysis of flow cytometry data

    Directory of Open Access Journals (Sweden)

    Weng Andrew

    2010-11-01

    Full Text Available Abstract Background In a high throughput setting, effective flow cytometry data analysis depends heavily on proper data preprocessing. While usual preprocessing steps of quality assessment, outlier removal, normalization, and gating have received considerable scrutiny from the community, the influence of data transformation on the output of high throughput analysis has been largely overlooked. Flow cytometry measurements can vary over several orders of magnitude, cell populations can have variances that depend on their mean fluorescence intensities, and may exhibit heavily-skewed distributions. Consequently, the choice of data transformation can influence the output of automated gating. An appropriate data transformation aids in data visualization and gating of cell populations across the range of data. Experience shows that the choice of transformation is data specific. Our goal here is to compare the performance of different transformations applied to flow cytometry data in the context of automated gating in a high throughput, fully automated setting. We examine the most common transformations used in flow cytometry, including the generalized hyperbolic arcsine, biexponential, linlog, and generalized Box-Cox, all within the BioConductor flowCore framework that is widely used in high throughput, automated flow cytometry data analysis. All of these transformations have adjustable parameters whose effects upon the data are non-intuitive for most users. By making some modelling assumptions about the transformed data, we develop maximum likelihood criteria to optimize parameter choice for these different transformations. Results We compare the performance of parameter-optimized and default-parameter (in flowCore data transformations on real and simulated data by measuring the variation in the locations of cell populations across samples, discovered via automated gating in both the scatter and fluorescence channels. We find that parameter

  15. Absolute counting of neutrophils in whole blood using flow cytometry.

    Science.gov (United States)

    Brunck, Marion E G; Andersen, Stacey B; Timmins, Nicholas E; Osborne, Geoffrey W; Nielsen, Lars K

    2014-12-01

    Absolute neutrophil count (ANC) is used clinically to monitor physiological dysfunctions such as myelosuppression or infection. In the research laboratory, ANC is a valuable measure to monitor the evolution of a wide range of disease states in disease models. Flow cytometry (FCM) is a fast, widely used approach to confidently identify thousands of cells within minutes. FCM can be optimised for absolute counting using spiked-in beads or by measuring the sample volume analysed. Here we combine the 1A8 antibody, specific for the mouse granulocyte protein Ly6G, with flow cytometric counting in straightforward FCM assays for mouse ANC, easily implementable in the research laboratory. Volumetric and Trucount™ bead assays were optimized for mouse neutrophils, and ANC values obtained with these protocols were compared to ANC measured by a dual-platform assay using the Orphee Mythic 18 veterinary haematology analyser. The single platform assays were more precise with decreased intra-assay variability compared with ANC obtained using the dual protocol. Defining ANC based on Ly6G expression produces a 15% higher estimate than the dual protocol. Allowing for this difference in ANC definition, the flow cytometry counting assays using Ly6G can be used reliably in the research laboratory to quantify mouse ANC from a small volume of blood. We demonstrate the utility of the volumetric protocol in a time-course study of chemotherapy induced neutropenia using four drug regimens. © 2014 International Society for Advancement of Cytometry.

  16. DNA-Cytometry of Progressive and Regressive Cervical Intraepithelial Neoplasia

    Directory of Open Access Journals (Sweden)

    Antonius G. J. M. Hanselaar

    1998-01-01

    Full Text Available A retrospective analysis was performed on archival cervical smears from a group of 56 women with cervical intraepithelial neoplasia (CIN, who had received follow‐up by cytology only. Automated image cytometry of Feulgen‐stained DNA was used to determine the differences between progressive and regressive lesions. The first group of 30 smears was from women who had developed cancer after initial smears with dysplastic changes (progressive group. The second group of 26 smears with dysplastic changes had shown regression to normal (regressive group. The goal of the study was to determine if differences in cytometric features existed between the progressive and regressive groups. CIN categories I, II and III were represented in both groups, and measurements were pooled across diagnostic categories. Images of up to 700 intermediate cells were obtained from each slide, and cells were scanned exhaustively for the detection of diagnostic cells. Discriminant function analysis was performed for both intermediate and diagnostic cells. The most significant differences between the groups were found for diagnostic cells, with a cell classification accuracy of 82%. Intermediate cells could be classified with 60% accuracy. Cytometric features which afforded the best discrimination were characteristic of the chromatin organization in diagnostic cells (nuclear texture. Slide classification was performed by thresholding the number of cells which exhibited progression associated changes (PAC in chromatin configuration, with an accuracy of 93 and 73% for diagnostic and intermediate cells, respectively. These results indicate that regardless of the extent of nuclear atypia as reflected in the CIN category, features of chromatin organization can potentially be used to predict the malignant or progressive potential of CIN lesions.

  17. Multiplex polymerase chain reaction on FTA cards vs. flow cytometry for B-lymphocyte clonality.

    Science.gov (United States)

    Dictor, Michael; Skogvall, Ingela; Warenholt, Janina; Rambech, Eva

    2007-01-01

    Two-colour flow cytometry was compared with multiplex PCR with capillary electrophoresis for clonality determination in specific categories of B-cell lymphoma. FTA cards were evaluated for preserving DNA from node imprints and expediting molecular analysis. A single-tube multiplex PCR targeted IGH and lymphoma-specific translocations in DNA extracted from 180 frozen lymphoid tissues and DNA bound to FTA cards from 192 fresh tissues and 137 aspirates. PCR results were compared with flow cytometry in the extracted and aspirated samples. Overall, single-tube multiplex PCR sensitivity was equivalent in the sample groups (intergroup range 79%-91%). False negatives were associated with tumour origin in the follicle centre. Multiplex PCR and flow cytometry were equally sensitive and together detected 98% of B-cell lymphomas. Additional two-tube targeting of IGK suggested an overall molecular sensitivity >90%. False positive (pseudoclonal) single-tube multiplex PCR was associated with necrosis and sparse lymphocytes. Multiplex PCR using template DNA bound to an FTA card effectively detects B-lymphocyte clonality, obviates DNA extraction and refrigeration, and can be used without diminished sensitivity in fine needle aspirates or node imprints as a replacement for or complement to flow cytometry at any point in the diagnostic work-up.

  18. A model of selective visual attention for a stereo pair of images

    Science.gov (United States)

    Park, Min Chul; Kim, Sung Kyu; Son, Jung-Young

    2005-11-01

    Human visual attention system has a remarkable ability to interpret complex scenes with the ease and simplicity by selecting or focusing on a small region of visual field without scanning the whole images. In this paper, a novel selective visual attention model by using 3D image display system for a stereo pair of images is proposed. It is based on the feature integration theory and locates ROI(region of interest) or FOA(focus of attention). The disparity map obtained from a stereo pair of images is exploited as one of spatial visual features to form a set of topographic feature maps in our approach. Though the true human cognitive mechanism on the analysis and integration process might be different from our assumption the proposed attention system matches well with the results found by human observers.

  19. A pilot study for the integration of cytometry reports in digital cytology telemedicine applications.

    Science.gov (United States)

    Giansanti, Daniele; Cerroni, Fabio; Amodeo, Rachele; Filoni, Marco; Giovagnoli, Maria Rosaria

    2010-01-01

    Up to date, tele-pathology in the three different forms of application, "dynamic", "static" and "virtual microscopy" has been mainly based on tele-hystology remote consulting. Today the diffusion of specialized WAN connections is guiding the research of new applications of tele-pathology. A specific analysis has been conducted, focused on digital cytology, in the biomedical laboratory of Sant'Andrea Hospital to investigate the technologies potentially useful to integrate in the LAN/WAN for telemedicine applications. Among the possible tools useful to be integrated in the LAN/WAN for telemedicine applications, the cytometry equipment available in the technical unity of cytometry has been considered important. The study finally provides a proposal for a tele-consulting architecture for the integration of cytometry reports both in the hospital LAN and the WAN for possible cooperative diagnosis and second opinion support.

  20. A pilot study for the integration of cytometry reports in digital cytology telemedicine applications

    Directory of Open Access Journals (Sweden)

    Daniele Giansanti

    2010-06-01

    Full Text Available Up to date, tele-pathology in the three different forms of application, "dynamic", "static" and "virtual microscopy" has been mainly based on tele-hystology remote consulting. Today the diffusion of specialized WAN connections is guiding the research of new applications of tele-pathology. A specific analysis has been conducted, focused on digital cytology, in the biomedical laboratory of Sant'Andrea Hospital to investigate the technologies potentially useful to integrate in the LAN/WAN for telemedicine applications. Among the possible tools useful to be integrated in the LAN/WAN for telemedicine applications, the cytometry equipment available in the technical unity of cytometry has been considered important. The study finally provides a proposal for a tele-consulting architecture for the integration of cytometry reports both in the hospital LAN and the WAN for possible cooperative diagnosis and second opinion support.

  1. Emotional textile image classification based on cross-domain convolutional sparse autoencoders with feature selection

    Science.gov (United States)

    Li, Zuhe; Fan, Yangyu; Liu, Weihua; Yu, Zeqi; Wang, Fengqin

    2017-01-01

    We aim to apply sparse autoencoder-based unsupervised feature learning to emotional semantic analysis for textile images. To tackle the problem of limited training data, we present a cross-domain feature learning scheme for emotional textile image classification using convolutional autoencoders. We further propose a correlation-analysis-based feature selection method for the weights learned by sparse autoencoders to reduce the number of features extracted from large size images. First, we randomly collect image patches on an unlabeled image dataset in the source domain and learn local features with a sparse autoencoder. We then conduct feature selection according to the correlation between different weight vectors corresponding to the autoencoder's hidden units. We finally adopt a convolutional neural network including a pooling layer to obtain global feature activations of textile images in the target domain and send these global feature vectors into logistic regression models for emotional image classification. The cross-domain unsupervised feature learning method achieves 65% to 78% average accuracy in the cross-validation experiments corresponding to eight emotional categories and performs better than conventional methods. Feature selection can reduce the computational cost of global feature extraction by about 50% while improving classification performance.

  2. Pre-processing, registration and selection of adaptive optics corrected retinal images.

    Science.gov (United States)

    Ramaswamy, Gomathy; Devaney, Nicholas

    2013-07-01

    In this paper, the aim is to demonstrate enhanced processing of sequences of fundus images obtained using a commercial AO flood illumination system. The purpose of the work is to (1) correct for uneven illumination at the retina (2) automatically select the best quality images and (3) precisely register the best images. Adaptive optics corrected retinal images are pre-processed to correct uneven illumination using different methods; subtracting or dividing by the average filtered image, homomorphic filtering and a wavelet based approach. These images are evaluated to measure the image quality using various parameters, including sharpness, variance, power spectrum kurtosis and contrast. We have carried out the registration in two stages; a coarse stage using cross-correlation followed by fine registration using two approaches; parabolic interpolation on the peak of the cross-correlation and maximum-likelihood estimation. The angle of rotation of the images is measured using a combination of peak tracking and Procrustes transformation. We have found that a wavelet approach (Daubechies 4 wavelet at 6th level decomposition) provides good illumination correction with clear improvement in image sharpness and contrast. The assessment of image quality using a 'Designer metric' works well when compared to visual evaluation, although it is highly correlated with other metrics. In image registration, sub-pixel translation measured using parabolic interpolation on the peak of the cross-correlation function and maximum-likelihood estimation are found to give very similar results (RMS difference 0.047 pixels). We have confirmed that correcting rotation of the images provides a significant improvement, especially at the edges of the image. We observed that selecting the better quality frames (e.g. best 75% images) for image registration gives improved resolution, at the expense of poorer signal-to-noise. The sharpness map of the registered and de-rotated images shows increased

  3. Determination of total bacterial count in raw milk by flow cytometry

    Directory of Open Access Journals (Sweden)

    Dubravka Samaržija

    2004-01-01

    Full Text Available The automatic flow cytometry as routine method for total bacterial count determination of raw ex-farm milk has recently been accepted in Croatia. This method significantly differs from the reference method (Standard Plate Count mostly in the presentation of the results obtained. Therefore, this paper summarized experiences in the application of flow cytometry in the dairy laboratories practice. The principle and the practice of the method, methodological details and factors influencing the results were described. In order to avoid problems regarding the interpretation of the results, which aregeneral problems of the quantitative microbiology, this article try to explain an appropriate conversion of the results with regards to SPC/ml, as an official method for the bacteriological quality proposal by the national legislation.

  4. Installation of spectrally selective imaging system in RF negative ion source

    International Nuclear Information System (INIS)

    Ikeda, K.; Kisaki, M.; Nagaoka, K.; Nakano, H.; Osakabe, M.; Tsumori, K.; Kaneko, O.; Takeiri, Y.; Wünderlich, D.; Fantz, U.; Heinemann, B.; Geng, S.

    2016-01-01

    A spectrally selective imaging system has been installed in the RF negative ion source in the International Thermonuclear Experimental Reactor-relevant negative ion beam test facility ELISE (Extraction from a Large Ion Source Experiment) to investigate distribution of hydrogen Balmer-α emission (H α ) close to the production surface of hydrogen negative ion. We selected a GigE vision camera coupled with an optical band-path filter, which can be controlled remotely using high speed network connection. A distribution of H α emission near the bias plate has been clearly observed. The same time trend on H α intensities measured by the imaging diagnostic and the optical emission spectroscopy is confirmed

  5. Cytometry of mammalian sperm

    Energy Technology Data Exchange (ETDEWEB)

    Gledhill, B.L.

    1983-10-11

    Male germ cells respond dramatically to a variety of insults and are important reproductive dosimeters. Semen analyses are very useful in studies on the effects of drugs, chemicals, and environmental hazards on testicular function, male fertility and heritable germinal mutations. The accessibility of male cells makes them well suited for analytical cytology. We might automate the process of determining sperm morphology but should not do so solely for increased speed. Rather, richer tangible benefits will derive from cytometric evaluation through increased sensitivity, reduced subjectivity, standardization between investigators and laboratories, enhanced archival systems, and the benefits of easily exchanged standardized data. Inroads on the standardization of assays for motility and functional integrity are being made. Flow cytometric analysis of total DNA content of individual sperm is an insensitive means to detect exposure to reproductive toxins because of the small size and low frequency of the DNA content errors. Flow cytometry can be applied to determine the proportions of X- and Y-sperm in semen samples.

  6. Cytometry of mammalian sperm

    International Nuclear Information System (INIS)

    Gledhill, B.L.

    1983-01-01

    Male germ cells respond dramatically to a variety of insults and are important reproductive dosimeters. Semen analyses are very useful in studies on the effects of drugs, chemicals, and environmental hazards on testicular function, male fertility and heritable germinal mutations. The accessibility of male cells makes them well suited for analytical cytology. We might automate the process of determining sperm morphology but should not do so solely for increased speed. Rather, richer tangible benefits will derive from cytometric evaluation through increased sensitivity, reduced subjectivity, standardization between investigators and laboratories, enhanced archival systems, and the benefits of easily exchanged standardized data. Inroads on the standardization of assays for motility and functional integrity are being made. Flow cytometric analysis of total DNA content of individual sperm is an insensitive means to detect exposure to reproductive toxins because of the small size and low frequency of the DNA content errors. Flow cytometry can be applied to determine the proportions of X- and Y-sperm in semen samples

  7. Mobile phone based imaging system for selected tele-healthcare applications

    OpenAIRE

    Condominas Guàrdia, Jordi

    2013-01-01

    A mobile phone based telemedicine study is developed to see how feasible phone usage is in selected health care applications. The research is divided into three different objectives. The first objective is to compile the technical characteristics of selected mobile phones from telemedicine perspective. The second objective is to develop techniques to acquire quality images of skin with mobile phones. Finally a smartphone based telemedicine application will be developed to asses...

  8. Biology and flow cytometry of proangiogenic hematopoietic progenitors cells.

    Science.gov (United States)

    Rose, Jonathan A; Erzurum, Serpil; Asosingh, Kewal

    2015-01-01

    During development, hematopoiesis and neovascularization are closely linked to each other via a common bipotent stem cell called the hemangioblast that gives rise to both hematopoietic cells and endothelial cells. In postnatal life, this functional connection between the vasculature and hematopoiesis is maintained by a subset of hematopoietic progenitor cells endowed with the capacity to differentiate into potent proangiogenic cells. These proangiogenic hematopoietic progenitors comprise a specific subset of bone marrow (BM)-derived cells that homes to sites of neovascularization and possess potent paracrine angiogenic activity. There is emerging evidence that this subpopulation of hematopoietic progenitors plays a critical role in vascular health and disease. Their angiogenic activity is distinct from putative "endothelial progenitor cells" that become structural cells of the endothelium by differentiation into endothelial cells. Proangiogenic hematopoietic progenitor cell research requires multidisciplinary expertise in flow cytometry, hematology, and vascular biology. This review provides a comprehensive overview of proangiogenic hematopoietic progenitor cell biology and flow cytometric methods to detect these cells in the peripheral blood circulation and BM. © 2014 International Society for Advancement of Cytometry.

  9. Lipid nanoparticles assessment by flow cytometry.

    Science.gov (United States)

    Bryła, Anna; Juzwa, Wojciech; Weiss, Marek; Lewandowicz, Grażyna

    2017-03-30

    Liposomes are promising carriers for drugs and bioactive compounds. Size and structure are their crucial parameters. Thus, it is essential to assess individual vesicles as prepared. Currently available techniques fail to measure liposome's size and structure simultaneously, with a high throughput. To solve this problem, we have developed a novel, flow cytometric method quantifying liposomes. Firstly, the following fluorescent staining combinations were tested: DiD/TO, Rh123/DiD, Syto9/DiD. Further, chosen fluorochromes were used to compare three populations of vesicles: raw (R), obtained by thin film hydration and extruded ones (populations E10 and E21). Dynamic light scattering (DLS) was used for determination of average diameter and size distribution of nanocarriers. Structural differences between the raw and the extruded liposomes, as well as additional information concerning vesicles size were acquired employing atomic force microscopy (AFM). DLS analysis indicated that, three distinct populations of vesicles were obtained. Liposomes were characterized by mean diameter of 323nm, 220nm and 170nm for population R, E10 and E21 respectively. All the populations were stable and revealed zeta potential of -29mV. AFM confirmed that raw and extruded liposomes were differed in structure. DiD/TO was the optimal fluorochrome combination that enabled to resolve distinctly the sub-populations of liposomes. Results obtained by flow cytometry were in a good agreement with those from DLS and AFM. It was proved that, flow cytometry, when proper fluorescent dyes are used, is an adequate method for liposomes assessment. The proposed method enables fast and reliable analysis of liposomes in their native environment. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Hyperexpansion of wheat chromosomes sorted by flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Endo, Takashi R.; Kubaláková, Marie; Vrána, Jan; Doležel, Jaroslav

    2014-01-01

    Roč. 89, č. 4 (2014), s. 181-185 ISSN 1341-7568 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : flow cytometry * flow sorting * chromosome Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.930, year: 2014 http://gateway.isiknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcAuth=Alerting&SrcApp=Alerting&DestApp=MEDLINE&DestLinkType=FullRecord&UT=25747042

  11. 3D material cytometry (3DMaC): a very high-replicate, high-throughput analytical method using microfabricated, shape-specific, cell-material niches.

    Science.gov (United States)

    Parratt, Kirsten; Jeong, Jenny; Qiu, Peng; Roy, Krishnendu

    2017-08-08

    Studying cell behavior within 3D material niches is key to understanding cell biology in health and diseases, and developing biomaterials for regenerative medicine applications. Current approaches to studying these cell-material niches have low throughput and can only analyze a few replicates per experiment resulting in reduced measurement assurance and analytical power. Here, we report 3D material cytometry (3DMaC), a novel high-throughput method based on microfabricated, shape-specific 3D cell-material niches and imaging cytometry. 3DMaC achieves rapid and highly multiplexed analyses of very high replicate numbers ("n" of 10 4 -10 6 ) of 3D biomaterial constructs. 3DMaC overcomes current limitations of low "n", low-throughput, and "noisy" assays, to provide rapid and simultaneous analyses of potentially hundreds of parameters in 3D biomaterial cultures. The method is demonstrated here for a set of 85 000 events containing twelve distinct cell-biomaterial micro-niches along with robust, customized computational methods for high-throughput analytics with potentially unprecedented statistical power.

  12. Flow cytometry with gold nanoparticlesand their clusters as scattering contrast agents: FDTD simulation of light-cell interaction

    DEFF Research Database (Denmark)

    Tanev, Stoyan; Sun, Wenbo; Pond, James

    2009-01-01

    refractive index matching conditions and by cells labeled by gold nanoparticles. The optical schematics including phase contrast (OPCM) microscopy as a prospective modality for in vivo flow cytometry is also analyzed. The validation of the FDTD approach for the simulation of flow cytometry may open a new...

  13. Organizing the Cellular and Molecular Heterogeneity in High Grade Serous Ovarian Cancer by Mass Cytometry

    Science.gov (United States)

    2015-10-01

    about more informed changes to treatment modalities. To accomplish this vision with HG-SOC, we are using a single cell technology , mass cytometry... extracted from the composite MST. Clusters are represented as bubbles, the size of which corresponds to the number of cells in the cluster. The level of...Fantl WJ, Nolan GP. Transient partial permeabilization with saponin enables cellular barcoding prior to surface marker staining. Cytometry A. 2014 Dec;85

  14. Flow Cytometry Method as a Diagnostic Tool for Pleural Fluid Involvement in a Patient with Multiple Myeloma

    Directory of Open Access Journals (Sweden)

    Muzaffer Keklik

    2012-10-01

    Full Text Available Multiple myeloma is a malignant proliferation of plasma cells that mainly affects bone marrow. Pleural effusions secondary to pleural myelomatous involvement have rarely been reported in the literature. As it is rarely detected, we aimed to report a case in which pleural effusion of a multiple myeloma was confirmed as true myelomatous involvement by flow cytometry method. A 52-years old man presented to our clinic with chest and back pain lasting for 3 months. On the chest radiography, pleural fluid was detected in left hemithorax. Pleural fluid flow cytometry was performed. In the flow cytometry, CD56, CD38 and CD138 found to be positive, while CD19 was negative. True myelomatous pleural effusions are very uncommon, with fewer than 100 cases reported worldwide. Flow cytometry is a potentially useful diagnostic tool for clinical practice. We presented our case; as it has been rarely reported, although flow cytometer is a simple method for detection of pleural fluid involvement in multiple myeloma.

  15. Using image processing technology and mathematical algorithm in the automatic selection of vocal cord opening and closing images from the larynx endoscopy video.

    Science.gov (United States)

    Kuo, Chung-Feng Jeffrey; Chu, Yueng-Hsiang; Wang, Po-Chun; Lai, Chun-Yu; Chu, Wen-Lin; Leu, Yi-Shing; Wang, Hsing-Won

    2013-12-01

    The human larynx is an important organ for voice production and respiratory mechanisms. The vocal cord is approximated for voice production and open for breathing. The videolaryngoscope is widely used for vocal cord examination. At present, physicians usually diagnose vocal cord diseases by manually selecting the image of the vocal cord opening to the largest extent (abduction), thus maximally exposing the vocal cord lesion. On the other hand, the severity of diseases such as vocal palsy, atrophic vocal cord is largely dependent on the vocal cord closing to the smallest extent (adduction). Therefore, diseases can be assessed by the image of the vocal cord opening to the largest extent, and the seriousness of breathy voice is closely correlated to the gap between vocal cords when closing to the smallest extent. The aim of the study was to design an automatic vocal cord image selection system to improve the conventional selection process by physicians and enhance diagnosis efficiency. Also, due to the unwanted fuzzy images resulting from examination process caused by human factors as well as the non-vocal cord images, texture analysis is added in this study to measure image entropy to establish a screening and elimination system to effectively enhance the accuracy of selecting the image of the vocal cord closing to the smallest extent. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  16. Flow cytometry-assisted rapid isolation of recombinant Plasmodium berghei parasites exemplified by functional analysis of aquaglyceroporin

    Science.gov (United States)

    Kenthirapalan, Sanketha; Waters, Andrew P.; Matuschewski, Kai; Kooij, Taco W.A.

    2012-01-01

    The most critical bottleneck in the generation of recombinant Plasmodium berghei parasites is the mandatory in vivo cloning step following successful genetic manipulation. This study describes a new technique for rapid selection of recombinant P. berghei parasites. The method is based on flow cytometry to isolate isogenic parasite lines and represents a major advance for the field, in that it will speed the generation of recombinant parasites as well as cut down on animal use significantly. High expression of GFP during blood infection, a prerequisite for robust separation of transgenic lines by flow cytometry, was achieved. Isogenic recombinant parasite populations were isolated even in the presence of a 100-fold excess of wild-type (WT) parasites. Aquaglyceroporin (AQP) loss-of-function mutants and parasites expressing a tagged AQP were generated to validate this approach. aqp− parasites grow normally within the WT phenotypic range during blood infection of NMRI mice. Similarly, colonization of the insect vector and establishment of an infection after mosquito transmission were unaffected, indicating that AQP is dispensable for life cycle progression in vivo under physiological conditions, refuting its use as a suitable drug target. Tagged AQP localized to perinuclear structures and not the parasite plasma membrane. We suggest that flow-cytometric isolation of isogenic parasites overcomes the major roadblock towards a genome-scale repository of mutant and transgenic malaria parasite lines. PMID:23137753

  17. Integration of lyoplate based flow cytometry and computational analysis for standardized immunological biomarker discovery.

    Directory of Open Access Journals (Sweden)

    Federica Villanova

    Full Text Available Discovery of novel immune biomarkers for monitoring of disease prognosis and response to therapy in immune-mediated inflammatory diseases is an important unmet clinical need. Here, we establish a novel framework for immunological biomarker discovery, comparing a conventional (liquid flow cytometry platform (CFP and a unique lyoplate-based flow cytometry platform (LFP in combination with advanced computational data analysis. We demonstrate that LFP had higher sensitivity compared to CFP, with increased detection of cytokines (IFN-γ and IL-10 and activation markers (Foxp3 and CD25. Fluorescent intensity of cells stained with lyophilized antibodies was increased compared to cells stained with liquid antibodies. LFP, using a plate loader, allowed medium-throughput processing of samples with comparable intra- and inter-assay variability between platforms. Automated computational analysis identified novel immunophenotypes that were not detected with manual analysis. Our results establish a new flow cytometry platform for standardized and rapid immunological biomarker discovery with wide application to immune-mediated diseases.

  18. Integration of lyoplate based flow cytometry and computational analysis for standardized immunological biomarker discovery.

    Science.gov (United States)

    Villanova, Federica; Di Meglio, Paola; Inokuma, Margaret; Aghaeepour, Nima; Perucha, Esperanza; Mollon, Jennifer; Nomura, Laurel; Hernandez-Fuentes, Maria; Cope, Andrew; Prevost, A Toby; Heck, Susanne; Maino, Vernon; Lord, Graham; Brinkman, Ryan R; Nestle, Frank O

    2013-01-01

    Discovery of novel immune biomarkers for monitoring of disease prognosis and response to therapy in immune-mediated inflammatory diseases is an important unmet clinical need. Here, we establish a novel framework for immunological biomarker discovery, comparing a conventional (liquid) flow cytometry platform (CFP) and a unique lyoplate-based flow cytometry platform (LFP) in combination with advanced computational data analysis. We demonstrate that LFP had higher sensitivity compared to CFP, with increased detection of cytokines (IFN-γ and IL-10) and activation markers (Foxp3 and CD25). Fluorescent intensity of cells stained with lyophilized antibodies was increased compared to cells stained with liquid antibodies. LFP, using a plate loader, allowed medium-throughput processing of samples with comparable intra- and inter-assay variability between platforms. Automated computational analysis identified novel immunophenotypes that were not detected with manual analysis. Our results establish a new flow cytometry platform for standardized and rapid immunological biomarker discovery with wide application to immune-mediated diseases.

  19. Hybrid Imaging Labels: Providing the Link Between Mass Spectrometry-Based Molecular Pathology and Theranostics

    Science.gov (United States)

    Buckle, Tessa; van der Wal, Steffen; van Malderen, Stijn J.M.; Müller, Larissa; Kuil, Joeri; van Unen, Vincent; Peters, Ruud J.B.; van Bemmel, Margaretha E.M.; McDonnell, Liam A.; Velders, Aldrik H.; Koning, Frits; Vanhaeke, Frank; van Leeuwen, Fijs W. B.

    2017-01-01

    Background: Development of theranostic concepts that include inductively coupled plasma mass spectrometry (ICP-MS) and laser ablation ICP-MS (LA-ICP-MS) imaging can be hindered by the lack of a direct comparison to more standardly used methods for in vitro and in vivo evaluation; e.g. fluorescence or nuclear medicine. In this study a bimodal (or rather, hybrid) tracer that contains both a fluorescent dye and a chelate was used to evaluate the existence of a direct link between mass spectrometry (MS) and in vitro and in vivo molecular imaging findings using fluorescence and radioisotopes. At the same time, the hybrid label was used to determine whether the use of a single isotope label would allow for MS-based diagnostics. Methods: A hybrid label that contained both a DTPA chelate (that was coordinated with either 165Ho or 111In) and a Cy5 fluorescent dye was coupled to the chemokine receptor 4 (CXCR4) targeting peptide Ac-TZ14011 (hybrid-Cy5-Ac-TZ4011). This receptor targeting tracer was used to 1) validate the efficacy of (165Ho-based) mass-cytometry in determining the receptor affinity via comparison with fluorescence-based flow cytometry (Cy5), 2) evaluate the microscopic binding pattern of the tracer in tumor cells using both fluorescence confocal imaging (Cy5) and LA-ICP-MS-imaging (165Ho), 3) compare in vivo biodistribution patterns obtained with ICP-MS (165Ho) and radiodetection (111In) after intravenous administration of hybrid-Cy5-Ac-TZ4011 in tumor-bearing mice. Finally, LA-ICP-MS-imaging (165Ho) was linked to fluorescence-based analysis of excised tissue samples (Cy5). Results: Analysis with both mass-cytometry and flow cytometry revealed a similar receptor affinity, respectively 352 ± 141 nM and 245 ± 65 nM (p = 0.08), but with a much lower detection sensitivity for the first modality. In vitro LA-ICP-MS imaging (165Ho) enabled clear discrimination between CXCR4 positive and negative cells, but fluorescence microscopy was required to determine the

  20. Interfacing Lab-on-a-Chip Embryo Technology with High-Definition Imaging Cytometry.

    Science.gov (United States)

    Zhu, Feng; Hall, Christopher J; Crosier, Philip S; Wlodkowic, Donald

    2015-08-01

    To spearhead deployment of zebrafish embryo biotests in large-scale drug discovery studies, automated platforms are needed to integrate embryo in-test positioning and immobilization (suitable for high-content imaging) with fluidic modules for continuous drug and medium delivery under microperfusion to developing embryos. In this work, we present an innovative design of a high-throughput three-dimensional (3D) microfluidic chip-based device for automated immobilization and culture and time-lapse imaging of developing zebrafish embryos under continuous microperfusion. The 3D Lab-on-a-Chip array was fabricated in poly(methyl methacrylate) (PMMA) transparent thermoplastic using infrared laser micromachining, while the off-chip interfaces were fabricated using additive manufacturing processes (fused deposition modelling and stereolithography). The system's design facilitated rapid loading and immobilization of a large number of embryos in predefined clusters of traps during continuous microperfusion of drugs/toxins. It was conceptually designed to seamlessly interface with both upright and inverted fluorescent imaging systems and also to directly interface with conventional microtiter plate readers that accept 96-well plates. Compared with the conventional Petri dish assays, the chip-based bioassay was much more convenient and efficient as only small amounts of drug solutions were required for the whole perfusion system running continuously over 72 h. Embryos were spatially separated in the traps that assisted tracing single embryos, preventing interembryo contamination and improving imaging accessibility.

  1. Stochastic Individual-Based Modeling of Bacterial Growth and Division Using Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Míriam R. García

    2018-01-01

    Full Text Available A realistic description of the variability in bacterial growth and division is critical to produce reliable predictions of safety risks along the food chain. Individual-based modeling of bacteria provides the theoretical framework to deal with this variability, but it requires information about the individual behavior of bacteria inside populations. In this work, we overcome this problem by estimating the individual behavior of bacteria from population statistics obtained with flow cytometry. For this objective, a stochastic individual-based modeling framework is defined based on standard assumptions during division and exponential growth. The unknown single-cell parameters required for running the individual-based modeling simulations, such as cell size growth rate, are estimated from the flow cytometry data. Instead of using directly the individual-based model, we make use of a modified Fokker-Plank equation. This only equation simulates the population statistics in function of the unknown single-cell parameters. We test the validity of the approach by modeling the growth and division of Pediococcus acidilactici within the exponential phase. Estimations reveal the statistics of cell growth and division using only data from flow cytometry at a given time. From the relationship between the mother and daughter volumes, we also predict that P. acidilactici divide into two successive parallel planes.

  2. Flow Cytometry in Diagnosis of Myelomatous Pleural Effusion: A Case Report.

    Science.gov (United States)

    Arora, Parul; Gupta, Sanjeev Kumar; Mallik, Nabhajit; Mittal, Reena; Sharma, Om Dutt; Kumar, Lalit

    2016-06-01

    Plasma cell myeloma is a multifocal plasma cell neoplasm associated with increased monoclonal protein in serum and/or urine. Pleural effusions in patients with myeloma are uncommon (6 %). However, effusions due to direct infiltration of the pleura by plasma cells (myelomatous pleural effusion) are extremely rare (pleural fluid cytology, electrophoresis or pleural biopsy. We present a case of myelomatous pleural effusion diagnosed using flow cytometry immunophenotyping in addition to the pleural fluid cytology. A 45 year old female was diagnosed as plasma cell myeloma (IgG kappa) in 2007. She received multiple lines of therapy during the course of her treatment including thalidomide, dexamethasone, lenalidomide, bortezomib, and doxorubicin based regimens. However, the patient had progressive extramedullary disease and developed pleural effusion in 2014. Cytological examination of the pleural fluid showed degenerative changes. Few preserved areas showed mononuclear cells including morphologically abnormal plasma cells. Immunophenotyping of these cells by flow cytometry revealed a pattern indicating neoplastic plasma cells. There was expression of CD38, CD138, and CD56, with absence of CD19, CD10 and CD45. This confirmed the diagnosis of myelomatous pleural effusion. Subsequently, the patient was offered a dexamethasone, cyclophosphamide, etoposide and cisplatin based regimen but, she declined further treatment and succumbed to her disease 3 months later. Myelomatous pleural effusion is a rare complication of plasma cell myeloma. Flow cytometry can be used as an adjunctive technique in its diagnosis particularly in cases with equivocal cytology and electrophoresis findings.

  3. Rough-fuzzy clustering and unsupervised feature selection for wavelet based MR image segmentation.

    Directory of Open Access Journals (Sweden)

    Pradipta Maji

    Full Text Available Image segmentation is an indispensable process in the visualization of human tissues, particularly during clinical analysis of brain magnetic resonance (MR images. For many human experts, manual segmentation is a difficult and time consuming task, which makes an automated brain MR image segmentation method desirable. In this regard, this paper presents a new segmentation method for brain MR images, integrating judiciously the merits of rough-fuzzy computing and multiresolution image analysis technique. The proposed method assumes that the major brain tissues, namely, gray matter, white matter, and cerebrospinal fluid from the MR images are considered to have different textural properties. The dyadic wavelet analysis is used to extract the scale-space feature vector for each pixel, while the rough-fuzzy clustering is used to address the uncertainty problem of brain MR image segmentation. An unsupervised feature selection method is introduced, based on maximum relevance-maximum significance criterion, to select relevant and significant textural features for segmentation problem, while the mathematical morphology based skull stripping preprocessing step is proposed to remove the non-cerebral tissues like skull. The performance of the proposed method, along with a comparison with related approaches, is demonstrated on a set of synthetic and real brain MR images using standard validity indices.

  4. Gram-typing of mastitis bacteria in milk samples using flow cytometry

    DEFF Research Database (Denmark)

    Langerhuus, Sine Nygaard; Ingvartsen, Klaus Lønne; Bennedsgaard, Torben Werner

    2013-01-01

    Fast identification of pathogenic bacteria in milk samples from cows with clinical mastitis is central to proper treatment. In Denmark, time to bacterial diagnosis is typically 24 to 48 h when using traditional culturing methods. The PCR technique provides a faster and highly sensitive identifica......Fast identification of pathogenic bacteria in milk samples from cows with clinical mastitis is central to proper treatment. In Denmark, time to bacterial diagnosis is typically 24 to 48 h when using traditional culturing methods. The PCR technique provides a faster and highly sensitive...... cytometry-based method, which can detect and distinguish gram-negative and gram-positive bacteria in mastitis milk samples. The differentiation was based on bacterial fluorescence intensities upon labeling with biotin-conjugated wheat germ agglutinin and acridine orange. Initially 19 in-house bacterial...... characteristic curves for the 19 bacterial cultures. The method was then tested on 53 selected mastitis cases obtained from the department biobank (milk samples from 6 gram-negative and 47 gram-positive mastitis cases). Gram-negative bacteria in milk samples were detected with a sensitivity of 1...

  5. Mouse monoclonal antibodies against human c-Mpl and characterization for flow cytometry applications.

    Science.gov (United States)

    Abbott, Christina; Huang, Guo; Ellison, Aaron R; Chen, Ching; Arora, Taruna; Szilvassy, Stephen J; Wei, Ping

    2010-04-01

    Mouse monoclonal antibodies (MAbs) against human c-Mpl, the cognate receptor for thrombopoietin (TPO), were generated using hybridoma technology and characterized by various assays to demonstrate their specificity and affinity. Two such MAbs, 1.6 and 1.75, were determined to be superior for flow cytometry studies and exhibited double-digit picomolar (pM) affinities to soluble human c-Mpl protein. Both MAbs specifically bound to cells engineered to overexpress human c-Mpl protein, immortalized human hematopoietic cell lines that express endogenous c-Mpl, primary human bone marrow and peripheral blood-derived CD34(+) cells, and purified human platelets. No binding was detected on cell lines that did not express c-Mpl. Receptor competition and siRNA knock-down studies further confirmed the specificity of antibodies 1.6 and 1.75 for human c-Mpl. In contrast to these newly generated MAbs, none of eight commercially available anti-c-Mpl antibodies tested were found to bind specifically to human c-Mpl and were thus shown to be unsuitable for flow cytometry studies. Monoclonal antibodies 1.6 and 1.75 will therefore be useful flow cytometry reagents to detect cell surface c-Mpl expression.

  6. Metal-Containing Polystyrene Beads as Standards for Mass Cytometry.

    Science.gov (United States)

    Abdelrahman, Ahmed I; Ornatsky, Olga; Bandura, Dmitry; Baranov, Vladimir; Kinach, Robert; Dai, Sheng; Thickett, Stuart C; Tanner, Scott; Winnik, Mitchell A

    2010-01-01

    We examine the suitability of metal-containing polystyrene beads for the calibration of a mass cytometer instrument, a single particle analyser based on an inductively coupled plasma ion source and a time of flight mass spectrometer. These metal-containing beads are also verified for their use as internal standards for this instrument. These beads were synthesized by multiple-stage dispersion polymerization with acrylic acid as a comonomer. Acrylic acid acts as a ligand to anchor the metal ions within the interior of the beads. Mass cytometry enabled the bead-by-bead measurement of the metal-content and determination of the metal-content distribution. Beads synthesized by dispersion polymerization that involved three stages were shown to have narrower bead-to-bead variation in their lanthanide content than beads synthesized by 2-stage dispersion polymerization. The beads exhibited insignificant release of their lanthanide content to aqueous solutions of different pHs over a period of six months. When mixed with KG1a or U937 cell lines, metal-containing polymer beads were shown not to affect the mass cytometry response to the metal content of element-tagged antibodies specifically attached to these cells.

  7. Routine detection of Epstein-Barr virus specific T-cells in the peripheral blood by flow cytometry

    Science.gov (United States)

    Crucian, B. E.; Stowe, R. P.; Pierson, D. L.; Sams, C. F.

    2001-01-01

    The ability to detect cytomegalovirus-specific T-cells (CD4(+)) in the peripheral blood by flow cytometry has been recently described by Picker et al. In this method, cells are incubated with viral antigen and responding (cytokine producing) T-cells are then identified by flow cytometry. To date, this technique has not been reliably used to detect Epstein-Barr virus (EBV)-specific T-cells primarily due to the superantigen/mitogenic properties of the virus which non-specifically activate T-cells. By modifying culture conditions under which the antigens are presented, we have overcome this limitation and developed an assay to detect and quantitate EBV-specific T-cells. The detection of cytokine producing T-cells by flow cytometry requires an extremely strong signal (such as culture in the presence of PMA and ionomycin). Our data indicate that in modified culture conditions (early removal of viral antigen) the non-specific activation of T-cells by EBV is reduced, but antigen presentation will continue uninhibited. Using this method, EBV-specific T-cells may be legitimately detected using flow cytometry. No reduction in the numbers of antigen-specific T-cells was observed by the early removal of target antigen when verified using cytomegalovirus antigen (a virus with no non-specific T-cell activation properties). In EBV-seropositive individuals, the phenotype of the EBV-specific cytokine producing T-cells was evaluated using four-color flow cytometry and found to be CD45(+), CD3(+), CD4(+), CD45RA(-), CD69(+), CD25(-). This phenotype indicates the stimulation of circulating previously unactivated memory T-cells. No cytokine production was observed in CD4(+) T-cells from EBV-seronegative individuals, confirming the specificity of this assay. In addition, the use of four color cytometry (CD45, CD3, CD69, IFNgamma/IL-2) allows the total quantitative assessment of EBV-specific T-cells while monitoring the interference of EBV non-specific mitogenic activity. This method may

  8. [Hyperspectral remote sensing image classification based on SVM optimized by clonal selection].

    Science.gov (United States)

    Liu, Qing-Jie; Jing, Lin-Hai; Wang, Meng-Fei; Lin, Qi-Zhong

    2013-03-01

    Model selection for support vector machine (SVM) involving kernel and the margin parameter values selection is usually time-consuming, impacts training efficiency of SVM model and final classification accuracies of SVM hyperspectral remote sensing image classifier greatly. Firstly, based on combinatorial optimization theory and cross-validation method, artificial immune clonal selection algorithm is introduced to the optimal selection of SVM (CSSVM) kernel parameter a and margin parameter C to improve the training efficiency of SVM model. Then an experiment of classifying AVIRIS in India Pine site of USA was performed for testing the novel CSSVM, as well as a traditional SVM classifier with general Grid Searching cross-validation method (GSSVM) for comparison. And then, evaluation indexes including SVM model training time, classification overall accuracy (OA) and Kappa index of both CSSVM and GSSVM were all analyzed quantitatively. It is demonstrated that OA of CSSVM on test samples and whole image are 85.1% and 81.58, the differences from that of GSSVM are both within 0.08% respectively; And Kappa indexes reach 0.8213 and 0.7728, the differences from that of GSSVM are both within 0.001; While the ratio of model training time of CSSVM and GSSVM is between 1/6 and 1/10. Therefore, CSSVM is fast and accurate algorithm for hyperspectral image classification and is superior to GSSVM.

  9. Sensitive and selective tumor imaging with novel and highly activatable fluorescence probes

    International Nuclear Information System (INIS)

    Urano, Yasuteru

    2008-01-01

    Selective and sensitive tumor imaging in vivo is one of the most requested methodologies in medical sciences. Although several imaging modalities have been developed including positron emission tomography (PET) and magnetic resonance (MR) imaging for the detection of tumors, none of these modalities can activate the signals upon being accumulated or uptaken to tumor sites. Among these modalities, only optical fluorescence imaging has a marked advantage, that is, their signals can be dramatically increased upon detecting some biological features. In this short review, I will introduce some recent strategies for activatable optical fluorescence imaging of tumors, and discuss their advantages over other modalities. (author)

  10. A new and fast image feature selection method for developing an optimal mammographic mass detection scheme.

    Science.gov (United States)

    Tan, Maxine; Pu, Jiantao; Zheng, Bin

    2014-08-01

    Selecting optimal features from a large image feature pool remains a major challenge in developing computer-aided detection (CAD) schemes of medical images. The objective of this study is to investigate a new approach to significantly improve efficacy of image feature selection and classifier optimization in developing a CAD scheme of mammographic masses. An image dataset including 1600 regions of interest (ROIs) in which 800 are positive (depicting malignant masses) and 800 are negative (depicting CAD-generated false positive regions) was used in this study. After segmentation of each suspicious lesion by a multilayer topographic region growth algorithm, 271 features were computed in different feature categories including shape, texture, contrast, isodensity, spiculation, local topological features, as well as the features related to the presence and location of fat and calcifications. Besides computing features from the original images, the authors also computed new texture features from the dilated lesion segments. In order to select optimal features from this initial feature pool and build a highly performing classifier, the authors examined and compared four feature selection methods to optimize an artificial neural network (ANN) based classifier, namely: (1) Phased Searching with NEAT in a Time-Scaled Framework, (2) A sequential floating forward selection (SFFS) method, (3) A genetic algorithm (GA), and (4) A sequential forward selection (SFS) method. Performances of the four approaches were assessed using a tenfold cross validation method. Among these four methods, SFFS has highest efficacy, which takes 3%-5% of computational time as compared to GA approach, and yields the highest performance level with the area under a receiver operating characteristic curve (AUC) = 0.864 ± 0.034. The results also demonstrated that except using GA, including the new texture features computed from the dilated mass segments improved the AUC results of the ANNs optimized

  11. Use of Multicolor Flow Cytometry for Isolation of Specific Cell Populations Deriving from Differentiated Human Embryonic Stem Cells

    NARCIS (Netherlands)

    Mengarelli, Isabella; Fryga, Andrew; Barberi, Tiziano

    2016-01-01

    Flow Cytometry-Sorting (FCM-Sorting) is a technique commonly used to identify and isolate specific types of cells from a heterogeneous population of live cells. Here we describe a multicolor flow cytometry technique that uses five distinct cell surface antigens to isolate four live populations with

  12. Applications of flow cytometry to toxicological mycotoxin effects in cultured mammalian cells: a review.

    Science.gov (United States)

    Juan-García, Ana; Manyes, Lara; Ruiz, María-José; Font, Guillermina

    2013-06-01

    This review gives an overview of flow cytometry applications to toxicological studies of several physiological target sites of mycotoxins on different mammalian cell lines. Mycotoxins are secondary metabolites of fungi that may be present in food, feed, air and water. The increasing presence of mycotoxins in crops, their wide distribution in the food chain, and their potential for toxicity demonstrate the need for further knowledge. Flow cytometry has become a valuable tool in mycotoxin studies in recent years for the rapid analysis of single cells in a mixture. In toxicology, the power of these methods lies in the possibility of determining a wide range of cell parameters, providing valuable information to elucidate cell growth and viability, metabolic activity, mitochondrial membrane potential and membrane integrity mechanisms. There are studies using flow cytometry technique on Alternaria, Aspergillus, Fusarium and Penicillium mycotoxins including information about cell type, assay conditions and functional parameters. Most of the studies collected in the literature are on deoxynivalenol and zearalenone mycotoxins. Cell cycle analysis and apoptosis are the processes more widely investigated. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Detection of an Abnormal Myeloid Clone by Flow Cytometry in Familial Platelet Disorder With Propensity to Myeloid Malignancy.

    Science.gov (United States)

    Ok, Chi Young; Leventaki, Vasiliki; Wang, Sa A; Dinardo, Courtney; Medeiros, L Jeffrey; Konoplev, Sergej

    2016-02-01

    To report aberrant myeloblasts detected by flow cytometry immunophenotypic studies in an asymptomatic patient with familial platelet disorder with propensity to myeloid malignancy, a rare autosomal dominant disease caused by germline heterozygous mutations in Runt-related transcription factor 1. Morphologic evaluation, flow cytometry immunophenotypic studies, nanofluidics-based qualitative multiplex reverse transcriptase polymerase chain reaction, Sanger sequencing, and next-generation sequencing-based mutational hotspot analysis of 53 genes were performed on bone marrow biopsy and aspirate samples. Flow cytometry immunophenotypic analysis showed 0.6% CD34+ blasts with an abnormal immunophenotype: CD13 increased, CD33+, CD38 decreased, CD117 increased, and CD123 increased. The acquisition of new phenotypic aberrancies in myeloblasts as detected by flow cytometry immunophenotypic studies might be a harbinger of impending myelodysplastic syndrome or acute myeloid leukemia in a patient with familial platelet disorder with propensity to myeloid malignancy. © American Society for Clinical Pathology, 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. The use of flow cytometry to monitor chitin synthesis in regenerating protoplasts of Candida albicans.

    Science.gov (United States)

    Hector, R F; Braun, P C; Hart, J T; Kamarck, M E

    1990-01-01

    Flow cytometry was used to monitor chitin synthesis in regenerating protoplasts of the yeast Candida albicans. Comparisons of cells stained with Calcofluor White, a fluorochrome with known affinity for chitin, and cells incubated in the presence of N-[3H]-acetylglucosamine, the precursor substrate for chitin, showed a linear relationship between fluorescence and incorporation of label over time. Changes in both the fluorescence and light scatter of regenerating protoplasts treated with inhibitors of fungal chitin synthase were also quantitated by flow cytometry.

  15. Analysis of immunophenotype in acute myeloid leukemia by multiparameter flow cytometry

    International Nuclear Information System (INIS)

    Gao Yanqun; Jin Haijie; Yan Pei; Wang Feifei; Li Xiaohong; Gao Chunji

    2005-01-01

    To evaluate the immunophenotype of acute leukemia patients, the surface and cytoplasmic antigen expression in 162 cases of acute leukemia were analyzed by multiparameter flow cytometry and CD45/SSC gating. The results showed that CDl17 (94.9%), CD13 (88.5%) and CD33(70.5%) were mainly expressed in ANLL patients; cCD79a(100%), CD19(92.1%) were chiefly expressed in B-ALL patients, and in T-ALL patients, cCD3(100%) and CD2(83.3%) were expressed; For the expression of lymphoid differentiation antigen Ly+ANLL, CD7 (56.2%) and CD19(31.2%) were chiefly found, and for myeloid antigen My+ALL, CD13(88. 9%) and CD33 (27.8%) were detected. In conclusion, multiparameter flow cytometry and three-color direct immunofluorescence staining methods may be of important clinical significance in diagnosis, therapy and prognosis of acute leukemia. (authors)

  16. Effective delivery of hydrophobic drugs to breast (MCF-7) and Liver (HepG2) cancer cells: A detailed investigation using Cytotoxicity assays, fluorescence imaging and flow cytometry.

    Science.gov (United States)

    Manatunga, Danushika C; de Silva, Rohini M; Nalin de Silva, K M; Neelika Malavige, Gathsaurie; Wijeratne, Dulharie T; Williams, Gareth R; Jayasinghe, Chanika D; Udagama, Preethi V

    2018-04-03

    This study aimed to develop a drug carrier system consisting of a polymer containing hydroxyapatite (HAp) shell and a magnetic core of iron oxide nanoparticles. Doxorubicin and/or curcumin were loaded into the carrier via a simple diffusion deposition approach, with encapsulation efficiencies (EE) for curcumin and doxorubicin of 93.03 ± 0.3% and 97.37 ± 0.12% respectively. The co-loading of curcumin and doxorubicin led to a total EE of 76.02 ± 0.48%. Release studies were carried out at pH 7.4 and 5.3, and revealed higher release was at pH 5.3 expressing the potential application in tumor microenvironments. Cytotoxicity assays, fluorescence imaging and flow cytometry showed the formulations could effectively inhibit the growth of MCF-7 and HEpG2 cancer cells, being more potent than the free drug molecules both in dose and time dependent manner. Additionally, hemolysis tests and cytotoxicity evaluations determined the drug-loaded carriers to be non-toxic towards non-cancerous cells. These formulations thus have great potential in the development of new cancer therapeutics. Copyright © 2018. Published by Elsevier B.V.

  17. European canine lymphoma network consensus recommendations for reporting flow cytometry in canine hematopoietic neoplasms.

    Science.gov (United States)

    Comazzi, S; Avery, P R; Garden, O A; Riondato, F; Rütgen, B; Vernau, W

    2017-09-01

    Flow cytometry (FC) is assuming increasing importance in diagnosis in veterinary oncology. The European Canine Lymphoma Network (ECLN) is an international cooperation of different institutions working on canine lymphoma diagnosis and therapy. The ECLN panel of experts on FC has defined the issue of reporting FC on canine lymphoma and leukemia as their first hot topic, since a standardized report that includes all the important information is still lacking in veterinary medicine. The flow cytometry panel of the ECLN started a consensus initiative using the Delphi approach. Clinicians were considered the main target of FC reports. A panel of experts in FC was interrogated about the important information needed from a report. Using the feedback from clinicians and subsequent discussion, a list of information to be included in the report was made, with four different levels of recommendation. The final report should include both a quantitative part and a qualitative or descriptive part with interpretation of the salient results. Other items discussed included the necessity of reporting data regarding the quality of samples, use of absolute numbers of positive cells, cutoff values, the intensity of fluorescence, and possible aberrant patterns of antigen expression useful from a clinical point of view. The consensus initiative is a first step toward standardization of diagnostic approach to canine hematopoietic neoplasms among different institutions and countries. This harmonization will improve communication and patient care and also facilitate the multicenter studies necessary to further our knowledge of canine hematopoietic neoplasms. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

  18. Radon-induced DNA damage and apoptosis analyzed by flow cytometry

    International Nuclear Information System (INIS)

    Meenakshi, C.; Mohankumar, Mary N.

    2012-01-01

    Natural radiation is the major source of human exposure to ionizing radiation and its largest contributing component to effective doses arises from inhalation of 222 Rn and its radioactive progeny. 222 Rn, a chemically inert gas produced naturally from radium in rocks and soil is a proven source of lung cancer especially in closed environments such as mines and in poorly ventilated homes. Much of the data on the effect of radon in humans comes from epidemiological studies, often masked by confounding factors such as age, smoking and lifestyle. Radiation carcinogenesis is initiated by DNA damage and flow cytometry is a versatile, fast and accurate technique for the analysis of DNA damage as it offers the analysis of high number of individual cells in few minutes. An attempt was made to detect DNA damage and apoptosis after exposing human blood cells in vitro to radon by flow cytometry. Blood samples were collected from apparently healthy individuals and exposed in vitro to radon ranging between 1-5 mGy using a simple, portable irradiation assembly designed and tested at the Radiological Safety Division of Indira Gandhi Centre for Atomic Research. Cultures were initiated by the addition of phytohemagglutinin and cells were processed stained and analyzed for DNA damage and apoptosis by flow cytometry. CV values indicative of DNA damage were plotted against dose and were observed to increase in a dose dependent manner 3h after of irradiation. However no such response was observed at 24h and 48h. Nevertheless, the percentage of apoptotic cells increased steadily with dose after 24 and 48h post exposure. DNA breaks appear to be rejoined after about 24h of irradiation. However apoptotic cells increased with time and dose, suggesting elimination of highly damaged cells. Further experiments are needed to identify apoptotic cells as a biomarker of radiation exposure and risk. (author)

  19. Stochastic Measurement Models for Quantifying Lymphocyte Responses Using Flow Cytometry

    Science.gov (United States)

    Kan, Andrey; Pavlyshyn, Damian; Markham, John F.; Dowling, Mark R.; Heinzel, Susanne; Zhou, Jie H. S.; Marchingo, Julia M.; Hodgkin, Philip D.

    2016-01-01

    Adaptive immune responses are complex dynamic processes whereby B and T cells undergo division and differentiation triggered by pathogenic stimuli. Deregulation of the response can lead to severe consequences for the host organism ranging from immune deficiencies to autoimmunity. Tracking cell division and differentiation by flow cytometry using fluorescent probes is a major method for measuring progression of lymphocyte responses, both in vitro and in vivo. In turn, mathematical modeling of cell numbers derived from such measurements has led to significant biological discoveries, and plays an increasingly important role in lymphocyte research. Fitting an appropriate parameterized model to such data is the goal of these studies but significant challenges are presented by the variability in measurements. This variation results from the sum of experimental noise and intrinsic probabilistic differences in cells and is difficult to characterize analytically. Current model fitting methods adopt different simplifying assumptions to describe the distribution of such measurements and these assumptions have not been tested directly. To help inform the choice and application of appropriate methods of model fitting to such data we studied the errors associated with flow cytometry measurements from a wide variety of experiments. We found that the mean and variance of the noise were related by a power law with an exponent between 1.3 and 1.8 for different datasets. This violated the assumptions inherent to commonly used least squares, linear variance scaling and log-transformation based methods. As a result of these findings we propose a new measurement model that we justify both theoretically, from the maximum entropy standpoint, and empirically using collected data. Our evaluation suggests that the new model can be reliably used for model fitting across a variety of conditions. Our work provides a foundation for modeling measurements in flow cytometry experiments thus

  20. Biomass measurement by flow cytometry during solid-state fermentation of basidiomycetes.

    Science.gov (United States)

    Steudler, Susanne; Böhmer, Ulrike; Weber, Jost; Bley, Thomas

    2015-02-01

    Solid-state fermentation (SSF) is a robust process that is well suited to the on-site cultivation of basidiomycetes that produce enzymes for the treatment of lignocellulosics. Reliable methods for biomass quantification are essential for the analysis of fungal growth kinetics. However, direct biomass determination is not possible during SSF because the fungi grow into the substrate and use it as a nutrient source. This necessitates the use of indirect methods that are either very laborious and time consuming or can only provide biomass measurements during certain growth periods. Here, we describe the development and optimization of a new rapid method for fungal biomass determination during SSF that is based on counting fungal nuclei by flow cytometry. Fungal biomass was grown on an organic substrate and its concentration was measured by isolating the nuclei from the fungal hyphae after cell disruption, staining them with SYTOX(®) Green, and then counting them using a flow cytometer. A calibration curve relating the dry biomass of the samples to their concentrations of nuclei was established. Multiple buffers and disruption methods were tested. The results obtained were compared with values determined using the method of ergosterol determination, a classical technique for fungal biomass measurement during SSF. Our new approach can be used to measure fungal biomass on a range of different scales, from 15 mL cultures to a laboratory reactor with a working volume of 10 L (developed by the Research Center for Medical Technology and Biotechnology (fzmb GmbH)). © 2014 International Society for Advancement of Cytometry. © 2014 International Society for Advancement of Cytometry.

  1. A general method for bead-enhanced quantitation by flow cytometry

    Science.gov (United States)

    Montes, Martin; Jaensson, Elin A.; Orozco, Aaron F.; Lewis, Dorothy E.; Corry, David B.

    2009-01-01

    Flow cytometry provides accurate relative cellular quantitation (percent abundance) of cells from diverse samples, but technical limitations of most flow cytometers preclude accurate absolute quantitation. Several quantitation standards are now commercially available which, when added to samples, permit absolute quantitation of CD4+ T cells. However, these reagents are limited by their cost, technical complexity, requirement for additional software and/or limited applicability. Moreover, few studies have validated the use of such reagents in complex biological samples, especially for quantitation of non-T cells. Here we show that addition to samples of known quantities of polystyrene fluorescence standardization beads permits accurate quantitation of CD4+ T cells from complex cell samples. This procedure, here termed single bead-enhanced cytofluorimetry (SBEC), was equally capable of enumerating eosinophils as well as subcellular fragments of apoptotic cells, moieties with very different optical and fluorescent characteristics. Relative to other proprietary products, SBEC is simple, inexpensive and requires no special software, suggesting that the method is suitable for the routine quantitation of most cells and other particles by flow cytometry. PMID:17067632

  2. Radiation-induced apoptosis in thymocytes as determined by flow cytometry

    International Nuclear Information System (INIS)

    Su Xu; Zhang Yingchun; Liu Shuzheng

    1995-01-01

    Programmed cell death (PCD), or apoptosis, is a conceptually different way of cell death from necrosis. PCD plays an important role in immunologic regulation, PCD in thymocytes was analyzed by flow cytometry following in vitro X-irradiation. It was found that culturing of thymocytes could induce PCD which showed a time dependent increase. Four hours after culturing, 16% of thymocytes was found in the Ao region (PCD is shown in the Ao region of the histogram of flow cytometry). PCD in thymocytes showed a time dependent increase after 2.0 Gy X-irradiation, being significantly higher than that in the control at the same culturing time. 24 hours after X-irradiation in vitro, it was found that with doses below 100 mGy PCD was not significantly different from the control at the same culturing time. But when the doses were above 100 mGy, PCD showed a dose dependent increase, being significantly higher than that of the control at the same culturing time. These results are important in the understanding of the biological effects of low dose radiation

  3. Research Techniques Made Simple: Experimental Methodology for Single-Cell Mass Cytometry

    NARCIS (Netherlands)

    Matos, Tiago R.; Liu, Hongye; Ritz, Jerome

    2017-01-01

    Growing recognition of the complexity of interactions within cellular systems has fueled the development of mass cytometry. The precision of time-of-flight mass spectrometry combined with the labeling of specific ligands with mass tags enables detection and quantification of more than 40 markers at

  4. Immune Response to Mycobacterial Infection: Lessons from Flow Cytometry

    OpenAIRE

    Rovina, Nikoletta; Panagiotou, Marios; Pontikis, Konstantinos; Kyriakopoulou, Magdalini; Koulouris, Nikolaos G.; Koutsoukou, Antonia

    2013-01-01

    Detecting and treating active and latent tuberculosis are pivotal elements for effective infection control; yet, due to their significant inherent limitations, the diagnostic means for these two stages of tuberculosis (TB) to date remain suboptimal. This paper reviews the current diagnostic tools for mycobacterial infection and focuses on the application of flow cytometry as a promising method for rapid and reliable diagnosis of mycobacterial infection as well as discrimination between active...

  5. Stage-specific activity of potential antimalarial compounds measured in vitro by flow cytometry in comparison to optical microscopy and hypoxanthine uptake

    Directory of Open Access Journals (Sweden)

    Carmen E Contreras

    2004-03-01

    Full Text Available The evaluation of new antimalarial agents using older methods of monitoring sensitivity to antimalarial drugs are laborious and poorly suited to discriminate stage-specific activity. We used flow cytometry to study the effect of established antimalarial compounds, cysteine protease inhibitors, and a quinolone against asexual stages of Plasmodium falciparum. Cultured P. falciparum parasites were treated for 48 h with different drug concentrations and the parasitemia was determined by flow cytometry methods after DNA staining with propidium iodide. P. falciparum erythrocytic life cycle stages were readily distinguished by flow cytometry. Activities of established and new antimalarial compounds measured by flow cytometry were equivalent to results obtained with microscopy and metabolite uptake assays. The antimalarial activity of all compounds was higher against P. falciparum trophozoite stages. Advantages of flow cytometry analysis over traditional assays included higher throughput for data collection, insight into the stage-specificity of antimalarial activity avoiding use of radioactive isotopes.

  6. Preparation of rat islet B-cell-enriched fractions by light-scatter flow cytometry

    International Nuclear Information System (INIS)

    Rabinovitch, A.; Russell, T.; Shienvold, F.; Noel, J.; Files, N.; Patel, Y.; Ingram, M.

    1982-01-01

    Flow cytometry has been examined as a method to separate islet cells into homogeneous subpopulations. Collagenase-isolated rat islets were dissociated into single cells and these were analyzed and sorted according to their low forward angle light scattering properties by using automated flow cytometry. Light scatter histograms showed two peaks of viable cells. Radioimmunoassay of hormone content in cell fractions collected across the the two peaks showed that glucagon-containing cells were concentrated towards the left side of the left peak and somatostatin-containing cells were concentrated towards the right side of the left peak, whereas insulin-containing cells were clearly enriched in the right peak. The B-cell-enriched fraction (90% B cells, 3% A cells, 2% D cells) exhibited significant insulin secretory responses to glucose (16.7 mM), and 3-isobutyl-1-methylxanthine (0.1 mM), during a 24-h culture period, and these responses were slightly greater than those observed in the original mixed islet cell preparation (66% B cells, 14% A cells, and 4% D cells). These results indicate that flow cytometry can be applied to sort pancreatic islet cells into populations enriched in specific endocrine cell types for further study of the functions of individual cell types

  7. SU-C-209-05: Monte Carlo Model of a Prototype Backscatter X-Ray (BSX) Imager for Projective and Selective Object-Plane Imaging

    International Nuclear Information System (INIS)

    Rolison, L; Samant, S; Baciak, J; Jordan, K

    2016-01-01

    Purpose: To develop a Monte Carlo N-Particle (MCNP) model for the validation of a prototype backscatter x-ray (BSX) imager, and optimization of BSX technology for medical applications, including selective object-plane imaging. Methods: BSX is an emerging technology that represents an alternative to conventional computed tomography (CT) and projective digital radiography (DR). It employs detectors located on the same side as the incident x-ray source, making use of backscatter and avoiding ring geometry to enclose the imaging object. Current BSX imagers suffer from low spatial resolution. A MCNP model was designed to replicate a BSX prototype used for flaw detection in industrial materials. This prototype consisted of a 1.5mm diameter 60kVp pencil beam surrounded by a ring of four 5.0cm diameter NaI scintillation detectors. The imaging phantom consisted of a 2.9cm thick aluminum plate with five 0.6cm diameter holes drilled halfway. The experimental image was created using a raster scanning motion (in 1.5mm increments). Results: A qualitative comparison between the physical and simulated images showed very good agreement with 1.5mm spatial resolution in plane perpendicular to incident x-ray beam. The MCNP model developed the concept of radiography by selective plane detection (RSPD) for BSX, whereby specific object planes can be imaged by varying kVp. 10keV increments in mean x-ray energy yielded 4mm thick slice resolution in the phantom. Image resolution in the MCNP model can be further increased by increasing the number of detectors, and decreasing raster step size. Conclusion: MCNP modelling was used to validate a prototype BSX imager and introduce the RSPD concept, allowing for selective object-plane imaging. There was very good visual agreement between the experimental and MCNP imaging. Beyond optimizing system parameters for the existing prototype, new geometries can be investigated for volumetric image acquisition in medical applications. This material is

  8. SU-C-209-05: Monte Carlo Model of a Prototype Backscatter X-Ray (BSX) Imager for Projective and Selective Object-Plane Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Rolison, L; Samant, S; Baciak, J; Jordan, K [University of Florida, Gainesville, FL (United States)

    2016-06-15

    Purpose: To develop a Monte Carlo N-Particle (MCNP) model for the validation of a prototype backscatter x-ray (BSX) imager, and optimization of BSX technology for medical applications, including selective object-plane imaging. Methods: BSX is an emerging technology that represents an alternative to conventional computed tomography (CT) and projective digital radiography (DR). It employs detectors located on the same side as the incident x-ray source, making use of backscatter and avoiding ring geometry to enclose the imaging object. Current BSX imagers suffer from low spatial resolution. A MCNP model was designed to replicate a BSX prototype used for flaw detection in industrial materials. This prototype consisted of a 1.5mm diameter 60kVp pencil beam surrounded by a ring of four 5.0cm diameter NaI scintillation detectors. The imaging phantom consisted of a 2.9cm thick aluminum plate with five 0.6cm diameter holes drilled halfway. The experimental image was created using a raster scanning motion (in 1.5mm increments). Results: A qualitative comparison between the physical and simulated images showed very good agreement with 1.5mm spatial resolution in plane perpendicular to incident x-ray beam. The MCNP model developed the concept of radiography by selective plane detection (RSPD) for BSX, whereby specific object planes can be imaged by varying kVp. 10keV increments in mean x-ray energy yielded 4mm thick slice resolution in the phantom. Image resolution in the MCNP model can be further increased by increasing the number of detectors, and decreasing raster step size. Conclusion: MCNP modelling was used to validate a prototype BSX imager and introduce the RSPD concept, allowing for selective object-plane imaging. There was very good visual agreement between the experimental and MCNP imaging. Beyond optimizing system parameters for the existing prototype, new geometries can be investigated for volumetric image acquisition in medical applications. This material is

  9. Frequency Selective Non-Linear Blending to Improve Image Quality in Liver CT.

    Science.gov (United States)

    Bongers, M N; Bier, G; Kloth, C; Schabel, C; Fritz, J; Nikolaou, K; Horger, M

    2016-12-01

    Purpose: To evaluate the effects of a new frequency selective non-linear blending (NLB) algorithm on the contrast resolution of liver CT with low intravascular concentration of iodine contrast. Materials and Methods: Our local ethics committee approved this retrospective study. The informed consent requirement was waived. CT exams of 25 patients (60 % female, mean age: 65 ± 16 years of age) with late phase CT scans of the liver were included as a model for poor intrahepatic vascular contrast enhancement. Optimal post-processing settings to enhance the contrast of hepatic vessels were determined. Outcome variables included signal-to-noise (SNR) and contrast-to-noise ratios (CNR) of hepatic vessels and SNR of liver parenchyma of standard and post-processed images. Image quality was quantified by two independent readers using Likert scales. Results: The post-processing settings for the visualization of hepatic vasculature were optimal at a center of 115HU, delta of 25HU, and slope of 5. Image noise was statistically indifferent between standard and post-processed images. The CNR between the hepatic vasculature (HV) and liver parenchyma could be significantly increased for liver veins (CNR Standard 1.62 ± 1.10, CNR NLB 3.6 ± 2.94, p = 0.0002) and portal veins (CNR Standard 1.31 ± 0.85, CNR NLB 2.42 ± 3.03, p = 0.046). The SNR of liver parenchyma was significantly higher on post-processed images (SNR NLB 11.26 ± 3.16, SNR Standard 8.85 ± 2.27, p = 0.008). The overall image quality and depiction of HV were significantly higher on post-processed images (NLB DHV : 4 [3 - 4.75], S tandardDHV : 2 [1.3 - 2.5], p = algorithm increases the contrast resolution of liver CT and can improve the visibility of the hepatic vasculature in the setting of a low contrast ratio between vessels and the parenchyma. Key Points: • Using the new frequency selective non-linear blending algorithm is feasible in contrast

  10. Using neutrosophic graph cut segmentation algorithm for qualified rendering image selection in thyroid elastography video.

    Science.gov (United States)

    Guo, Yanhui; Jiang, Shuang-Quan; Sun, Baiqing; Siuly, Siuly; Şengür, Abdulkadir; Tian, Jia-Wei

    2017-12-01

    Recently, elastography has become very popular in clinical investigation for thyroid cancer detection and diagnosis. In elastogram, the stress results of the thyroid are displayed using pseudo colors. Due to variation of the rendering results in different frames, it is difficult for radiologists to manually select the qualified frame image quickly and efficiently. The purpose of this study is to find the qualified rendering result in the thyroid elastogram. This paper employs an efficient thyroid ultrasound image segmentation algorithm based on neutrosophic graph cut to find the qualified rendering images. Firstly, a thyroid ultrasound image is mapped into neutrosophic set, and an indeterminacy filter is constructed to reduce the indeterminacy of the spatial and intensity information in the image. A graph is defined on the image and the weight for each pixel is represented using the value after indeterminacy filtering. The segmentation results are obtained using a maximum-flow algorithm on the graph. Then the anatomic structure is identified in thyroid ultrasound image. Finally the rendering colors on these anatomic regions are extracted and validated to find the frames which satisfy the selection criteria. To test the performance of the proposed method, a thyroid elastogram dataset is built and totally 33 cases were collected. An experienced radiologist manually evaluates the selection results of the proposed method. Experimental results demonstrate that the proposed method finds the qualified rendering frame with 100% accuracy. The proposed scheme assists the radiologists to diagnose the thyroid diseases using the qualified rendering images.

  11. UNLABELED SELECTED SAMPLES IN FEATURE EXTRACTION FOR CLASSIFICATION OF HYPERSPECTRAL IMAGES WITH LIMITED TRAINING SAMPLES

    Directory of Open Access Journals (Sweden)

    A. Kianisarkaleh

    2015-12-01

    Full Text Available Feature extraction plays a key role in hyperspectral images classification. Using unlabeled samples, often unlimitedly available, unsupervised and semisupervised feature extraction methods show better performance when limited number of training samples exists. This paper illustrates the importance of selecting appropriate unlabeled samples that used in feature extraction methods. Also proposes a new method for unlabeled samples selection using spectral and spatial information. The proposed method has four parts including: PCA, prior classification, posterior classification and sample selection. As hyperspectral image passes these parts, selected unlabeled samples can be used in arbitrary feature extraction methods. The effectiveness of the proposed unlabeled selected samples in unsupervised and semisupervised feature extraction is demonstrated using two real hyperspectral datasets. Results show that through selecting appropriate unlabeled samples, the proposed method can improve the performance of feature extraction methods and increase classification accuracy.

  12. Flow cytometry with gold nanoparticles and their clusters as scattering contrast agents: FDTD simulation of light-cell interaction.

    Science.gov (United States)

    Tanev, Stoyan; Sun, Wenbo; Pond, James; Tuchin, Valery V; Zharov, Vladimir P

    2009-09-01

    The formulation of the finite-difference time-domain (FDTD) approach is presented in the framework of its potential applications to in-vivo flow cytometry based on light scattering. The consideration is focused on comparison of light scattering by a single biological cell alone in controlled refractive-index matching conditions and by cells labeled by gold nanoparticles. The optical schematics including phase contrast (OPCM) microscopy as a prospective modality for in-vivo flow cytometry is also analyzed. The validation of the FDTD approach for the simulation of flow cytometry may open up a new avenue in the development of advanced cytometric techniques based on scattering effects from nanoscale targets. 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

  13. Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trapping

    Science.gov (United States)

    Labib, Mahmoud; Mohamadi, Reza M.; Poudineh, Mahla; Ahmed, Sharif U.; Ivanov, Ivaylo; Huang, Ching-Lung; Moosavi, Maral; Sargent, Edward H.; Kelley, Shana O.

    2018-05-01

    Cell-to-cell variation in gene expression creates a need for techniques that can characterize expression at the level of individual cells. This is particularly true for rare circulating tumour cells, in which subtyping and drug resistance are of intense interest. Here we describe a method for cell analysis—single-cell mRNA cytometry—that enables the isolation of rare cells from whole blood as a function of target mRNA sequences. This approach uses two classes of magnetic particles that are labelled to selectively hybridize with different regions of the target mRNA. Hybridization leads to the formation of large magnetic clusters that remain localized within the cells of interest, thereby enabling the cells to be magnetically separated. Targeting specific intracellular mRNAs enablescirculating tumour cells to be distinguished from normal haematopoietic cells. No polymerase chain reaction amplification is required to determine RNA expression levels and genotype at the single-cell level, and minimal cell manipulation is required. To demonstrate this approach we use single-cell mRNA cytometry to detect clinically important sequences in prostate cancer specimens.

  14. National flow cytometry and sorting research resource. Annual progress report, July, 1, 1994--June 30, 1995, Year 12

    Energy Technology Data Exchange (ETDEWEB)

    Jett, J.H.

    1995-04-27

    Research progress utilizing flow cytometry is described. Topics include: rapid kinetics flow cytometry; characterization of size determinations for small DNA fragments; statistical analysis; energy transfer measurements of molecular confirmation in micelles; and enrichment of Mus spretus chromosomes by dual parameter flow sorting and identification of sorted fractions by fluorescence in-situ hybridization onto G-banded mouse metaphase spreads.

  15. Monitoring hyperproliferative disorders in human skin: flow cytometry of changing cytokeratin expression.

    NARCIS (Netherlands)

    Franssen, M.E.J.; Boezeman, J.B.M.; Kerkhof, P.C.M. van de; Erp, P.E.J. van

    2004-01-01

    BACKGROUND: Monitoring dynamics of different cell populations in solid tissues using flow cytometry has several limitations. The interaction and changes in epidermal subpopulations in hyperproliferative skin disorders such as psoriasis, a very common chronic inflammatory skin disease, may, however,

  16. External quality assessment in flow cytometry: educational aspects and trends toward improvement

    NARCIS (Netherlands)

    W.H.B.M. Levering

    2007-01-01

    textabstractFlow cytometry (FCM) uses the principles of hydro- dynamic focusing, light scattering, light excitation, and emission of fluorochrome molecules to generate specific multi-parameter data from particles and cells. FCM became rapidly a routine method for clinical decision-making in

  17. Analyse de données de cytometrie de flux pour un grand nombre d'échantillons

    OpenAIRE

    Chen , Xiaoyi

    2015-01-01

    In the course of my Ph.D. work, I have developed and applied two new computational approaches for automatic identification of cell populations in multi-parameter flow cytometry across a large number of samples. Both approaches were motivated and taken by the LabEX "Milieu Intérieur" study (hereafter MI study). In this project, ten 8-color flow cytometry panels were standardized for assessment of the major and minor cell populations present in peripheral whole blood, and data were collected an...

  18. Detection of circulating breast cancer cells using photoacoustic flow cytometry

    Science.gov (United States)

    Bhattacharyya, Kiran

    According to the American Cancer Society, more than 200,000 new cases of breast cancer are expected to be diagnosed this year. Moreover, about 40,000 women died from breast cancer last year alone. As breast cancer progresses in an individual, it can transform from a localized state to a metastatic one with multiple tumors distributed through the body, not necessarily contained within the breast. Metastasis is the spread of cancer through the body by circulating tumor cells (CTCs) which can be found in the blood and lymph of the diagnosed patient. Diagnosis of a metastatic state by the discovery of a secondary tumor can often come too late and hence, significantly reduce the patient's chance of survival. There is a current need for a CTC detection method which would diagnose metastasis before the secondary tumor occurs or reaches a size resolvable by current imaging systems. Since earlier detection would improve prognosis, this study proposes a method of labeling of breast cancer cells for detection with a photoacoustic flow cytometry system as a model for CTC detection in human blood. Gold nanoparticles and fluorescent polystyrene nanoparticles are proposed as contrast agents for T47D, the breast cancer cell line of choice. The labeling, photoacoustic detection limit, and sensitivity are first characterized and then applied to a study to show detection from human blood.

  19. In vitro flow cytometry-based screening platform for cellulase engineering

    Science.gov (United States)

    Körfer, Georgette; Pitzler, Christian; Vojcic, Ljubica; Martinez, Ronny; Schwaneberg, Ulrich

    2016-01-01

    Ultrahigh throughput screening (uHTS) plays an essential role in directed evolution for tailoring biocatalysts for industrial applications. Flow cytometry-based uHTS provides an efficient coverage of the generated protein sequence space by analysis of up to 107 events per hour. Cell-free enzyme production overcomes the challenge of diversity loss during the transformation of mutant libraries into expression hosts, enables directed evolution of toxic enzymes, and holds the promise to efficiently design enzymes of human or animal origin. The developed uHTS cell-free compartmentalization platform (InVitroFlow) is the first report in which a flow cytometry-based screened system has been combined with compartmentalized cell-free expression for directed cellulase enzyme evolution. InVitroFlow was validated by screening of a random cellulase mutant library employing a novel screening system (based on the substrate fluorescein-di-β-D-cellobioside), and yielded significantly improved cellulase variants (e.g. CelA2-H288F-M1 (N273D/H288F/N468S) with 13.3-fold increased specific activity (220.60 U/mg) compared to CelA2 wildtype: 16.57 U/mg). PMID:27184298

  20. Combinatorial selection of aptamers: new radioligands for in vivo molecular imaging

    International Nuclear Information System (INIS)

    Pestourie, C.

    2005-10-01

    Aptamers are oligonucleotide structures selected for their capacity to bind to a desired target. The first part of this work focuses on the selection of aptamers directed against the oncogenic form of the tyrosine kinase receptor Ret (RetC634Y). We compared different selection protocols: i) selection against the purified RetC634Y recombinant protein, ii) selection against whole living cells which express RetC634Y and iii) a crossover selection alternating between cells and recombinant protein. One aptamer, D4, was found to be able to inhibit Ret and to reverse the cell phenotype induced by the activation of the receptor. Then, we developed the in vivo use of the selected aptamers. Finally, we used the whole living cells selection protocol to develop aptamers against HLA-G. This protein is characterised by its function in immuno tolerance. Taken together, these studies should pave the way for the in vivo use of aptamers as new therapeutic and diagnostic agents for in vivo PET imaging. (author)

  1. Q selection for an electro-optical earth imaging system: theoretical and experimental results.

    Science.gov (United States)

    Cochrane, Andy; Schulz, Kevin; Kendrick, Rick; Bell, Ray

    2013-09-23

    This paper explores practical design considerations for selecting Q for an electro-optical earth imaging system, where Q is defined as (λ FN) / pixel pitch. Analytical methods are used to show that, under imaging conditions with high SNR, increasing Q with fixed aperture cannot lead to degradation of image quality regardless of the angular smear rate of the system. The potential for degradation of image quality under low SNR is bounded by an increase of the detector noise scaling as Q. An imaging test bed is used to collect representative imagery for various Q configurations. The test bed includes real world errors such as image smear and haze. The value of Q is varied by changing the focal length of the imaging system. Imagery is presented over a broad range of parameters.

  2. Assessing the Performance of Imaging Health Systems in Five Selected Hospitals in Uganda

    Directory of Open Access Journals (Sweden)

    Michael G Kawooya

    2012-01-01

    Full Text Available Objectives : The first objective of the study was to develop an index termed as the ′Imaging Coverage′ (IC, for measuring the performance of the imaging health systems. This index together with the Hospital-Based Utilization (HBU would then be calculated for five Ugandan hospitals. Second, was to relate the financial resources and existing health policy to the performance of the imaging systems. Materials and Methods: This was a cross-sectional survey employing the triangulation methodology, conducted in Mulago National Referral Hospital. The qualitative study used cluster sampling, in-depth interviews, focus group discussions, and self-administered questionnaires to explore the non-measurable aspects of the imaging systems′ performances. Results: The IC developed and tested as an index for the imaging system′s performance was 36%. General X-rays had the best IC followed by ultrasound. The Hospital-Based Utilization for the five selected hospitals was 186 per thousand and was the highest for general radiography followed by ultrasound. Conclusion: The IC for the five selected hospitals was 36% and the HBU was 186 per thousand, reflecting low performance levels, largely attributable to inadequate funding. There were shortfalls in imaging requisitions and inefficiencies in the imaging systems, financing, and health policy. Although the proportion of inappropriate imaging was small, reducing this inappropriateness even further would lead to a significant total saving, which could be channeled into investigating more patients. Financial resources stood out as the major limitation in attaining the desired performance and there is a need to increase budget funding so as to improve the performance of the imaging health systems.

  3. A Flow Cytometry Protocol to Estimate DNA Content in the Yellowtail Tetra Astyanax altiparanae

    Directory of Open Access Journals (Sweden)

    Pedro L. P. Xavier

    2017-09-01

    Full Text Available The production of triploid yellowtail tetra Astyanax altiparanae is a key factor to obtain permanently sterile individuals by chromosome set manipulation. Flow cytometric analysis is the main tool for confirmation of the resultant triploids individuals, but very few protocols are specific for A. altiparanae species. The current study has developed a protocol to estimate DNA content in this species. Furthermore, a protocol for long-term storage of dorsal fins used for flow cytometry analysis was established. The combination of five solutions with three detergents (Nonidet P-40 Substitute, Tween 20, and Triton X-100 at 0.1, 0.2, and 0.4% concentration was evaluated. Using the best solution from this first experiment, the addition of trypsin (0.125, 0.25, and 0.5% and sucrose (74 mM and the effects of increased concentrations of the detergents at 0.6 and 1.2% concentration were also evaluated. After adjustment of the protocol for flow cytometry, preservation of somatic tissue or isolated nuclei was also evaluated by freezing (at −20°C and fixation in saturated NaCl solution, acetic methanol (1:3, ethanol, and formalin at 10% for 30 or 60 days of storage at 25°C. Flow cytometry analysis in yellowtail tetra species was optimized using the following conditions: lysis solution: 9.53 mM MgCl2.7H20; 47.67 mM KCl; 15 mM Tris; 74 mM sucrose, 0.6% Triton X-100, pH 8.0; staining solution: Dulbecco's PBS with DAPI 1 μg mL−1; preservation procedure: somatic cells (dorsal fin samples frozen at −20°C. Using this protocol, samples may be stored up to 60 days with good accuracy for flow cytometry analysis.

  4. A novel method for measuring cellular antibody uptake using imaging flow cytometry reveals distinct uptake rates for two different monoclonal antibodies targeting L1.

    Science.gov (United States)

    Hazin, John; Moldenhauer, Gerhard; Altevogt, Peter; Brady, Nathan R

    2015-08-01

    Monoclonal antibodies (mAbs) have emerged as a promising tool for cancer therapy. Differing approaches utilize mAbs to either deliver a drug to the tumor cells or to modulate the host's immune system to mediate tumor kill. The rate by which a therapeutic antibody is being internalized by tumor cells is a decisive feature for choosing the appropriate treatment strategy. We herein present a novel method to effectively quantitate antibody uptake of tumor cells by using image-based flow cytometry, which combines image analysis with high throughput of sample numbers and sample size. The use of this method is established by determining uptake rate of an anti-EpCAM antibody (HEA125), from single cell measurements of plasma membrane versus internalized antibody, in conjunction with inhibitors of endocytosis. The method is then applied to two mAbs (L1-9.3, L1-OV52.24) targeting the neural cell adhesion molecule L1 (L1CAM) at two different epitopes. Based on median cell population responses, we find that mAb L1-OV52.24 is rapidly internalized by the ovarian carcinoma cell line SKOV3ip while L1 mAb 9.3 is mainly retained at the cell surface. These findings suggest the L1 mAb OV52.24 as a candidate to be further developed for drug-delivery to cancer cells, while L1-9.3 may be optimized to tag the tumor cells and stimulate immunogenic cancer cell killing. Furthermore, when analyzing cell-to-cell variability, we observed L1 mAb OV52.24 rapidly transition into a subpopulation with high-internalization capacity. In summary, this novel high-content method for measuring antibody internalization rate provides a high level of accuracy and sensitivity for cell population measurements and reveals further biologically relevant information when taking into account cellular heterogeneity. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Tumor recognition in wireless capsule endoscopy images using textural features and SVM-based feature selection.

    Science.gov (United States)

    Li, Baopu; Meng, Max Q-H

    2012-05-01

    Tumor in digestive tract is a common disease and wireless capsule endoscopy (WCE) is a relatively new technology to examine diseases for digestive tract especially for small intestine. This paper addresses the problem of automatic recognition of tumor for WCE images. Candidate color texture feature that integrates uniform local binary pattern and wavelet is proposed to characterize WCE images. The proposed features are invariant to illumination change and describe multiresolution characteristics of WCE images. Two feature selection approaches based on support vector machine, sequential forward floating selection and recursive feature elimination, are further employed to refine the proposed features for improving the detection accuracy. Extensive experiments validate that the proposed computer-aided diagnosis system achieves a promising tumor recognition accuracy of 92.4% in WCE images on our collected data.

  6. Dynamic measurements of flowing cells labeled by gold nanoparticles using full-field photothermal interferometric imaging

    Science.gov (United States)

    Turko, Nir A.; Roitshtain, Darina; Blum, Omry; Kemper, Björn; Shaked, Natan T.

    2017-06-01

    We present highly dynamic photothermal interferometric phase microscopy for quantitative, selective contrast imaging of live cells during flow. Gold nanoparticles can be biofunctionalized to bind to specific cells, and stimulated for local temperature increase due to plasmon resonance, causing a rapid change of the optical phase. These phase changes can be recorded by interferometric phase microscopy and analyzed to form an image of the binding sites of the nanoparticles in the cells, gaining molecular specificity. Since the nanoparticle excitation frequency might overlap with the sample dynamics frequencies, photothermal phase imaging was performed on stationary or slowly dynamic samples. Furthermore, the computational analysis of the photothermal signals is time consuming. This makes photothermal imaging unsuitable for applications requiring dynamic imaging or real-time analysis, such as analyzing and sorting cells during fast flow. To overcome these drawbacks, we utilized an external interferometric module and developed new algorithms, based on discrete Fourier transform variants, enabling fast analysis of photothermal signals in highly dynamic live cells. Due to the self-interference module, the cells are imaged with and without excitation in video-rate, effectively increasing signal-to-noise ratio. Our approach holds potential for using photothermal cell imaging and depletion in flow cytometry.

  7. Combination of CD157 and FLAER to Detect Peripheral Blood Eosinophils by Multiparameter Flow Cytometry.

    Science.gov (United States)

    Carulli, Giovanni; Marini, Alessandra; Sammuri, Paola; Domenichini, Cristiana; Ottaviano, Virginia; Pacini, Simone; Petrini, Mario

    2015-01-01

    The identification of eosinophils by flow cytometry is difficult because most of the surface antigens expressed by eosinophils are shared with neutrophils. Some methods have been proposed, generally based on differential light scatter properties, enhanced autofluorescence, lack of CD16 or selective positivity of CD52. Such methods, however, show several limitations. In the present study we report a novel method based on the analysis of glycosylphosphatidylinositol (GPI)-linked molecules. The combination of CD157 and FLAER was used, since FLAER recognizes all GPI-linked molecules, while CD157 is absent on the membrane of eosinophils and expressed by neutrophils. Peripheral blood samples from normal subjects and patients with variable percentages of eosinophils (n = 31), and without any evidence for circulating immature myeloid cells, were stained with the combination of FLAER-Alexa Fluor and CD157-PE. A FascCanto II cytometer was used. Granulocytes were gated after CD33 staining and eosinophils were identified as CD157(-)/FLAER(+) events. Neutrophils were identified as CD157(+)/FLAER(+) events. The percentages of eosinophils detected by this method showed a very significant correlation both with automated counting and with manual counting (r = 0.981 and 0.989, respectively). Sorting assays were carried out by a S3 Cell Sorter: cytospins obtained from CD157(-)/FLAER(+) events consisted of 100% eosinophils, while samples from CD157(+)/FLAER(+) events were represented only by neutrophils. In conclusion, this method shows high sensitivity and specificity in order to distinguish eosinophils from neutrophils by flow cytometry. However, since CD157 is gradually up-regulated throughout bone marrow myeloid maturation, our method cannot be applied to cases characterized by immature myeloid cells.

  8. TU-CD-BRA-05: Atlas Selection for Multi-Atlas-Based Image Segmentation Using Surrogate Modeling

    International Nuclear Information System (INIS)

    Zhao, T; Ruan, D

    2015-01-01

    Purpose: The growing size and heterogeneity in training atlas necessitates sophisticated schemes to identify only the most relevant atlases for the specific multi-atlas-based image segmentation problem. This study aims to develop a model to infer the inaccessible oracle geometric relevance metric from surrogate image similarity metrics, and based on such model, provide guidance to atlas selection in multi-atlas-based image segmentation. Methods: We relate the oracle geometric relevance metric in label space to the surrogate metric in image space, by a monotonically non-decreasing function with additive random perturbations. Subsequently, a surrogate’s ability to prognosticate the oracle order for atlas subset selection is quantified probabilistically. Finally, important insights and guidance are provided for the design of fusion set size, balancing the competing demands to include the most relevant atlases and to exclude the most irrelevant ones. A systematic solution is derived based on an optimization framework. Model verification and performance assessment is performed based on clinical prostate MR images. Results: The proposed surrogate model was exemplified by a linear map with normally distributed perturbation, and verified with several commonly-used surrogates, including MSD, NCC and (N)MI. The derived behaviors of different surrogates in atlas selection and their corresponding performance in ultimate label estimate were validated. The performance of NCC and (N)MI was similarly superior to MSD, with a 10% higher atlas selection probability and a segmentation performance increase in DSC by 0.10 with the first and third quartiles of (0.83, 0.89), compared to (0.81, 0.89). The derived optimal fusion set size, valued at 7/8/8/7 for MSD/NCC/MI/NMI, agreed well with the appropriate range [4, 9] from empirical observation. Conclusion: This work has developed an efficacious probabilistic model to characterize the image-based surrogate metric on atlas selection

  9. Fusion of remote sensing images based on pyramid decomposition with Baldwinian Clonal Selection Optimization

    Science.gov (United States)

    Jin, Haiyan; Xing, Bei; Wang, Lei; Wang, Yanyan

    2015-11-01

    In this paper, we put forward a novel fusion method for remote sensing images based on the contrast pyramid (CP) using the Baldwinian Clonal Selection Algorithm (BCSA), referred to as CPBCSA. Compared with classical methods based on the transform domain, the method proposed in this paper adopts an improved heuristic evolutionary algorithm, wherein the clonal selection algorithm includes Baldwinian learning. In the process of image fusion, BCSA automatically adjusts the fusion coefficients of different sub-bands decomposed by CP according to the value of the fitness function. BCSA also adaptively controls the optimal search direction of the coefficients and accelerates the convergence rate of the algorithm. Finally, the fusion images are obtained via weighted integration of the optimal fusion coefficients and CP reconstruction. Our experiments show that the proposed method outperforms existing methods in terms of both visual effect and objective evaluation criteria, and the fused images are more suitable for human visual or machine perception.

  10. Determination of P-Glycoprotein Expression by Flow Cytometry in Hematological Malignancies

    Directory of Open Access Journals (Sweden)

    Berkay Saraymen

    2016-03-01

    Full Text Available Objective: Determination the expression of P-glycoprotein is especially problematic for normal tissues because immuno­logical methods are limited in terms of sensitivity. We aimed to determine the expression of P-glycoprotein and CD34 by flow cytometry, and to evaluate the level of expression of P-glycoprotein and CD34 with unresponsive to treatment in pa­tients diagnosed with hematologic malignancy. Methods: Our study included fifty patients diagnosed with acute myeloblastic leukemia and acute lymphoblastic leuke­mia, and twenty healthy controls who were admitted to Erci­yes University Hematology-Oncology Hospital. The suspend­ed cells from bone marrow samples of patients and the pe­ripheral blood samples of healthy people were marked with P-glycoprotein phycoerythrin and CD34 FITC or PerCP Cy 5.5; and then surface expression was measured by means of flow cytometry. Results: In 6 of 30 acute myeloblastic leukemia patients P-glycoprotein and CD34 expression, in 6 of 20 acute lympho­blastic leukemia patients P-glycoprotein, in 5 of them CD34 expression were determined. A significant relation between P-glycoprotein and CD34 expressions in acute myeloblas­tic leukemia and acute lymphoblastic leukemia bone marrow samples was reported. Conclusion: Our data indicate that flow cytometry is more reliable, precise and faster than molecular methods for mea­suring P-glycoprotein expression and suggests the pos­sibility of a significant relationship between P-glycoprotein and CD34 expressions in acute myeloblastic leukemia and acute lymphoblastic leukemia bone marrow samples. The blast cells expressing CD34 on their surface along with P-glycoprotein simultaneously show that multi drug resistance 1 gene is mostly active in immature cells.

  11. A liposome-based size calibration method for measuring microvesicles by flow cytometry

    DEFF Research Database (Denmark)

    Simonsen, Jens Bæk

    2016-01-01

    BACKGROUND: Over the last years the need for a gold standard to determine the sizes of extracellular vesicles including microvesicles by flow cytometry has been emphasized. METHODS: This work suggests to use artificial vesicles as calibrators to ascertain the size of microvesicles from the side...

  12. Novel quantitative autophagy analysis by organelle flow cytometry after cell sonication.

    Directory of Open Access Journals (Sweden)

    Michael Degtyarev

    Full Text Available Autophagy is a dynamic process of bulk degradation of cellular proteins and organelles in lysosomes. Current methods of autophagy measurement include microscopy-based counting of autophagic vacuoles (AVs in cells. We have developed a novel method to quantitatively analyze individual AVs using flow cytometry. This method, OFACS (organelle flow after cell sonication, takes advantage of efficient cell disruption with a brief sonication, generating cell homogenates with fluorescently labeled AVs that retain their integrity as confirmed with light and electron microscopy analysis. These AVs could be detected directly in the sonicated cell homogenates on a flow cytometer as a distinct population of expected organelle size on a cytometry plot. Treatment of cells with inhibitors of autophagic flux, such as chloroquine or lysosomal protease inhibitors, increased the number of particles in this population under autophagy inducing conditions, while inhibition of autophagy induction with 3-methyladenine or knockdown of ATG proteins prevented this accumulation. This assay can be easily performed in a high-throughput format and opens up previously unexplored avenues for autophagy analysis.

  13. Adaptive Spot Detection With Optimal Scale Selection in Fluorescence Microscopy Images.

    Science.gov (United States)

    Basset, Antoine; Boulanger, Jérôme; Salamero, Jean; Bouthemy, Patrick; Kervrann, Charles

    2015-11-01

    Accurately detecting subcellular particles in fluorescence microscopy is of primary interest for further quantitative analysis such as counting, tracking, or classification. Our primary goal is to segment vesicles likely to share nearly the same size in fluorescence microscopy images. Our method termed adaptive thresholding of Laplacian of Gaussian (LoG) images with autoselected scale (ATLAS) automatically selects the optimal scale corresponding to the most frequent spot size in the image. Four criteria are proposed and compared to determine the optimal scale in a scale-space framework. Then, the segmentation stage amounts to thresholding the LoG of the intensity image. In contrast to other methods, the threshold is locally adapted given a probability of false alarm (PFA) specified by the user for the whole set of images to be processed. The local threshold is automatically derived from the PFA value and local image statistics estimated in a window whose size is not a critical parameter. We also propose a new data set for benchmarking, consisting of six collections of one hundred images each, which exploits backgrounds extracted from real microscopy images. We have carried out an extensive comparative evaluation on several data sets with ground-truth, which demonstrates that ATLAS outperforms existing methods. ATLAS does not need any fine parameter tuning and requires very low computation time. Convincing results are also reported on real total internal reflection fluorescence microscopy images.

  14. Development of EMab-51, a Sensitive and Specific Anti-Epidermal Growth Factor Receptor Monoclonal Antibody in Flow Cytometry, Western Blot, and Immunohistochemistry.

    Science.gov (United States)

    Itai, Shunsuke; Kaneko, Mika K; Fujii, Yuki; Yamada, Shinji; Nakamura, Takuro; Yanaka, Miyuki; Saidoh, Noriko; Handa, Saori; Chang, Yao-Wen; Suzuki, Hiroyoshi; Harada, Hiroyuki; Kato, Yukinari

    2017-10-01

    The epidermal growth factor receptor (EGFR) is a member of the human epidermal growth factor receptor (HER) family of receptor tyrosine kinases and is involved in cell growth and differentiation. EGFR homodimers or heterodimers with other HER members, such as HER2 and HER3, activate downstream signaling cascades in many cancers. In this study, we developed novel anti-EGFR monoclonal antibodies (mAbs) and characterized their efficacy in flow cytometry, Western blot, and immunohistochemical analyses. First, we expressed the full-length or ectodomain of EGFR in LN229 glioblastoma cells and then immunized mice with LN229/EGFR or ectodomain of EGFR, and performed the first screening using enzyme-linked immunosorbent assays. Subsequently, we selected mAbs according to their efficacy in flow cytometry (second screening), Western blot (third screening), and immunohistochemical (fourth screening) analyses. Among 100 mAbs, only one clone EMab-51 (IgG 1 , kappa) reacted with EGFR in Western blot analysis. Finally, immunohistochemical analyses with EMab-51 showed sensitive and specific reactions against oral cancer cells, warranting the use of EMab-51 to detect EGFR in pathological analyses of EGFR-expressing cancers.

  15. Novel high-throughput cell-based hybridoma screening methodology using the Celigo Image Cytometer.

    Science.gov (United States)

    Zhang, Haohai; Chan, Leo Li-Ying; Rice, William; Kassam, Nasim; Longhi, Maria Serena; Zhao, Haitao; Robson, Simon C; Gao, Wenda; Wu, Yan

    2017-08-01

    Hybridoma screening is a critical step for antibody discovery, which necessitates prompt identification of potential clones from hundreds to thousands of hybridoma cultures against the desired immunogen. Technical issues associated with ELISA- and flow cytometry-based screening limit accuracy and diminish high-throughput capability, increasing time and cost. Conventional ELISA screening with coated antigen is also impractical for difficult-to-express hydrophobic membrane antigens or multi-chain protein complexes. Here, we demonstrate novel high-throughput screening methodology employing the Celigo Image Cytometer, which avoids nonspecific signals by contrasting antibody binding signals directly on living cells, with and without recombinant antigen expression. The image cytometry-based high-throughput screening method was optimized by detecting the binding of hybridoma supernatants to the recombinant antigen CD39 expressed on Chinese hamster ovary (CHO) cells. Next, the sensitivity of the image cytometer was demonstrated by serial dilution of purified CD39 antibody. Celigo was used to measure antibody affinities of commercial and in-house antibodies to membrane-bound CD39. This cell-based screening procedure can be completely accomplished within one day, significantly improving throughput and efficiency of hybridoma screening. Furthermore, measuring direct antibody binding to living cells eliminated both false positive and false negative hits. The image cytometry method was highly sensitive and versatile, and could detect positive antibody in supernatants at concentrations as low as ~5ng/mL, with concurrent K d binding affinity coefficient determination. We propose that this screening method will greatly facilitate antibody discovery and screening technologies. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Automatic selection of resting-state networks with functional magnetic resonance imaging

    Directory of Open Access Journals (Sweden)

    Silvia Francesca eStorti

    2013-05-01

    Full Text Available Functional magnetic resonance imaging (fMRI during a resting-state condition can reveal the co-activation of specific brain regions in distributed networks, called resting-state networks, which are selected by independent component analysis (ICA of the fMRI data. One of the major difficulties with component analysis is the automatic selection of the ICA features related to brain activity. In this study we describe a method designed to automatically select networks of potential functional relevance, specifically, those regions known to be involved in motor function, visual processing, executive functioning, auditory processing, memory, and the default-mode network. To do this, image analysis was based on probabilistic ICA as implemented in FSL software. After decomposition, the optimal number of components was selected by applying a novel algorithm which takes into account, for each component, Pearson's median coefficient of skewness of the spatial maps generated by FSL, followed by clustering, segmentation, and spectral analysis. To evaluate the performance of the approach, we investigated the resting-state networks in 25 subjects. For each subject, three resting-state scans were obtained with a Siemens Allegra 3 T scanner (NYU data set. Comparison of the visually and the automatically identified neuronal networks showed that the algorithm had high accuracy (first scan: 95%, second scan: 95%, third scan: 93% and precision (90%, 90%, 84%. The reproducibility of the networks for visual and automatic selection was very close: it was highly consistent in each subject for the default-mode network (≥ 92% and the occipital network, which includes the medial visual cortical areas (≥ 94%, and consistent for the attention network (≥ 80%, the right and/or left lateralized frontoparietal attention networks, and the temporal-motor network (≥ 80%. The automatic selection method may be used to detect neural networks and reduce subjectivity in ICA

  17. SPITZER IMAGING OF STRONGLY LENSED HERSCHEL-SELECTED DUSTY STAR-FORMING GALAXIES

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Brian; Cooray, Asantha; Calanog, J. A.; Nayyeri, H.; Timmons, N.; Casey, C. [Department of Physics and Astronomy, University of California, Irvine, CA 92697 (United States); Baes, M. [Sterrenkundig Observatorium, Universiteit Gent, Krijgslaan 281 S9, B-9000 Gent (Belgium); Chapman, S. [Department of Physics and Atmospheric Science, Dalhousie University, Halifax, Nova Scotia, B3H 4R2 (Canada); Dannerbauer, H. [Laboratoire AIM-Paris-Saclay, CEA/DSM/Irfu-CNRS-Université Paris Diderot, CE-Saclay, pt courrier 131, F-91191 Gif-sur-Yvette (France); Da Cunha, E. [Center for Astrophysics and Supercomputing, Swinburne University of Technology, Hawthorn VIC 3122 (Australia); De Zotti, G. [INAF-Osservatorio Astronomico di Padova, Vicolo Osservatorio 5, I-35122 Padova (Italy); Dunne, L.; Michałowski, M. J.; Oteo, I. [Institute for Astronomy, University of Edinburgh, Royal Observatory, Blackford Hill, Edinburgh, EH9 3HJ (United Kingdom); Farrah, D. [Department of Physics, Virginia Tech, Blacksburg, VA 24061 (United States); Fu, Hai [Department of Physics and Astronomy, University of Iowa, Van Allen Hall, Iowa City, IA 52242 (United States); Gonzalez-Nuevo, J. [Departamento de Fisica, Universidad de Oviedo C/ Calvo Sotelo, s/n, E-33007 Oviedo (Spain); Magdis, G. [Department of Astrophysics, Denys Wilkinson Building, University of Oxford, Keble Road, Oxford OX1 3RH (United Kingdom); Riechers, D. A. [Department of Astronomy, Cornell University, 220 Space Sciences Building, Ithaca, NY 14853 (United States); Scott, D. [Department of Physics and Astronomy, University of British Columbia, 6224 Agricultural Road, Vancouver, BC V6T 1Z1 (Canada); and others

    2015-11-20

    We present the rest-frame optical spectral energy distribution (SED) and stellar masses of six Herschel-selected gravitationally lensed dusty, star-forming galaxies (DSFGs) at 1 < z < 3. These galaxies were first identified with Herschel/SPIRE imaging data from the Herschel Astrophysical Terahertz Large Area Survey (H-ATLAS) and the Herschel Multi-tiered Extragalactic Survey (HerMES). The targets were observed with Spitzer/IRAC at 3.6 and 4.5 μm. Due to the spatial resolution of the IRAC observations at the level of 2″, the lensing features of a background DSFG in the near-infrared are blended with the flux from the foreground lensing galaxy in the IRAC imaging data. We make use of higher resolution Hubble/WFC3 or Keck/NIRC2 Adaptive Optics imaging data to fit light profiles of the foreground lensing galaxy (or galaxies) as a way to model the foreground components, in order to successfully disentangle the foreground lens and background source flux densities in the IRAC images. The flux density measurements at 3.6 and 4.5 μm, once combined with Hubble/WFC3 and Keck/NIRC2 data, provide important constraints on the rest-frame optical SED of the Herschel-selected lensed DSFGs. We model the combined UV- to millimeter-wavelength SEDs to establish the stellar mass, dust mass, star formation rate, visual extinction, and other parameters for each of these Herschel-selected DSFGs. These systems have inferred stellar masses in the range 8 × 10{sup 10}–4 × 10{sup 11} M{sub ⊙} and star formation rates of around 100 M{sub ⊙} yr{sup −1}. This puts these lensed submillimeter systems well above the SFR-M* relation observed for normal star-forming galaxies at similar redshifts. The high values of SFR inferred for these systems are consistent with a major merger-driven scenario for star formation.

  18. Peering beneath the surface: novel imaging techniques to noninvasively select gametes and embryos for ART.

    Science.gov (United States)

    Jasensky, Joshua; Swain, Jason E

    2013-10-01

    Embryo imaging has long been a critical tool for in vitro fertilization laboratories, aiding in morphological assessment of embryos, which remains the primary tool for embryo selection. With the recent emergence of clinically applicable real-time imaging systems to assess embryo morphokinetics, a renewed interest has emerged regarding noninvasive methods to assess gamete and embryo development as a means of inferring quality. Several studies exist that utilize novel imaging techniques to visualize or quantify intracellular components of gametes and embryos with the intent of correlating localization of organelles or molecular constitution with quality or outcome. However, the safety of these approaches varies due to the potential detrimental impact of light exposure or other variables. Along with complexity of equipment and cost, these drawbacks currently limit clinical application of these novel microscopes and imaging techniques. However, as evidenced by clinical incorporation of some real-time imaging devices as well as use of polarized microscopy, some of these imaging approaches may prove to be useful. This review summarizes the existing literature on novel imaging approaches utilized to examine gametes and embryos. Refinement of some of these imaging systems may permit clinical application and serve as a means to offer new, noninvasive selection tools to improve outcomes for various assisted reproductive technology procedures.

  19. Advances in feature selection methods for hyperspectral image processing in food industry applications: a review.

    Science.gov (United States)

    Dai, Qiong; Cheng, Jun-Hu; Sun, Da-Wen; Zeng, Xin-An

    2015-01-01

    There is an increased interest in the applications of hyperspectral imaging (HSI) for assessing food quality, safety, and authenticity. HSI provides abundance of spatial and spectral information from foods by combining both spectroscopy and imaging, resulting in hundreds of contiguous wavebands for each spatial position of food samples, also known as the curse of dimensionality. It is desirable to employ feature selection algorithms for decreasing computation burden and increasing predicting accuracy, which are especially relevant in the development of online applications. Recently, a variety of feature selection algorithms have been proposed that can be categorized into three groups based on the searching strategy namely complete search, heuristic search and random search. This review mainly introduced the fundamental of each algorithm, illustrated its applications in hyperspectral data analysis in the food field, and discussed the advantages and disadvantages of these algorithms. It is hoped that this review should provide a guideline for feature selections and data processing in the future development of hyperspectral imaging technique in foods.

  20. Flow Cytometry Method as a Diagnostic Tool for Pleural Fluid Involvement in a Patient with Multiple Myeloma

    Directory of Open Access Journals (Sweden)

    MUZAFFER KEKLIK

    2012-01-01

    Full Text Available

    Multiple myeloma is a malignant proliferation of plasma cells that mainly affects bone marrow. Pleural effusions secondary to pleural myelomatous involvement have rarely been reported in the literature. As it is rarely detected, we aimed to report a case in which pleural effusion of a multiple myeloma was confirmed as true myelomatous involvement by flow cytometry method. A 52-years old man presented to our clinic with chest and back pain lasting for 3 months. On the chest radiography, pleural fluid was detected in left hemithorax. Pleural fluid flow cytometry was performed. In the flow cytometry, CD56, CD38 and CD138 found to be positive, while CD19 was negative. True myelomatous pleural effusions are very uncommon, with fewer than 100 cases reported worldwide. Flow cytometry is a potentially useful diagnostic tool for clinical practice. We presented our case; as it has been rarely reported, although flow cytometer is a simple method for detection of pleural fluid involvement in multiple myeloma.

  1. Investigation of psychophysical similarity measures for selection of similar images in the diagnosis of clustered microcalcifications on mammograms

    International Nuclear Information System (INIS)

    Muramatsu, Chisako; Li Qiang; Schmidt, Robert; Shiraishi, Junji; Doi, Kunio

    2008-01-01

    The presentation of images with lesions of known pathology that are similar to an unknown lesion may be helpful to radiologists in the diagnosis of challenging cases for improving the diagnostic accuracy and also for reducing variation among different radiologists. The authors have been developing a computerized scheme for automatically selecting similar images with clustered microcalcifications on mammograms from a large database. For similar images to be useful, they must be similar from the point of view of the diagnosing radiologists. In order to select such images, subjective similarity ratings were obtained for a number of pairs of clustered microcalcifications by breast radiologists for establishment of a ''gold standard'' of image similarity, and the gold standard was employed for determination and evaluation of the selection of similar images. The images used in this study were obtained from the Digital Database for Screening Mammography developed by the University of South Florida. The subjective similarity ratings for 300 pairs of images with clustered microcalcifications were determined by ten breast radiologists. The authors determined a number of image features which represent the characteristics of clustered microcalcifications that radiologists would use in their diagnosis. For determination of objective similarity measures, an artificial neural network (ANN) was employed. The ANN was trained with the average subjective similarity ratings as teacher and selected image features as input data. The ANN was trained to learn the relationship between the image features and the radiologists' similarity ratings; therefore, once the training was completed, the ANN was able to determine the similarity, called a psychophysical similarity measure, which was expected to be close to radiologists' impressions, for an unknown pair of clustered microcalcifications. By use of a leave-one-out test method, the best combination of features was selected. The correlation

  2. Investigation of psychophysical similarity measures for selection of similar images in the diagnosis of clustered microcalcifications on mammograms

    Energy Technology Data Exchange (ETDEWEB)

    Muramatsu, Chisako; Li Qiang; Schmidt, Robert; Shiraishi, Junji; Doi, Kunio [Department of Radiology, University of Chicago, 5841 South Maryland Avenue, Chicago, Illinois 60637 (United States) and Department of Intelligent Image Information, Gifu University, 1-1 Yanagido, Gifu (Japan); Department of Radiology, Duke Advanced Imaging Labs, Duke University, 2424 Erwin Road, Suite 302, Durham, North Carolina 27705 (United States); Department of Radiology, University of Chicago, 5841 South Maryland Avenue, Chicago, Illinois 60637 (United States)

    2008-12-15

    The presentation of images with lesions of known pathology that are similar to an unknown lesion may be helpful to radiologists in the diagnosis of challenging cases for improving the diagnostic accuracy and also for reducing variation among different radiologists. The authors have been developing a computerized scheme for automatically selecting similar images with clustered microcalcifications on mammograms from a large database. For similar images to be useful, they must be similar from the point of view of the diagnosing radiologists. In order to select such images, subjective similarity ratings were obtained for a number of pairs of clustered microcalcifications by breast radiologists for establishment of a ''gold standard'' of image similarity, and the gold standard was employed for determination and evaluation of the selection of similar images. The images used in this study were obtained from the Digital Database for Screening Mammography developed by the University of South Florida. The subjective similarity ratings for 300 pairs of images with clustered microcalcifications were determined by ten breast radiologists. The authors determined a number of image features which represent the characteristics of clustered microcalcifications that radiologists would use in their diagnosis. For determination of objective similarity measures, an artificial neural network (ANN) was employed. The ANN was trained with the average subjective similarity ratings as teacher and selected image features as input data. The ANN was trained to learn the relationship between the image features and the radiologists' similarity ratings; therefore, once the training was completed, the ANN was able to determine the similarity, called a psychophysical similarity measure, which was expected to be close to radiologists' impressions, for an unknown pair of clustered microcalcifications. By use of a leave-one-out test method, the best combination of features

  3. An optical authentication system based on imaging of excitation-selected lanthanide luminescence.

    Science.gov (United States)

    Carro-Temboury, Miguel R; Arppe, Riikka; Vosch, Tom; Sørensen, Thomas Just

    2018-01-01

    Secure data encryption relies heavily on one-way functions, and copy protection relies on features that are difficult to reproduce. We present an optical authentication system based on lanthanide luminescence from physical one-way functions or physical unclonable functions (PUFs). They cannot be reproduced and thus enable unbreakable encryption. Further, PUFs will prevent counterfeiting if tags with unique PUFs are grafted onto products. We have developed an authentication system that comprises a hardware reader, image analysis, and authentication software and physical keys that we demonstrate as an anticounterfeiting system. The physical keys are PUFs made from random patterns of taggants in polymer films on glass that can be imaged following selected excitation of particular lanthanide(III) ions doped into the individual taggants. This form of excitation-selected imaging ensures that by using at least two lanthanide(III) ion dopants, the random patterns cannot be copied, because the excitation selection will fail when using any other emitter. With the developed reader and software, the random patterns are read and digitized, which allows a digital pattern to be stored. This digital pattern or digital key can be used to authenticate the physical key in anticounterfeiting or to encrypt any message. The PUF key was produced with a staggering nominal encoding capacity of 7 3600 . Although the encoding capacity of the realized authentication system reduces to 6 × 10 104 , it is more than sufficient to completely preclude counterfeiting of products.

  4. Noninvasive imaging systems for gametes and embryo selection in IVF programs: a review.

    Science.gov (United States)

    Omidi, Marjan; Faramarzi, Azita; Agharahimi, Azam; Khalili, Mohammad Ali

    2017-09-01

    Optimizing the efficiency of the in vitro fertilization procedure by improving pregnancy rates and reducing the risks of multiple pregnancies simultaneously are the primary goals of the current assisted reproductive technology program. With the move to single embryo transfers, the need for more cost-effective and noninvasive methods for embryo selection prior to transfer is paramount. These aims require advancement in a more acquire gametes/embryo testing and selection procedures using high-tech devices. Therefore, the aim of the present review is to evaluate the efficacy of noninvasive imaging systems in the current literatures, focusing on the potential clinical application in infertile patients undergoing assisted reproductive technology treatments. In this regards, three advanced imaging systems of motile sperm organelle morphology examination, polarization microscopy and time-lapse monitoring for the best selection of the gametes and preimplantation embryos are introduced in full. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

  5. Threshold selection for classification of MR brain images by clustering method

    Energy Technology Data Exchange (ETDEWEB)

    Moldovanu, Simona [Faculty of Sciences and Environment, Department of Chemistry, Physics and Environment, Dunărea de Jos University of Galaţi, 47 Domnească St., 800008, Romania, Phone: +40 236 460 780 (Romania); Dumitru Moţoc High School, 15 Milcov St., 800509, Galaţi (Romania); Obreja, Cristian; Moraru, Luminita, E-mail: luminita.moraru@ugal.ro [Faculty of Sciences and Environment, Department of Chemistry, Physics and Environment, Dunărea de Jos University of Galaţi, 47 Domnească St., 800008, Romania, Phone: +40 236 460 780 (Romania)

    2015-12-07

    Given a grey-intensity image, our method detects the optimal threshold for a suitable binarization of MR brain images. In MR brain image processing, the grey levels of pixels belonging to the object are not substantially different from the grey levels belonging to the background. Threshold optimization is an effective tool to separate objects from the background and further, in classification applications. This paper gives a detailed investigation on the selection of thresholds. Our method does not use the well-known method for binarization. Instead, we perform a simple threshold optimization which, in turn, will allow the best classification of the analyzed images into healthy and multiple sclerosis disease. The dissimilarity (or the distance between classes) has been established using the clustering method based on dendrograms. We tested our method using two classes of images: the first consists of 20 T2-weighted and 20 proton density PD-weighted scans from two healthy subjects and from two patients with multiple sclerosis. For each image and for each threshold, the number of the white pixels (or the area of white objects in binary image) has been determined. These pixel numbers represent the objects in clustering operation. The following optimum threshold values are obtained, T = 80 for PD images and T = 30 for T2w images. Each mentioned threshold separate clearly the clusters that belonging of the studied groups, healthy patient and multiple sclerosis disease.

  6. Using Lanthanide Nanoparticles as Isotopic Tags for Biomarker Detection by Mass Cytometry

    Science.gov (United States)

    Cao, Pengpeng

    The development of robust, versatile, and high-throughput biosensing techniques has widespread implications for early disease detection and accurate diagnosis. An innovative technology, mass cytometry, has been developed to use isotopically-labelled antibodies to simultaneously study multiple parameters of single cells. The current detection sensitivity of mass cytometry is limited by the number of copies of a given isotope that can be attached to a given antibody. This thesis describes research on the synthesis, characterization, and bioconjugation of a new class of nanoparticle-based labelling agents to be employed for the detection of low-abundance biomarkers by mass cytometry. Hydrophobic lanthanide nanoparticles (Ln NPs) have been prepared by the Winnik group. To render the NPs water-soluble for biological applications, we coated the NP surface with a first generation of multidentate poly(ethylene glycol) (PEG)-based ligands via ligand exchange. We measured the size, morphology, and polydispersity of these hydrophilic NPs by transmission electron microscopy (TEM) and dynamic light scattering (DLS). The colloidal stability of the NPs was determined at various pH and in phosphate buffered saline (PBS) solutions. Tetradentate-PEG-coated NPs (Tetra-NPs) exhibited the best stability at pH 3 to 9, and in PBS. However, when cells were treated with Tetra-NPs in preliminary in vitro studies, significant undesirable non-specific binding (NSB) was observed. In order to tackle the NSB issue presented in the Tetra-NPs, we prepared a second generation of polymer-based ligands using ring-opening metathesis polymerization (ROMP). A small library of ROMP polymers was synthesized, characterized, and used to stabilize NPs in aqueous solutions. The ROMP-NPs were found to have significantly reduced NSB to cells by inductively coupled plasma-mass spectrometry (ICP-MS). To further modify the NPs, amine groups were introduced as functional handles to both the tetradentate-PEG and

  7. The impact of digital imaging in the field of cytopathology.

    Science.gov (United States)

    Pantanowitz, Liron; Hornish, Maryanne; Goulart, Robert A

    2009-03-06

    With the introduction of digital imaging, pathology is undergoing a digital transformation. In the field of cytology, digital images are being used for telecytology, automated screening of Pap test slides, training and education (e.g. online digital atlases), and proficiency testing. To date, there has been no systematic review on the impact of digital imaging on the practice of cytopathology. This article critically addresses the emerging role of computer-assisted screening and the application of digital imaging to the field of cytology, including telecytology, virtual microscopy, and the impact of online cytology resources. The role of novel diagnostic techniques like image cytometry is also reviewed.

  8. Selecting electrode configurations for image-guided cochlear implant programming using template matching.

    Science.gov (United States)

    Zhang, Dongqing; Zhao, Yiyuan; Noble, Jack H; Dawant, Benoit M

    2018-04-01

    Cochlear implants (CIs) are neural prostheses that restore hearing using an electrode array implanted in the cochlea. After implantation, the CI processor is programmed by an audiologist. One factor that negatively impacts outcomes and can be addressed by programming is cross-electrode neural stimulation overlap (NSO). We have proposed a system to assist the audiologist in programming the CI that we call image-guided CI programming (IGCIP). IGCIP permits using CT images to detect NSO and recommend deactivation of a subset of electrodes to avoid NSO. We have shown that IGCIP significantly improves hearing outcomes. Most of the IGCIP steps are robustly automated but electrode configuration selection still sometimes requires manual intervention. With expertise, distance-versus-frequency curves, which are a way to visualize the spatial relationship learned from CT between the electrodes and the nerves they stimulate, can be used to select the electrode configuration. We propose an automated technique for electrode configuration selection. A comparison between this approach and one we have previously proposed shows that our method produces results that are as good as those obtained with our previous method while being generic and requiring fewer parameters.

  9. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Deng-Liang [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Yang, Hai-Tao; Wang, Jiang-Jie [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yao, Pei-Sen [Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Pan, Ru-Jun [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yang, Chaoyong James, E-mail: cyyang@xmu.edu.cn [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Kang, De-Zhi, E-mail: kdzy99988@163.com [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

    2014-10-31

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

  10. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    International Nuclear Information System (INIS)

    Wang, Deng-Liang; Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan; Yang, Hai-Tao; Wang, Jiang-Jie; Yao, Pei-Sen; Pan, Ru-Jun; Yang, Chaoyong James; Kang, De-Zhi

    2014-01-01

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K d 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K d 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy

  11. OpenCyto: an open source infrastructure for scalable, robust, reproducible, and automated, end-to-end flow cytometry data analysis.

    Directory of Open Access Journals (Sweden)

    Greg Finak

    2014-08-01

    Full Text Available Flow cytometry is used increasingly in clinical research for cancer, immunology and vaccines. Technological advances in cytometry instrumentation are increasing the size and dimensionality of data sets, posing a challenge for traditional data management and analysis. Automated analysis methods, despite a general consensus of their importance to the future of the field, have been slow to gain widespread adoption. Here we present OpenCyto, a new BioConductor infrastructure and data analysis framework designed to lower the barrier of entry to automated flow data analysis algorithms by addressing key areas that we believe have held back wider adoption of automated approaches. OpenCyto supports end-to-end data analysis that is robust and reproducible while generating results that are easy to interpret. We have improved the existing, widely used core BioConductor flow cytometry infrastructure by allowing analysis to scale in a memory efficient manner to the large flow data sets that arise in clinical trials, and integrating domain-specific knowledge as part of the pipeline through the hierarchical relationships among cell populations. Pipelines are defined through a text-based csv file, limiting the need to write data-specific code, and are data agnostic to simplify repetitive analysis for core facilities. We demonstrate how to analyze two large cytometry data sets: an intracellular cytokine staining (ICS data set from a published HIV vaccine trial focused on detecting rare, antigen-specific T-cell populations, where we identify a new subset of CD8 T-cells with a vaccine-regimen specific response that could not be identified through manual analysis, and a CyTOF T-cell phenotyping data set where a large staining panel and many cell populations are a challenge for traditional analysis. The substantial improvements to the core BioConductor flow cytometry packages give OpenCyto the potential for wide adoption. It can rapidly leverage new developments in

  12. Selective principal component regression analysis of fluorescence hyperspectral image to assess aflatoxin contamination in corn

    Science.gov (United States)

    Selective principal component regression analysis (SPCR) uses a subset of the original image bands for principal component transformation and regression. For optimal band selection before the transformation, this paper used genetic algorithms (GA). In this case, the GA process used the regression co...

  13. High-throughput tri-colour flow cytometry technique to assess Plasmodium falciparum parasitaemia in bioassays

    DEFF Research Database (Denmark)

    Tiendrebeogo, Regis W; Adu, Bright; Singh, Susheel K

    2014-01-01

    BACKGROUND: Unbiased flow cytometry-based methods have become the technique of choice in many laboratories for high-throughput, accurate assessments of malaria parasites in bioassays. A method to quantify live parasites based on mitotracker red CMXRos was recently described but consistent...... distinction of early ring stages of Plasmodium falciparum from uninfected red blood cells (uRBC) remains a challenge. METHODS: Here, a high-throughput, three-parameter (tri-colour) flow cytometry technique based on mitotracker red dye, the nucleic acid dye coriphosphine O (CPO) and the leucocyte marker CD45...... for enumerating live parasites in bioassays was developed. The technique was applied to estimate the specific growth inhibition index (SGI) in the antibody-dependent cellular inhibition (ADCI) assay and compared to parasite quantification by microscopy and mitotracker red staining. The Bland-Altman analysis...

  14. Frequency selective non-linear blending to improve image quality in liver CT

    International Nuclear Information System (INIS)

    Bongers, M.N.; Bier, G.; Kloth, C.; Schabel, C.; Nikolaou, K.; Horger, M.; Fritz, J.

    2016-01-01

    To evaluate the effects of a new frequency selective non-linear blending (NLB) algorithm on the contrast resolution of liver CT with low intravascular concentration of iodine contrast. Our local ethics committee approved this retrospective study. The informed consent requirement was waived. CT exams of 25 patients (60% female, mean age: 65±16 years of age) with late phase CT scans of the liver were included as a model for poor intrahepatic vascular contrast enhancement. Optimal post-processing settings to enhance the contrast of hepatic vessels were determined. Outcome variables included signal-to-noise (SNR) and contrast-to-noise ratios (CNR) of hepatic vessels and SNR of liver parenchyma of standard and post-processed images. Image quality was quantified by two independent readers using Likert scales. The post-processing settings for the visualization of hepatic vasculature were optimal at a center of 115HU, delta of 25HU, and slope of 5. Image noise was statistically indifferent between standard and post-processed images. The CNR between the hepatic vasculature (HV) and liver parenchyma could be significantly increased for liver veins (CNR Standard 1.62±1.10, CNR NLB 3.6±2.94, p=0.0002) and portal veins (CNR Standard 1.31±0.85, CNR NLB 2.42±3.03, p=0.046). The SNR of liver parenchyma was significantly higher on post-processed images (SNR NLB 11.26±3.16, SNR Standard 8.85± 2.27, p=0.008). The overall image quality and depiction of HV were significantly higher on post-processed images (NLB DHV : 4 [3-4.75], S tandardDHV : 2 [1.3-2.5], p=<0.0001; NLBIQ : 4 [4-4], StandardIQ : 2 [2-3], p=<0.0001). The use of a frequency selective non-linear blending algorithm increases the contrast resolution of liver CT and can improve the visibility of the hepatic vasculature in the setting of a low contrast ratio between vessels and the parenchyma.

  15. Frequency selective non-linear blending to improve image quality in liver CT

    Energy Technology Data Exchange (ETDEWEB)

    Bongers, M.N.; Bier, G.; Kloth, C.; Schabel, C.; Nikolaou, K.; Horger, M. [University Hospital of Tuebingen (Germany). Dept. of Diagnostic and Interventional Radiology; Fritz, J. [Johns Hopkins University School of Medicine, Baltimore, MD (United States). Russell H. Morgan Dept. of Radiology and Radiological Science

    2016-12-15

    To evaluate the effects of a new frequency selective non-linear blending (NLB) algorithm on the contrast resolution of liver CT with low intravascular concentration of iodine contrast. Our local ethics committee approved this retrospective study. The informed consent requirement was waived. CT exams of 25 patients (60% female, mean age: 65±16 years of age) with late phase CT scans of the liver were included as a model for poor intrahepatic vascular contrast enhancement. Optimal post-processing settings to enhance the contrast of hepatic vessels were determined. Outcome variables included signal-to-noise (SNR) and contrast-to-noise ratios (CNR) of hepatic vessels and SNR of liver parenchyma of standard and post-processed images. Image quality was quantified by two independent readers using Likert scales. The post-processing settings for the visualization of hepatic vasculature were optimal at a center of 115HU, delta of 25HU, and slope of 5. Image noise was statistically indifferent between standard and post-processed images. The CNR between the hepatic vasculature (HV) and liver parenchyma could be significantly increased for liver veins (CNR{sub Standard} 1.62±1.10, CNR{sub NLB} 3.6±2.94, p=0.0002) and portal veins (CNR{sub Standard} 1.31±0.85, CNR{sub NLB} 2.42±3.03, p=0.046). The SNR of liver parenchyma was significantly higher on post-processed images (SNR{sub NLB} 11.26±3.16, SNR{sub Standard} 8.85± 2.27, p=0.008). The overall image quality and depiction of HV were significantly higher on post-processed images (NLB{sub DHV}: 4 [3-4.75], S{sub tandardDHV}: 2 [1.3-2.5], p=<0.0001; {sub NLBIQ}: 4 [4-4], {sub StandardIQ}: 2 [2-3], p=<0.0001). The use of a frequency selective non-linear blending algorithm increases the contrast resolution of liver CT and can improve the visibility of the hepatic vasculature in the setting of a low contrast ratio between vessels and the parenchyma.

  16. Is image selection a useful strategy to decrease the transmission time in teleradiology? A study using 100 emergency cranial CTs

    International Nuclear Information System (INIS)

    Ludwig, K.; Bick, U.; Oelerich, M.; Schuierer, G.; Puskas, Z.; Nicolas, K.; Koch, A.; Lenzen, H.

    1998-01-01

    This study examines the suitability of working with a selection of images in a teleradiology consulting system in neurological or neurosurgical emergency situations. The teleradiology system was based on IBM-compatible personal computers, video digitization for data acquisition, and data transmission by Integrated System Digital Network. Forty normal and 60 abnormal emergency cranial computed tomograms were shown to a radiologist on call who presented all cases he regarded as pathologic to a neuroradiologic expert by teleradiology. To reduce transmission time, only a selection of images from the CT study was presented (up to four images per case). For each case the on-call radiologist's diagnosis (D on-call ), the expert's diagnosis on the teleradiology screen (D monitor ), and the expert's diagnosis on the original film (D original ) was documented, together with an estimation of the agreement between those diagnoses. There was clinically relevant disagreement between the on-call radiologist's diagnosis and the neuroradiologist's diagnosis based on the image selection on the teleradiology monitor in 23 % of cases. A clinically important discrepancy between the neuroradiologist's diagnosis based on the image selection and his diagnosis using the original films was found in 30 % of cases. This was due to the presence of clinically relevant information on images not transferred by the on-call radiologist. Image quality of the transferred images was sufficient in all cases. Drastic selection of images from a complete CT study leads to a high rate of incorrect diagnoses and is not appropriate to reduce transmission time in teleradiology. (orig.)

  17. Fluorescent genetic barcoding in mammalian cells for enhanced multiplexing capabilities in flow cytometry.

    Science.gov (United States)

    Smurthwaite, Cameron A; Hilton, Brett J; O'Hanlon, Ryan; Stolp, Zachary D; Hancock, Bryan M; Abbadessa, Darin; Stotland, Aleksandr; Sklar, Larry A; Wolkowicz, Roland

    2014-01-01

    The discovery of the green fluorescent protein from Aequorea victoria has revolutionized the field of cell and molecular biology. Since its discovery a growing panel of fluorescent proteins, fluorophores and fluorescent-coupled staining methodologies, have expanded the analytical capabilities of flow cytometry. Here, we exploit the power of genetic engineering to barcode individual cells with genes encoding fluorescent proteins. For genetic engineering, we utilize retroviral technology, which allows for the expression of ectopic genetic information in a stable manner in mammalian cells. We have genetically barcoded both adherent and nonadherent cells with different fluorescent proteins. Multiplexing power was increased by combining both the number of distinct fluorescent proteins, and the fluorescence intensity in each channel. Moreover, retroviral expression has proven to be stable for at least a 6-month period, which is critical for applications such as biological screens. We have shown the applicability of fluorescent barcoded multiplexing to cell-based assays that rely themselves on genetic barcoding, or on classical staining protocols. Fluorescent genetic barcoding gives the cell an inherited characteristic that distinguishes it from its counterpart. Once cell lines are developed, no further manipulation or staining is required, decreasing time, nonspecific background associated with staining protocols, and cost. The increasing number of discovered and/or engineered fluorescent proteins with unique absorbance/emission spectra, combined with the growing number of detection devices and lasers, increases multiplexing versatility, making fluorescent genetic barcoding a powerful tool for flow cytometry-based analysis. © 2013 International Society for Advancement of Cytometry.

  18. X-ray backscatter imaging for radiography by selective detection and snapshot: Evolution, development, and optimization

    Science.gov (United States)

    Shedlock, Daniel

    Compton backscatter imaging (CBI) is a single-sided imaging technique that uses the penetrating power of radiation and unique interaction properties of radiation with matter to image subsurface features. CBI has a variety of applications that include non-destructive interrogation, medical imaging, security and military applications. Radiography by selective detection (RSD), lateral migration radiography (LMR) and shadow aperture backscatter radiography (SABR) are different CBI techniques that are being optimized and developed. Radiography by selective detection (RSD) is a pencil beam Compton backscatter imaging technique that falls between highly collimated and uncollimated techniques. Radiography by selective detection uses a combination of single- and multiple-scatter photons from a projected area below a collimation plane to generate an image. As a result, the image has a combination of first- and multiple-scatter components. RSD techniques offer greater subsurface resolution than uncollimated techniques, at speeds at least an order of magnitude faster than highly collimated techniques. RSD scanning systems have evolved from a prototype into near market-ready scanning devices for use in a variety of single-sided imaging applications. The design has changed to incorporate state-of-the-art detectors and electronics optimized for backscatter imaging with an emphasis on versatility, efficiency and speed. The RSD system has become more stable, about 4 times faster, and 60% lighter while maintaining or improving image quality and contrast over the past 3 years. A new snapshot backscatter radiography (SBR) CBI technique, shadow aperture backscatter radiography (SABR), has been developed from concept and proof-of-principle to a functional laboratory prototype. SABR radiography uses digital detection media and shaded aperture configurations to generate near-surface Compton backscatter images without scanning, similar to how transmission radiographs are taken. Finally, a

  19. A CLIPS expert system for clinical flow cytometry data analysis

    Science.gov (United States)

    Salzman, G. C.; Duque, R. E.; Braylan, R. C.; Stewart, C. C.

    1990-01-01

    An expert system is being developed using CLIPS to assist clinicians in the analysis of multivariate flow cytometry data from cancer patients. Cluster analysis is used to find subpopulations representing various cell types in multiple datasets each consisting of four to five measurements on each of 5000 cells. CLIPS facts are derived from results of the clustering. CLIPS rules are based on the expertise of Drs. Stewart, Duque, and Braylan. The rules incorporate certainty factors based on case histories.

  20. Quantitative analysis of cellular glutathione by flow cytometry utilizing monochlorobimane: some applications to radiation and drug resistance in vitro and in vivo.

    Science.gov (United States)

    Rice, G C; Bump, E A; Shrieve, D C; Lee, W; Kovacs, M

    1986-12-01

    An assay using a bimane derivative has been developed to detect free glutathione (GSH) in individual viable cells by flow cytometry. Monochlorobimane [syn-(ClCH2CH3)-1,5-diazabicycla[3.30]acta-3,6-diene-2,8-dio ne], itself nonfluorescent, reacts with GSH to form a highly fluorescent derivative. High pressure liquid chromatography analysis showed that, using specific staining conditions, the only low molecular weight fluorescent derivative formed in Chinese hamster ovary cells was that formed with GSH. Very little reaction with protein sulfhydryls was observed. Rates of GSH depletion in Chinese hamster ovary cells exposed to diethylmaleate were essentially the same, whether measured by relative fluorescence intensity, by flow cytometry or by enzymatic assay on cellular extracts. This method was shown to be useful for measurement of GSH resynthesis, uptake, and depletion by prolonged hypoxia and misonidazole treatment. Since measurements are made on individual cells, cell-to-cell variation and populational heterogeneity in GSH content are revealed by flow cytometry. Although under most conditions in vitro GSH content is relatively homogeneous, under certain circumstances, such as release from hypoxia, heterogeneity in populational GSH levels was observed. The significance of this heterogeneity is discussed in regard to the induction of gene amplification and drug resistance by transient hypoxia. Numerous subclones of Chinese hamster ovary cells selected by growth in Adriamycin or methotrexate-containing medium express elevated levels of GSH per cell. The method was extended to quantitate the GSH content of cells excised from EMT-6/SF mouse tumors that had been treated in vivo with L-buthionine-S-R-sulfoximine, an inhibitor of GSH synthesis. The bivariate analysis (forward angle light scatter versus monochlorobimane fluorescence) of cells derived from these tumors gave excellent resolution of normal and tumor cells and demonstrated extensive heterogeneity in the tumor

  1. Registration for Optical Multimodal Remote Sensing Images Based on FAST Detection, Window Selection, and Histogram Specification

    Directory of Open Access Journals (Sweden)

    Xiaoyang Zhao

    2018-04-01

    Full Text Available In recent years, digital frame cameras have been increasingly used for remote sensing applications. However, it is always a challenge to align or register images captured with different cameras or different imaging sensor units. In this research, a novel registration method was proposed. Coarse registration was first applied to approximately align the sensed and reference images. Window selection was then used to reduce the search space and a histogram specification was applied to optimize the grayscale similarity between the images. After comparisons with other commonly-used detectors, the fast corner detector, FAST (Features from Accelerated Segment Test, was selected to extract the feature points. The matching point pairs were then detected between the images, the outliers were eliminated, and geometric transformation was performed. The appropriate window size was searched and set to one-tenth of the image width. The images that were acquired by a two-camera system, a camera with five imaging sensors, and a camera with replaceable filters mounted on a manned aircraft, an unmanned aerial vehicle, and a ground-based platform, respectively, were used to evaluate the performance of the proposed method. The image analysis results showed that, through the appropriate window selection and histogram specification, the number of correctly matched point pairs had increased by 11.30 times, and that the correct matching rate had increased by 36%, compared with the results based on FAST alone. The root mean square error (RMSE in the x and y directions was generally within 0.5 pixels. In comparison with the binary robust invariant scalable keypoints (BRISK, curvature scale space (CSS, Harris, speed up robust features (SURF, and commercial software ERDAS and ENVI, this method resulted in larger numbers of correct matching pairs and smaller, more consistent RMSE. Furthermore, it was not necessary to choose any tie control points manually before registration

  2. Fiber array based hyperspectral Raman imaging for chemical selective analysis of malaria-infected red blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Brückner, Michael [Leibniz Institute of Photonic Technology, 07745 Jena (Germany); Becker, Katja [Justus Liebig University Giessen, Biochemistry and Molecular Biology, 35392 Giessen (Germany); Popp, Jürgen [Leibniz Institute of Photonic Technology, 07745 Jena (Germany); Friedrich Schiller University Jena, Institute for Physical Chemistry, 07745 Jena (Germany); Friedrich Schiller University Jena, Abbe Centre of Photonics, 07745 Jena (Germany); Frosch, Torsten, E-mail: torsten.frosch@uni-jena.de [Leibniz Institute of Photonic Technology, 07745 Jena (Germany); Friedrich Schiller University Jena, Institute for Physical Chemistry, 07745 Jena (Germany); Friedrich Schiller University Jena, Abbe Centre of Photonics, 07745 Jena (Germany)

    2015-09-24

    A new setup for Raman spectroscopic wide-field imaging is presented. It combines the advantages of a fiber array based spectral translator with a tailor-made laser illumination system for high-quality Raman chemical imaging of sensitive biological samples. The Gaussian-like intensity distribution of the illuminating laser beam is shaped by a square-core optical multimode fiber to a top-hat profile with very homogeneous intensity distribution to fulfill the conditions of Koehler. The 30 m long optical fiber and an additional vibrator efficiently destroy the polarization and coherence of the illuminating light. This homogeneous, incoherent illumination is an essential prerequisite for stable quantitative imaging of complex biological samples. The fiber array translates the two-dimensional lateral information of the Raman stray light into separated spectral channels with very high contrast. The Raman image can be correlated with a corresponding white light microscopic image of the sample. The new setup enables simultaneous quantification of all Raman spectra across the whole spatial area with very good spectral resolution and thus outperforms other Raman imaging approaches based on scanning and tunable filters. The unique capabilities of the setup for fast, gentle, sensitive, and selective chemical imaging of biological samples were applied for automated hemozoin analysis. A special algorithm was developed to generate Raman images based on the hemozoin distribution in red blood cells without any influence from other Raman scattering. The new imaging setup in combination with the robust algorithm provides a novel, elegant way for chemical selective analysis of the malaria pigment hemozoin in early ring stages of Plasmodium falciparum infected erythrocytes. - Highlights: • Raman hyperspectral imaging allows for chemical selective analysis of biological samples with spatial heterogeneity. • A homogeneous, incoherent illumination is essential for reliable

  3. Fiber array based hyperspectral Raman imaging for chemical selective analysis of malaria-infected red blood cells

    International Nuclear Information System (INIS)

    Brückner, Michael; Becker, Katja; Popp, Jürgen; Frosch, Torsten

    2015-01-01

    A new setup for Raman spectroscopic wide-field imaging is presented. It combines the advantages of a fiber array based spectral translator with a tailor-made laser illumination system for high-quality Raman chemical imaging of sensitive biological samples. The Gaussian-like intensity distribution of the illuminating laser beam is shaped by a square-core optical multimode fiber to a top-hat profile with very homogeneous intensity distribution to fulfill the conditions of Koehler. The 30 m long optical fiber and an additional vibrator efficiently destroy the polarization and coherence of the illuminating light. This homogeneous, incoherent illumination is an essential prerequisite for stable quantitative imaging of complex biological samples. The fiber array translates the two-dimensional lateral information of the Raman stray light into separated spectral channels with very high contrast. The Raman image can be correlated with a corresponding white light microscopic image of the sample. The new setup enables simultaneous quantification of all Raman spectra across the whole spatial area with very good spectral resolution and thus outperforms other Raman imaging approaches based on scanning and tunable filters. The unique capabilities of the setup for fast, gentle, sensitive, and selective chemical imaging of biological samples were applied for automated hemozoin analysis. A special algorithm was developed to generate Raman images based on the hemozoin distribution in red blood cells without any influence from other Raman scattering. The new imaging setup in combination with the robust algorithm provides a novel, elegant way for chemical selective analysis of the malaria pigment hemozoin in early ring stages of Plasmodium falciparum infected erythrocytes. - Highlights: • Raman hyperspectral imaging allows for chemical selective analysis of biological samples with spatial heterogeneity. • A homogeneous, incoherent illumination is essential for reliable

  4. An observer study comparing spot imaging regions selected by radiologists and a computer for an automated stereo spot mammography technique

    International Nuclear Information System (INIS)

    Goodsitt, Mitchell M.; Chan, Heang-Ping; Lydick, Justin T.; Gandra, Chaitanya R.; Chen, Nelson G.; Helvie, Mark A.; Bailey, Janet E.; Roubidoux, Marilyn A.; Paramagul, Chintana; Blane, Caroline E.; Sahiner, Berkman; Petrick, Nicholas A.

    2004-01-01

    We are developing an automated stereo spot mammography technique for improved imaging of suspicious dense regions within digital mammograms. The technique entails the acquisition of a full-field digital mammogram, automated detection of a suspicious dense region within that mammogram by a computer aided detection (CAD) program, and acquisition of a stereo pair of images with automated collimation to the suspicious region. The latter stereo spot image is obtained within seconds of the original full-field mammogram, without releasing the compression paddle. The spot image is viewed on a stereo video display. A critical element of this technique is the automated detection of suspicious regions for spot imaging. We performed an observer study to compare the suspicious regions selected by radiologists with those selected by a CAD program developed at the University of Michigan. True regions of interest (TROIs) were separately determined by one of the radiologists who reviewed the original mammograms, biopsy images, and histology results. We compared the radiologist and computer-selected regions of interest (ROIs) to the TROIs. Both the radiologists and the computer were allowed to select up to 3 regions in each of 200 images (mixture of 100 CC and 100 MLO views). We computed overlap indices (the overlap index is defined as the ratio of the area of intersection to the area of interest) to quantify the agreement between the selected regions in each image. The averages of the largest overlap indices per image for the 5 radiologist-to-computer comparisons were directly related to the average number of regions per image traced by the radiologists (about 50% for 1 region/image, 84% for 2 regions/image and 96% for 3 regions/image). The average of the overlap indices with all of the TROIs was 73% for CAD and 76.8%+/-10.0% for the radiologists. This study indicates that the CAD determined ROIs could potentially be useful for a screening technique that includes stereo spot

  5. Assessing the Diagnostic Imaging needs for Five Selected Hospitals in Uganda

    Directory of Open Access Journals (Sweden)

    Michael G Kawooya

    2011-01-01

    Full Text Available Introduction: Uganda has limited health resources. It is important to measure the need for imaging in order to set policy and plan for imaging services. Objectives: The first specific objective was to develop and apply four imaging needs indices on a case study basis, in five selected Ugandan hospitals. The indices were: Imaging Load (IL, Imaging Burden (IB, Type Specific Imaging Burden (TSIB, and Disease Specific Imaging Burden (DSIB. The second objective was to explore the perceptions of the patient, referring clinician, and radiologist regarding the values, meaning, and objective of imaging in patient care. Materials and Methods: This was a cross-sectional survey employing triangulation methodology, conducted in 5 Ugandan hospitals over a period of 3 years during 2005 - 2008. The subjects were divided into four clusters: Obstetrics and gynecology (obs/gynae, surgery, internal medicine, and pediatrics. For the quantitative component of the study, data from case notes was used to calculate the indices. The qualitative component explored the non-measurable aspects of imaging needs from the clinician′s, radiologist′s, and patient′s perspective. Results: A total of 1961 patient case notes were studied. The IB was 460 per 1000 hospital patients per year. The highest TSIB was for ultrasound at 232 per 1000 hospital patients per year, followed by 191 patients for general X-ray. The majority of the patients interviewed had special desires, expectations, and misconceptions. Conclusions: There is a high IB of 460 per thousand patient populations per year, mainly due to ultrasound. The majority of the patients have perceptions, misconceptions, beliefs, and values which influence the need for imaging. There is a need to address the medical and non-tangible imaging needs of the patient and to counteract imaging-related misconceptions and over-expectations. Public awareness of the value, capabilities, limitations, and adverse effects of various

  6. Variability-based active galactic nucleus selection using image subtraction in the SDSS and LSST era

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Yumi; Gibson, Robert R.; Becker, Andrew C.; Ivezić, Željko; Connolly, Andrew J.; Ruan, John J.; Anderson, Scott F. [Department of Astronomy, University of Washington, Box 351580, Seattle, WA 98195 (United States); MacLeod, Chelsea L., E-mail: ymchoi@astro.washington.edu [Physics Department, U.S. Naval Academy, 572 Holloway Road, Annapolis, MD 21402 (United States)

    2014-02-10

    With upcoming all-sky surveys such as LSST poised to generate a deep digital movie of the optical sky, variability-based active galactic nucleus (AGN) selection will enable the construction of highly complete catalogs with minimum contamination. In this study, we generate g-band difference images and construct light curves (LCs) for QSO/AGN candidates listed in Sloan Digital Sky Survey Stripe 82 public catalogs compiled from different methods, including spectroscopy, optical colors, variability, and X-ray detection. Image differencing excels at identifying variable sources embedded in complex or blended emission regions such as Type II AGNs and other low-luminosity AGNs that may be omitted from traditional photometric or spectroscopic catalogs. To separate QSOs/AGNs from other sources using our difference image LCs, we explore several LC statistics and parameterize optical variability by the characteristic damping timescale (τ) and variability amplitude. By virtue of distinguishable variability parameters of AGNs, we are able to select them with high completeness of 93.4% and efficiency (i.e., purity) of 71.3%. Based on optical variability, we also select highly variable blazar candidates, whose infrared colors are consistent with known blazars. One-third of them are also radio detected. With the X-ray selected AGN candidates, we probe the optical variability of X-ray detected optically extended sources using their difference image LCs for the first time. A combination of optical variability and X-ray detection enables us to select various types of host-dominated AGNs. Contrary to the AGN unification model prediction, two Type II AGN candidates (out of six) show detectable variability on long-term timescales like typical Type I AGNs. This study will provide a baseline for future optical variability studies of extended sources.

  7. Analysis of nuclear localization of interleukin-1 family cytokines by flow cytometry.

    Science.gov (United States)

    Ross, Ralf; Grimmel, Jan; Goedicke, Sybelle; Möbus, Anna M; Bulau, Ana-Maria; Bufler, Philip; Ali, Shafaqat; Martin, Michael U

    2013-01-31

    The dual function cytokines IL-1α, IL-33 and IL-37 are members of the IL-1 cytokine family. Besides of being able to bind to their cognate receptors on target cells, they can act intracellularly in the producing cell. All three are able to translocate to the nucleus and have been discussed to affect gene expression. In order to compare and quantitate nuclear translocation of these IL-1 family members we established a robust technique which enables to measure nuclear localization on a single cell level by flow cytometry. Vectors encoding fusion proteins of different IL-1 family members with enhanced green fluorescent protein were cloned and cell lines transiently transfected with these. Fluorescent fusion proteins in intact cells or in isolated nuclei were detected subsequently by fluorescence microscopy and flow cytometry, respectively. Depending on the cellular system, cells and nuclei were distinguishable by flow cytometry in forward scatter/sideward scatter. Fluorescent fusion proteins were detectable in isolated nuclei up to three days following preparation. Signal intensity of fusion proteins of IL-33 and IL-37 in isolated nuclei but not of IL-1α, was markedly increased by fixation with paraformaldehyde, directly following cell lysis, indicating that IL-1α binds stronger to nuclear structures than IL-33 and IL-37. Nuclear translocation of fluorescent IL-37 fusion proteins in a stably transfected RAW264.7 mouse macrophage cell line required stimulation with lipopolysaccharide. Applying this method we demonstrated that a prolonged lag phase of more than 15h before LPS-stimulated nuclear translocation was detected. In summary, we present a robust method to analyze and quantitate nuclear localization of IL-1 cytokine family members. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Sinularin Selectively Kills Breast Cancer Cells Showing G2/M Arrest, Apoptosis, and Oxidative DNA Damage

    Directory of Open Access Journals (Sweden)

    Hurng-Wern Huang

    2018-04-01

    Full Text Available The natural compound sinularin, isolated from marine soft corals, is antiproliferative against several cancers, but its possible selective killing effect has rarely been investigated. This study investigates the selective killing potential and mechanisms of sinularin-treated breast cancer cells. In 3-(4,5-dimethylthiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H- tetrazolium, inner salt (MTS assay, sinularin dose-responsively decreased the cell viability of two breast cancer (SKBR3 and MDA-MB-231 cells, but showed less effect on breast normal (M10 cells after a 24 h treatment. According to 7-aminoactinomycin D (7AAD flow cytometry, sinularin dose-responsively induced the G2/M cycle arrest of SKBR3 cells. Sinularin dose-responsively induced apoptosis on SKBR3 cells in terms of a flow cytometry-based annexin V/7AAD assay and pancaspase activity, as well as Western blotting for cleaved forms of poly(ADP-ribose polymerase (PARP, caspases 3, 8, and 9. These caspases and PARP activations were suppressed by N-acetylcysteine (NAC pretreatment. Moreover, sinularin dose-responsively induced oxidative stress and DNA damage according to flow cytometry analyses of reactive oxygen species (ROS, mitochondrial membrane potential (MitoMP, mitochondrial superoxide, and 8-oxo-2′-deoxyguanosine (8-oxodG. In conclusion, sinularin induces selective killing, G2/M arrest, apoptosis, and oxidative DNA damage of breast cancer cells.

  9. Selective detection of Escherichia coli by imaging of the light intensity transmitted through an optical disk

    Science.gov (United States)

    Shiramizu, Hideyuki; Kuroda, Chiaki; Ohki, Yoshimichi; Shima, Takayuki; Wang, Xiaomin; Fujimaki, Makoto

    2018-03-01

    We have developed an optical disk system for imaging transmitted light from Escherichia coli dispersed on an optical disk. When E. coli was stained using Bismarck brown, the transmittance was found to decrease in images obtained at λ = 405 nm. The results indicate that transmittance imaging is suitable for finding the difference in light intensity between stained and unstained E. coli, whereas the reflectance images were scarcely changed by staining. Therefore, E. coli can be selectively discriminated from abiotic contaminants using transmittance imaging.

  10. DNA index determination with Automated Cellular Imaging System (ACIS in Barrett's esophagus: Comparison with CAS 200

    Directory of Open Access Journals (Sweden)

    Klein Michael

    2005-08-01

    Full Text Available Abstract Background For solid tumors, image cytometry has been shown to be more sensitive for diagnosing DNA content abnormalities (aneuploidy than flow cytometry. Image cytometry has often been performed using the semi-automated CAS 200 system. Recently, an Automated Cellular Imaging System (ACIS was introduced to determine DNA content (DNA index, but it has not been validated. Methods Using the CAS 200 system and ACIS, we compared the DNA index (DI obtained from the same archived formalin-fixed and paraffin embedded tissue samples from Barrett's esophagus related lesions, including samples with specialized intestinal metaplasia without dysplasia, low-grade dysplasia, high-grade dysplasia and adenocarcinoma. Results Although there was a very good correlation between the DI values determined by ACIS and CAS 200, the former was 25% more sensitive in detecting aneuploidy. ACIS yielded a mean DI value 18% higher than that obtained by CAS 200 (p t test. In addition, the average time required to perform a DNA ploidy analysis was shorter with the ACIS (30–40 min than with the CAS 200 (40–70 min. Results obtained by ACIS gave excellent inter-and intra-observer variability (coefficient of correlation >0.9 for both, p Conclusion Compared with the CAS 200, the ACIS is a more sensitive and less time consuming technique for determining DNA ploidy. Results obtained by ACIS are also highly reproducible.

  11. Using flow cytometry for counting natural planktonic bacteria and understanding the structure of planktonic bacterial communities

    Directory of Open Access Journals (Sweden)

    Josep M. Gasol

    2000-06-01

    Full Text Available Flow cytometry is rapidly becoming a routine methodology in aquatic microbial ecology. The combination of simple to use bench-top flow cytometers and highly fluorescent nucleic acid stains allows fast and easy determination of microbe abundance in the plankton of lakes and oceans. The different dyes and protocols used to stain and count planktonic bacteria as well as the equipment in use are reviewed, with special attention to some of the problems encountered in daily routine practice such as fixation, staining and absolute counting. One of the main advantages of flow cytometry over epifluorescence microscopy is the ability to obtain cell-specific measurements in large numbers of cells with limited effort. We discuss how this characteristic has been used for differentiating photosynthetic from non-photosynthetic prokaryotes, for measuring bacterial cell size and nucleic acid content, and for estimating the relative activity and physiological state of each cell. We also describe how some of the flow cytometrically obtained data can be used to characterize the role of microbes on carbon cycling in the aquatic environment and we prospect the likely avenues of progress in the study of planktonic prokaryotes through the use of flow cytometry.

  12. Reticulocyte analysis using flow cytometry.

    Science.gov (United States)

    Corberand, J X

    1996-12-01

    Automation of the reticulocyte count by means of flow cytometry has considerably improved the quality of this investigation. This article deals firstly with the reasons for the poor performance of the microscopic technique and with the physiological principles underlying identification and classification of reticulocytes using RNA labeling. It then outlines the automated methods currently on the market, which can be classified in three categories: a) "general-purpose" cytofluorometers, which in clinical laboratories usually deal with lymphocyte immunophenotyping; b) the only commercially available cytofluorometer dedicated to the reticulocyte count; this automat has the advantage of requiring no human intervention as it merely needs to be fed with samples; c) hematology analyzers with specific modules for automatic counting of reticulocytes previously incubated with a non-fluorescent dye. Of the various fluorescent markers available, thiazole orange, DEQTC iodide and auramine are most often used for this basic hematology test. The quality of the count, the availability of new reticulocyte indices (maturation index, percentage of young reticulocytes) and rapidity of the count give this test renewed value in the practical approach to the diagnosis of anemia, and also open new perspectives in the surveillance of aplastic anemia after chemotherapy or bone marrow grafting.

  13. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity.

    Science.gov (United States)

    Wang, Deng-Liang; Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan; Yang, Hai-Tao; Wang, Jiang-Jie; Yao, Pei-Sen; Pan, Ru-Jun; Yang, Chaoyong James; Kang, De-Zhi

    2014-10-31

    Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher's attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with Kd 56±7.3nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Detection of endogenous alkaline phosphatase activity in intact cells by flow cytometry using the fluorogenic ELF-97 phosphatase substrate

    Science.gov (United States)

    Telford, W. G.; Cox, W. G.; Stiner, D.; Singer, V. L.; Doty, S. B.

    1999-01-01

    BACKGROUND: The alkaline phosphatase (AP) substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-(3H)-quinazolinone (ELF((R))-97 for enzyme-labeled fluorescence) has been found useful for the histochemical detection of endogenous AP activity and AP-tagged proteins and oligonucleotide probes. In this study, we evaluated its effectiveness at detecting endogenous AP activity by flow cytometry. METHODS: The ELF-97 phosphatase substrate was used to detect endogenous AP activity in UMR-106 rat osteosarcoma cells and primary cultures of chick chondrocytes. Cells were labeled with the ELF-97 reagent and analyzed by flow cytometry using an argon ultraviolet (UV) laser. For comparison purposes, cells were also assayed for AP using a Fast Red Violet LB azo dye assay previously described for use in detecting AP activity by flow cytometry. RESULTS: The ELF-97 phosphatase substrate effectively detected endogenous AP activity in UMR-106 cells, with over 95% of the resulting fluorescent signal resulting from AP-specific activity (as determined by levamisole inhibition of AP activity). In contrast, less than 70% of the fluorescent signal from the Fast Red Violet LB (FRV) assay was AP-dependent, reflecting the high intrinsic fluorescence of the unreacted components. The ELF-97 phosphatase assay was also able to detect very low AP activity in chick chondrocytes that was undetectable by the azo dye method. CONCLUSIONS: The ELF-97 phosphatase assay was able to detect endogenous AP activity in fixed mammalian and avian cells by flow cytometry with superior sensitivity to previously described assays. This work also shows the applicability of ELF-97 to flow cytometry, supplementing its previously demonstrated histochemical applications. Copyright 1999 Wiley-Liss, Inc.

  15. Body image concern and selective attention to disgusting and non-self appearance-related stimuli.

    Science.gov (United States)

    Onden-Lim, Melissa; Wu, Ray; Grisham, Jessica R

    2012-09-01

    Although selective attention to one's own appearance has been widely documented in studies of body dysmorphic disorder (BDD), little is known about attentional bias toward non-self appearance-related stimuli in BDD. Furthermore, despite reports of heightened experience of disgust in BDD, it is unknown whether these individuals differentially attend to disgusting stimuli and whether disgust is important in processing of unattractive stimuli. We used a dot probe procedure to investigate the relationship between dysmorphic concern, a defining feature of BDD, and selective attention to faces, attractive, unattractive and disgusting images in a female heterosexual student population (N=92). At the long stimulus presentation (1000 ms), dysmorphic concern was positively associated with attention to faces in general and attractive appearance-related images. In contrast, at the short stimulus presentation (200 ms), there was a positive association between dysmorphic concern and disgusting images. Implications for theoretical models of BDD are discussed. Copyright © 2012. Published by Elsevier Ltd.

  16. Bleaching response of Symbiodinium (zooxanthellae): determination by flow cytometry.

    Science.gov (United States)

    Lee, Co Sin; Yeo, Yin Sheng Wilson; Sin, Tsai Min

    2012-10-01

    Coral bleaching is of increasing concern to reef management and stakeholders. Thus far, quantification of coral bleaching tends to be heavily reliant on the enumeration of zooxanthellae, with less emphasis on assessment of photosynthetic or physiological condition, these being often assessed separately by techniques such as liquid chromatography. Traditional methods of enumeration using microscopy are time consuming, subjected to low precision and great observer error. In this study, we presented a method for the distinction of physoiological condition and rapid enumeration of zooxanthellae using flow cytometry (FCM). Microscopy verified that healthy looking/live versus damaged/dead zooxanthellae could be reliably and objectively distinguished and counted by FCM on the basis of red and green fluorescence and light scatter. Excellent correlations were also determined between FCM and microscopy estimates of cell concentrations of fresh zooxanthellae isolates from Pocillopora damicornis. The relative intensities of chlorophyll and β-carotene fluorescences were shown to be important in understanding the results of increased cell counts in freshly isolated zooxanthellae experimentally exposed to high temperatures (34, 36, and 38°C) over 24 h, with ambient temperature (29°C) used as controls. The ability to simultaneously identify and enumerate subpopulations of different physiological states in the same sample provides an enormous advantage in not just determining bleaching responses, but elucidating adaptive response and mechanisms for tolerance. Therefore, this approach might provide a rapid, convenient, and reproducible methodology for climate change studies and reef management programs. Copyright © 2012 International Society for Advancement of Cytometry.

  17. Contrast based band selection for optimized weathered oil detection in hyperspectral images

    Science.gov (United States)

    Levaux, Florian; Bostater, Charles R., Jr.; Neyt, Xavier

    2012-09-01

    Hyperspectral imagery offers unique benefits for detection of land and water features due to the information contained in reflectance signatures such as the bi-directional reflectance distribution function or BRDF. The reflectance signature directly shows the relative absorption and backscattering features of targets. These features can be very useful in shoreline monitoring or surveillance applications, for example to detect weathered oil. In real-time detection applications, processing of hyperspectral data can be an important tool and Optimal band selection is thus important in real time applications in order to select the essential bands using the absorption and backscatter information. In the present paper, band selection is based upon the optimization of target detection using contrast algorithms. The common definition of the contrast (using only one band out of all possible combinations available within a hyperspectral image) is generalized in order to consider all the possible combinations of wavelength dependent contrasts using hyperspectral images. The inflection (defined here as an approximation of the second derivative) is also used in order to enhance the variations in the reflectance spectra as well as in the contrast spectrua in order to assist in optimal band selection. The results of the selection in term of target detection (false alarms and missed detection) are also compared with a previous method to perform feature detection, namely the matched filter. In this paper, imagery is acquired using a pushbroom hyperspectral sensor mounted at the bow of a small vessel. The sensor is mechanically rotated using an optical rotation stage. This opto-mechanical scanning system produces hyperspectral images with pixel sizes on the order of mm to cm scales, depending upon the distance between the sensor and the shoreline being monitored. The motion of the platform during the acquisition induces distortions in the collected HSI imagery. It is therefore

  18. An inter-laboratory comparison of PNH clone detection by high-sensitivity flow cytometry in a Russian cohort.

    Science.gov (United States)

    Sipol, Alexandra A; Babenko, Elena V; Borisov, Vyacheslav I; Naumova, Elena V; Boyakova, Elena V; Yakunin, Dimitry I; Glazanova, Tatyana V; Chubukina, Zhanna V; Pronkina, Natalya V; Popov, Alexander M; Saveliev, Leonid I; Lugovskaya, Svetlana A; Lisukov, Igor A; Kulagin, Alexander D; Illingworth, Andrea J

    2015-01-01

    Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal stem cell disorder characterized by partial or absolute deficiency of glycophosphatidyl-inositol (GPI) anchor-linked surface proteins on blood cells. A lack of precise diagnostic standards for flow cytometry has hampered useful comparisons of data between laboratories. We report data from the first study evaluating the reproducibility of high-sensitivity flow cytometry for PNH in Russia. PNH clone sizes were determined at diagnosis in PNH patients at a central laboratory and compared with follow-up measurements in six laboratories across the country. Analyses in each laboratory were performed according to recommendations from the International Clinical Cytometry Society (ICCS) and the more recent 'practical guidelines'. Follow-up measurements were compared with each other and with the values determined at diagnosis. PNH clone size measurements were determined in seven diagnosed PNH patients (five females, two males: mean age 37 years); five had a history of aplastic anemia and three (one with and two without aplastic anemia) had severe hemolytic PNH and elevated plasma lactate dehydrogenase. PNH clone sizes at diagnosis were low in patients with less severe clinical symptoms (0.41-9.7% of granulocytes) and high in patients with severe symptoms (58-99%). There were only minimal differences in the follow-up clone size measurement for each patient between the six laboratories, particularly in those with high values at diagnosis. The ICCS-recommended high-sensitivity flow cytometry protocol was effective for detecting major and minor PNH clones in Russian PNH patients, and showed high reproducibility between laboratories.

  19. Imaging Primary Mouse Sarcomas After Radiation Therapy Using Cathepsin-Activatable Fluorescent Imaging Agents

    Energy Technology Data Exchange (ETDEWEB)

    Cuneo, Kyle C. [Department of Radiation Oncology, Duke University School of Medicine, Durham, North Carolina (United States); Mito, Jeffrey K.; Javid, Melodi P. [Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, North Carolina (United States); Ferrer, Jorge M. [Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Kim, Yongbaek [Department of Clinical Pathology, College of Veterinary Medicine, Seoul National University, Seoul (Korea, Republic of); Lee, W. David [The David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Bawendi, Moungi G. [Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Brigman, Brian E. [Department of Orthopedic Surgery, Duke University School of Medicine, Durham, North Carolina (United States); Kirsch, David G., E-mail: david.kirsch@duke.edu [Department of Radiation Oncology, Duke University School of Medicine, Durham, North Carolina (United States); Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, North Carolina (United States)

    2013-05-01

    Purpose: Cathepsin-activated fluorescent probes can detect tumors in mice and in canine patients. We previously showed that these probes can detect microscopic residual sarcoma in the tumor bed of mice during gross total resection. Many patients with soft tissue sarcoma (STS) and other tumors undergo radiation therapy (RT) before surgery. This study assesses the effect of RT on the ability of cathepsin-activated probes to differentiate between normal and cancerous tissue. Methods and Materials: A genetically engineered mouse model of STS was used to generate primary hind limb sarcomas that were treated with hypofractionated RT. Mice were injected intravenously with cathepsin-activated fluorescent probes, and various tissues, including the tumor, were imaged using a hand-held imaging device. Resected tumor and normal muscle samples were harvested to assess cathepsin expression by Western blot. Uptake of activated probe was analyzed by flow cytometry and confocal microscopy. Parallel in vitro studies using mouse sarcoma cells were performed. Results: RT of primary STS in mice and mouse sarcoma cell lines caused no change in probe activation or cathepsin protease expression. Increasing radiation dose resulted in an upward trend in probe activation. Flow cytometry and immunofluorescence showed that a substantial proportion of probe-labeled cells were CD11b-positive tumor-associated immune cells. Conclusions: In this primary murine model of STS, RT did not affect the ability of cathepsin-activated probes to differentiate between tumor and normal muscle. Cathepsin-activated probes labeled tumor cells and tumor-associated macrophages. Our results suggest that it would be feasible to include patients who have received preoperative RT in clinical studies evaluating cathepsin-activated imaging probes.

  20. Selective interferometric imaging of internal multiples

    KAUST Repository

    Zuberi, M. A H

    2013-01-01

    Internal multiples deteriorate the image when the imaging procedure assumes only single scattering, especially if the velocity model does not reproduce such scattering in the Green’s function. If properly imaged, internal multiples (and internally-scattered energy) can enhance the seismic image and illuminate areas otherwise neglected or poorly imaged by conventional single-scattering approaches. Conventionally, in order to image internal multiples, accurate, sharp contrasts in the velocity model are required to construct a Green’s function with all the scattered energy. As an alternative, we develop a three-step procedure, which images the first-order internal scattering using the background Green’s function (from the surface to each image point), constructed from a smooth velocity model: We first back-propagate the recorded surface data using the background Green’s function, then cross-correlate the back-propagated data with the recorded data and finally cross-correlate the result with the original background Green’s function. This procedure images the contribution of the recorded first-order internal multiples and is almost free of the single-scattering recorded energy. This image can be added to the conventional single-scattering image, obtained e.g. from Kirchhoff migration, to enhance the image. Application to synthetic data with reflectors illuminated by multiple scattering only demonstrates the effectiveness of the approach.

  1. Evaluation of cell proliferative activity after irradiation using immunohistochemical approach and flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Tamada, Takashi (Okayama Univ. (Japan). School of Medicine)

    1992-06-01

    To evaluate a proliferative activity of post-irradiated malignant cells, we studied the kinetics of HeLa cells using immunohistochemical approach and flow cytometry. HeLa cells were stained with two proliferation-associated monoclonal antibodies, Ki-67 and anti-DNA polymerase {alpha} antibody. Nucleoli of non-irradiated cells were granularly stained with Ki-67. After irradiation, only the center of nuclei was diffusely stained with Ki-67. One hundred forty-four hours after low-dose irradiation, the staining patterns became the same as the control. On the other hand, after high-dose irradiation, the center of nuclei was weakly stained. DNA polymerase {alpha} was diffusely labelled with nuclei of the control. It was located around the border of nuclei of low-dose irradiated cells like a ring. But after high-dose irradiation, it was granularly distributed in the periphery of nuclei. FITC conjugated Ki-67/PI two parameter analysis was done by a single laser flow cytometer. Twenty-four hours after irradiation, DNA-histograms showed the accumulation to G{sub 2}/M phase and the increase of DNA content of G{sub 2}/M cells, as exposure dose was increased. Two parameter analysis showed the increase of FITC uptake of G{sub 2}/M phase as dose increased. These changes of flow cytometry were remarkably observed after 24 hours' incubation. It was shown that the difference of Ki-67 antigen and DNA polymerase {alpha} appearance depended on the irradiation dose. These findings suggest that immunohistochemical staining with Ki-67 or anti-DNA polymerase {alpha} antibody and flow cytometry using Ki-67 are available to evaluate cell damages after irradiation. (author).

  2. Comparison of BALB/c and CBA/J mice for the local lymph node assay using bromodeoxyuridine with flow cytometry (LLNA: BrdU-FCM).

    Science.gov (United States)

    Lee, Yong Sun; Yi, Jung-Sun; Seo, Souk Jin; Kim, Joo Hwan; Jung, Mi-Sook; Seo, Im-Kwon; Ahn, Ilyoung; Ko, Kyungyuk; Kim, Tae Sung; Lim, Kyung Min; Sohn, Soojung

    2017-02-01

    The local lymph node assay using 5-bromo-2-deoxyuridine (BrdU) with flow cytometry (LLNA: BrdU-FCM) is a modified LLNA that is used to identify skin sensitizers by counting BrdU-incorporated lymph node cells (LNCs) with flow cytometry. Unlike other LLNA methods (OECD TG 429, 442A and 442B) in which the CBA/J mouse strain is used, LLNA: BrdU-FCM was originally designed to be compatible with BALB/c, a mouse strain that is more widely used in many countries. To justify the substitution of CBA/J for BALB/c, the equivalence of the test results between two strains shall be established prior to the official implementation of LLNA: BrdU-FCM. This study aims to compare the test results of LLNA: BrdU-FCM produced in BALB/c mice with those in CBA/J mice for 18 reference substances, including 13 sensitizers and 5 non-sensitizers, listed in OECD Test Guideline 429. Based on the LLNA: BrdU-FCM test procedure, we selected an appropriate solvent and then performed preliminary tests to determine the non-irritating dose ranges for the main study, which revealed the difference in the irritation responses to 8 of the 18 chemicals between the two strains. In the main study, we measured the changes in the number of total LNCs, which indicated differences in the responses to test chemicals between the two strains. However, the stimulation index obtained with the counts of BrdU-incorporated LNCs with 7-AAD using flow cytometry yielded comparable results and 100% concordance between the BALB/c and CBA/J mouse strains was achieved, suggesting that the performance of LLNA: BrdU-FCM using BALB/c mice was equivalent to that with CBA/J mice. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. A COMPARISON STUDY OF DIFFERENT MARKER SELECTION METHODS FOR SPECTRAL-SPATIAL CLASSIFICATION OF HYPERSPECTRAL IMAGES

    Directory of Open Access Journals (Sweden)

    D. Akbari

    2015-12-01

    Full Text Available An effective approach based on the Minimum Spanning Forest (MSF, grown from automatically selected markers using Support Vector Machines (SVM, has been proposed for spectral-spatial classification of hyperspectral images by Tarabalka et al. This paper aims at improving this approach by using image segmentation to integrate the spatial information into marker selection process. In this study, the markers are extracted from the classification maps, obtained by both SVM and segmentation algorithms, and then are used to build the MSF. The segmentation algorithms are the watershed, expectation maximization (EM and hierarchical clustering. These algorithms are used in parallel and independently to segment the image. Moreover, the pixels of each class, with the largest population in the classification map, are kept for each region of the segmentation map. Lastly, the most reliable classified pixels are chosen from among the exiting pixels as markers. Two benchmark urban hyperspectral datasets are used for evaluation: Washington DC Mall and Berlin. The results of our experiments indicate that, compared to the original MSF approach, the marker selection using segmentation algorithms leads in more accurate classification maps.

  4. Feature selection from a facial image for distinction of sasang constitution.

    Science.gov (United States)

    Koo, Imhoi; Kim, Jong Yeol; Kim, Myoung Geun; Kim, Keun Ho

    2009-09-01

    Recently, oriental medicine has received attention for providing personalized medicine through consideration of the unique nature and constitution of individual patients. With the eventual goal of globalization, the current trend in oriental medicine research is the standardization by adopting western scientific methods, which could represent a scientific revolution. The purpose of this study is to establish methods for finding statistically significant features in a facial image with respect to distinguishing constitution and to show the meaning of those features. From facial photo images, facial elements are analyzed in terms of the distance, angle and the distance ratios, for which there are 1225, 61 250 and 749 700 features, respectively. Due to the very large number of facial features, it is quite difficult to determine truly meaningful features. We suggest a process for the efficient analysis of facial features including the removal of outliers, control for missing data to guarantee data confidence and calculation of statistical significance by applying ANOVA. We show the statistical properties of selected features according to different constitutions using the nine distances, 10 angles and 10 rates of distance features that are finally established. Additionally, the Sasang constitutional meaning of the selected features is shown here.

  5. Neutron autoradiography imaging of selective boron uptake in human metastatic tumours

    Energy Technology Data Exchange (ETDEWEB)

    Altieri, S. [Department of Nuclear and Theoretical Physics, University of Pavia, Via Bassi 6, Pavia (Italy); National Institute of Nuclear Physics (INFN), Section of Pavia, Via Bassi 6, Pavia (Italy)], E-mail: saverio.altieri@pv.infn.it; Bortolussi, S. [Department of Nuclear and Theoretical Physics, University of Pavia, Via Bassi 6, Pavia (Italy); National Institute of Nuclear Physics (INFN), Section of Pavia, Via Bassi 6, Pavia (Italy); Bruschi, P.; Chiari, P.; Fossati, F.; Stella, S. [Department of Nuclear and Theoretical Physics, University of Pavia, Via Bassi 6, Pavia (Italy); Prati, U.; Roveda, L. [Unit of cancer surgery, Cancer Center of Excellence, Foundation T. Campanella, Catanzaro (Italy); Zonta, A.; Zonta, C.; Ferrari, C.; Clerici, A. [Department of Surgery, University of Pavia, Piazza Botta, Pavia (Italy); Nano, R. [Department of Animal Biology, University of Pavia, Piazza Botta, Pavia (Italy); Pinelli, T. [Department of Nuclear and Theoretical Physics, University of Pavia, Via Bassi 6, Pavia (Italy); National Institute of Nuclear Physics (INFN), Section of Pavia, Via Bassi 6, Pavia (Italy)

    2008-12-15

    The ability to selectively hit the tumour cells is an essential characteristic of an anti-tumour therapy. In boron neutron capture therapy (BNCT) this characteristic is based on the selective uptake of {sup 10}B in the tumour cells with respect to normal tissues. An important step in the BNCT planning is the measurement of the boron concentration in the tissue samples, both tumour and healthy. When the tumour is spread through the healthy tissue, as in the case of metastases, the knowledge of the different kinds of tissues in the sample being analysed is crucial. If the percentage of tumour and normal tissues cannot be evaluated, the obtained concentration is a mean value depending on the composition of the different samples being measured. In this case an imaging method that could give information both on the morphology and on the spatial distribution of boron concentration in the sample would be a fundamental support. In this paper, the results of the boron uptake analysis in the tumour and in the healthy samples taken from human livers after boron phenylalanine (BPA) infusion are shown; boron imaging was performed using neutron autoradiography.

  6. Antibacterial Activity of Commercial Dentine Bonding Systems against E. faecalis–Flow Cytometry Study

    Directory of Open Access Journals (Sweden)

    Monika Lukomska-Szymanska

    2017-04-01

    Full Text Available Literature presents inconsistent results on the antibacterial activity of dentine bonding systems (DBS. Antibacterial activity of adhesive systems depends on several factors, including composition and acidity. Flow cytometry is a novel detection method to measure multiple characteristics of a single cell: total cell number, structural (size, shape, and functional parameters (viability, cell cycle. The LIVE/DEAD® BacLightTM bacterial viability assay was used to evaluate an antibacterial activity of DBS by assessing physical membrane disruption of bacteria mediated by DBS. Ten commercial DBSs: four total-etching (TE, four self-etching (SE and two selective enamel etching (SEE were tested. Both total-etching DBS ExciTE F and OptiBond Solo Plus showed comparatively low antibacterial activity against E. faecalis. The lowest activity of all tested TE systems showed Te-Econom Bond. Among SE DBS, G-ænial Bond (92.24% dead cells followed by Clearfil S3 Bond Plus (88.02% and Panavia F 2.0 ED Primer II (86.67% showed the highest antibacterial activity against E. faecalis, which was comparable to isopropranol (positive control. In the present study, self-etching DBS exhibited higher antimicrobial activity than tested total-etching adhesives against E. faecalis.

  7. MO-FG-204-02: Reference Image Selection in the Presence of Multiple Scan Realizations

    Energy Technology Data Exchange (ETDEWEB)

    Ruan, D; Dou, T; Thomas, D; Low, D [Deparment of Radiation Oncology, University of California Los Angeles, Los Angeles, CA (United States)

    2015-06-15

    Purpose: Fusing information from multiple correlated realizations (e.g., 4DCT) can improve image quality. This process often involves ill-conditioned and asymmetric nonlinear registration and the proper selection of a reference image is important. This work proposes to examine post-registration variation indirectly for such selection, and develops further insights to reduce the number of cross-registrations needed. Methods: We consider each individual scan as a noisy point in the vicinity of an image manifold, related by motion. Nonrigid registration “transports” a scan along the manifold to the reference neighborhood, and the residual is a surrogate for local variation. To test this conjecture, 10 thoracic scans from the same session were reconstructed from a recently developed low-dose helical 4DCT protocol. Pairwise registration was repeated bi-directionally (81 times) and fusion was performed with each candidate reference. The fused image quality was assessed with SNR and CNR. Registration residuals in SSD, harmonic energy, and deformation Jacobian behavior were examined. The semi-symmetry is further utilized to reduce the number of registration needed. Results: The comparison of image quality between single image and fused ones identified reduction of local intensity variance as the major contributor of image quality, boosting SNR and CNR by 5 to 7 folds. This observation further suggests the criticality of good agreement across post-registration images. Triangle inequality on the SSD metric provides a proficient upper-bound and surrogate on such disagreement. Empirical observation also confirms that fused images with high residual SSD have lower SNR and CNR than the ones with low or intermediate SSDs. Registration SSD is structurally close enough to symmetry for reduced computation. Conclusion: Registration residual is shown to be a good predictor of post-fusion image quality and can be used to identify good reference centers. Semi-symmetry of the

  8. Selective Detection of Neurotransmitters by Fluorescence and Chemiluminescence Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Ziqiang Wang; Edward S. Yeung

    2001-08-06

    In recent years, luminescence imaging has been widely employed in neurochemical analysis. It has a number of advantages for the study of neuronal and other biological cells: (1) a particular molecular species or cellular constituent can be selectively visualized in the presence of a large excess of other species in a heterogeneous environment; (2) low concentration detection limits can be achieved because of the inherent sensitivity associated with fluorescence and chemiluminescence; (3) low excitation intensities can be used so that long-term observation can be realized while the viability of the specimen is preserved; and (4) excellent spatial resolution can be obtained with the light microscope so subcellular compartments can be identified. With good sensitivity, temporal and spatial resolution, the flux of ions and molecules and the distribution and dynamics of intracellular species can be measured in real time with specific luminescence probes, substrates, or with native fluorescence. A noninvasive detection scheme based on glutamate dehydrogenase (GDH) enzymatic assay combined with microscopy was developed to measure the glutamate release in cultured cells from the central nervous system (CNS). The enzyme reaction is very specific and sensitive. The detection limit with CCD imaging is down to {micro}M levels of glutamate with reasonable response time. They also found that chemiluminescence associated with the ATP-dependent reaction between luciferase and luciferin can be used to image ATP at levels down to 10 nM in the millisecond time scale. Similar imaging experiments should be feasible in a broad spectrum of biological systems.

  9. A hybrid peptide PTS that facilitates transmembrane delivery and its application for the rapid in vivo imaging via near-infrared fluorescence imaging

    Directory of Open Access Journals (Sweden)

    Xuejiao eYan

    2016-03-01

    Full Text Available Background and purpose: Intravital imaging provides invaluable readouts for clinical diagnoses and therapies and shows great potential in the design of individualized drug dosage regimes. Ts is a mammalian free cell membrane-penetrating peptide. This study aimed to introduce a novel approach to the design of a cancer-selective peptide on the basis of a membrane-penetrating peptide and to explore its potential as a carrier of medical substances. Experimental approach: Ts was linked with a αvβ3-binding peptide P1c to create a hybrid referred to as PTS. The hybrid was labeled with an FITC or Cy5.5 as an imaging indicator to evaluate its in vitro and in vivo bioactivity. Key results: Hemolysis tests proved that in comparison with Ts, PTS caused similar or even less leakage of human erythrocytes at concentrations of up to 1 mmol/L. Flow cytometry assay and confocal microscopy demonstrated the following. 1 P1c alone could target and mostly halt at the cancer cell membrane. 2 Ts alone could not bind to the membrane sufficiently. 3 P1c greatly enhanced the binding affinity of PTS with MDA-MB-231 breast cancer cells that upregulated αvβ3. 4 Ts conferred PTS with the ability to traverse a cell membrane and thus facilitate the transmembrane delivery of imaging probes. In vivo near-infrared fluorescence (NIRF imaging demonstrated that the imaging probes were rapidly concentrated in a MDA-MB-231 tumor tissue within 1 h after intravenous injection. Conclusions and implications: PTS exhibited the capability of targeting specific tumors and greatly facilitating the transmembrane delivery of imaging probes.

  10. Two-stage atlas subset selection in multi-atlas based image segmentation

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Tingting, E-mail: tingtingzhao@mednet.ucla.edu; Ruan, Dan, E-mail: druan@mednet.ucla.edu [The Department of Radiation Oncology, University of California, Los Angeles, California 90095 (United States)

    2015-06-15

    Purpose: Fast growing access to large databases and cloud stored data presents a unique opportunity for multi-atlas based image segmentation and also presents challenges in heterogeneous atlas quality and computation burden. This work aims to develop a novel two-stage method tailored to the special needs in the face of large atlas collection with varied quality, so that high-accuracy segmentation can be achieved with low computational cost. Methods: An atlas subset selection scheme is proposed to substitute a significant portion of the computationally expensive full-fledged registration in the conventional scheme with a low-cost alternative. More specifically, the authors introduce a two-stage atlas subset selection method. In the first stage, an augmented subset is obtained based on a low-cost registration configuration and a preliminary relevance metric; in the second stage, the subset is further narrowed down to a fusion set of desired size, based on full-fledged registration and a refined relevance metric. An inference model is developed to characterize the relationship between the preliminary and refined relevance metrics, and a proper augmented subset size is derived to ensure that the desired atlases survive the preliminary selection with high probability. Results: The performance of the proposed scheme has been assessed with cross validation based on two clinical datasets consisting of manually segmented prostate and brain magnetic resonance images, respectively. The proposed scheme demonstrates comparable end-to-end segmentation performance as the conventional single-stage selection method, but with significant computation reduction. Compared with the alternative computation reduction method, their scheme improves the mean and medium Dice similarity coefficient value from (0.74, 0.78) to (0.83, 0.85) and from (0.82, 0.84) to (0.95, 0.95) for prostate and corpus callosum segmentation, respectively, with statistical significance. Conclusions: The authors

  11. Two-stage atlas subset selection in multi-atlas based image segmentation.

    Science.gov (United States)

    Zhao, Tingting; Ruan, Dan

    2015-06-01

    Fast growing access to large databases and cloud stored data presents a unique opportunity for multi-atlas based image segmentation and also presents challenges in heterogeneous atlas quality and computation burden. This work aims to develop a novel two-stage method tailored to the special needs in the face of large atlas collection with varied quality, so that high-accuracy segmentation can be achieved with low computational cost. An atlas subset selection scheme is proposed to substitute a significant portion of the computationally expensive full-fledged registration in the conventional scheme with a low-cost alternative. More specifically, the authors introduce a two-stage atlas subset selection method. In the first stage, an augmented subset is obtained based on a low-cost registration configuration and a preliminary relevance metric; in the second stage, the subset is further narrowed down to a fusion set of desired size, based on full-fledged registration and a refined relevance metric. An inference model is developed to characterize the relationship between the preliminary and refined relevance metrics, and a proper augmented subset size is derived to ensure that the desired atlases survive the preliminary selection with high probability. The performance of the proposed scheme has been assessed with cross validation based on two clinical datasets consisting of manually segmented prostate and brain magnetic resonance images, respectively. The proposed scheme demonstrates comparable end-to-end segmentation performance as the conventional single-stage selection method, but with significant computation reduction. Compared with the alternative computation reduction method, their scheme improves the mean and medium Dice similarity coefficient value from (0.74, 0.78) to (0.83, 0.85) and from (0.82, 0.84) to (0.95, 0.95) for prostate and corpus callosum segmentation, respectively, with statistical significance. The authors have developed a novel two-stage atlas

  12. Two-stage atlas subset selection in multi-atlas based image segmentation

    International Nuclear Information System (INIS)

    Zhao, Tingting; Ruan, Dan

    2015-01-01

    Purpose: Fast growing access to large databases and cloud stored data presents a unique opportunity for multi-atlas based image segmentation and also presents challenges in heterogeneous atlas quality and computation burden. This work aims to develop a novel two-stage method tailored to the special needs in the face of large atlas collection with varied quality, so that high-accuracy segmentation can be achieved with low computational cost. Methods: An atlas subset selection scheme is proposed to substitute a significant portion of the computationally expensive full-fledged registration in the conventional scheme with a low-cost alternative. More specifically, the authors introduce a two-stage atlas subset selection method. In the first stage, an augmented subset is obtained based on a low-cost registration configuration and a preliminary relevance metric; in the second stage, the subset is further narrowed down to a fusion set of desired size, based on full-fledged registration and a refined relevance metric. An inference model is developed to characterize the relationship between the preliminary and refined relevance metrics, and a proper augmented subset size is derived to ensure that the desired atlases survive the preliminary selection with high probability. Results: The performance of the proposed scheme has been assessed with cross validation based on two clinical datasets consisting of manually segmented prostate and brain magnetic resonance images, respectively. The proposed scheme demonstrates comparable end-to-end segmentation performance as the conventional single-stage selection method, but with significant computation reduction. Compared with the alternative computation reduction method, their scheme improves the mean and medium Dice similarity coefficient value from (0.74, 0.78) to (0.83, 0.85) and from (0.82, 0.84) to (0.95, 0.95) for prostate and corpus callosum segmentation, respectively, with statistical significance. Conclusions: The authors

  13. 78 FR 5186 - Clinical Flow Cytometry in Hematologic Malignancies; Public Workshop; Request for Comments

    Science.gov (United States)

    2013-01-24

    ... need for such products to assist clinical laboratories in performing this testing. FDA has been working... this workshop include: (1) Overview of Quality control and standardization issues associated with Clinical Flow Cytometry (FCM), including discussion of a NIST traceable standard; (2) Biological controls...

  14. Sample handling for kinetics and molecular assembly in flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Sklar, L.A. [Los Alamos National Lab., NM (United States). National Flow Cytometry Resource]|[Univ. of New Mexico, Albuquerque, NM (United States). School of Medicine; Seamer, L.C.; Kuckuck, F.; Prossnitz, E.; Edwards, B. [Univ. of New Mexico, Albuquerque, NM (United States). School of Medicine; Posner, G. [Northern Arizona Univ., Flagstaff, AZ (United States). Dept. of Chemistry

    1998-07-01

    Flow cytometry discriminates particle associated fluorescence from the fluorescence of the surrounding medium. It permits assemblies of macromolecular complexes on beads or cells to be detected in real-time with precision and specificity. The authors have investigated two types of robust sample handling systems which provide sub-second resolution and high throughput: (1) mixers which use stepper-motor driven syringes to initiate chemical reactions in msec time frames; and (2) flow injection controllers with valves and automated syringes used in chemical process control. In the former system, the authors used fast valves to overcome the disparity between mixing 100 {micro}ls of sample in 100 msecs and delivering sample to a flow cytometer at 1 {micro}l/sec. Particles were detected within 100 msec after mixing, but turbulence was created which lasted for 1 sec after injection of the sample into the flow cytometer. They used optical criteria to discriminate particles which were out of alignment due to the turbulent flow. Complex sample handling protocols involving multiple mixing steps and sample dilution have also been achieved. With the latter system they were able to automate sample handling and delivery with intervals of a few seconds. The authors used a fluidic approach to defeat turbulence caused by sample introduction. By controlling both sheath and sample with individual syringes, the period of turbulence was reduced to {approximately} 200 msecs. Automated sample handling and sub-second resolution should permit broad analytical and diagnostic applications of flow cytometry.

  15. Evaluation of CD307a expression patterns during normal B-cell maturation and in B-cell malignancies by flow cytometry.

    Science.gov (United States)

    Auat, Mariangeles; Cardoso, Chandra Chiappin; Santos-Pirath, Iris Mattos; Rudolf-Oliveira, Renata Cristina Messores; Matiollo, Camila; Lange, Bárbara Gil; da Silva, Jessica Pires; Dametto, Gisele Cristina; Pirolli, Mayara Marin; Colombo, Maria Daniela Holthausen Perico; Santos-Silva, Maria Claudia

    2018-02-24

    Flow cytometric immunophenotyping is deemed a fundamental tool for the diagnosis of B-cell neoplasms. Currently, the investigation of novel immunophenotypic markers has gained importance, as they can assist in the precise subclassification of B-cell malignancies by flow cytometry. Therefore, the purpose of the present study was to evaluate the expression of CD307a during normal B-cell maturation and in B-cell malignancies as well as to investigate its potential role in the differential diagnosis of these entities. CD307a expression was assessed by flow cytometry in normal precursor and mature B cells and in 115 samples collected from patients diagnosed with precursor and mature B-cell neoplasms. CD307a expression was compared between neoplastic and normal B cells. B-acute lymphoblastic leukemia cases exhibited minimal expression of CD307a, displaying a similar expression pattern to that of normal B-cell precursors. Mantle cell lymphoma (MCL) cases showed the lowest levels of CD307a among mature B-cell neoplasms. CD307a expression was statistically lower in MCL cases than in chronic B lymphocytic leukemia (CLL) and marginal zone lymphoma (MZL) cases. No statistical differences were observed between CD307a expression in neoplastic and normal plasma cells. These results indicate that the assessment of CD307a expression by flow cytometry could be helpful to distinguish CLL from MCL, and the latter from MZL. Although these results are not entirely conclusive, they provide a basis for further studies in a larger cohort of patients. © 2018 International Clinical Cytometry Society. © 2018 International Clinical Cytometry Society.

  16. EMS mutant spectra generated by multi-parameter flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Keysar, Stephen B. [Cell and Molecular Biology Graduate Program, Colorado State University, Fort Collins, CO (United States); Fox, Michael H., E-mail: michael.fox@colostate.edu [Cell and Molecular Biology Graduate Program, Colorado State University, Fort Collins, CO (United States); Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO (United States)

    2009-12-01

    The CHO A{sub L} cell line contains a single copy of human chromosome 11 that encodes several cell surface proteins including glycosyl phosphatidylinositol (GPI) linked CD59 and CD90, as well as CD98, CD44 and CD151 which are not GPI-linked. The flow cytometry mutation assay (FCMA) measures mutations of the CD59 gene by the absence of fluorescence when stained with antibodies against the CD59 cell surface protein. We have measured simultaneous mutations in CD59, CD44, CD90, CD98 and CD151 to generate a mutant spectrum for ionizing radiation. After treatment with ethyl methanesulfonate (EMS) many cells have an intermediate level of CD59 staining. Single cells were sorted from CD59{sup -} regions with varying levels of fluorescence and the resulting clonal populations had a stable phenotype for CD59 expression. Mutant spectra were generated by flow cytometry using the isolated clones and nearly all clones were mutated in CD59 only. Interestingly, about 60% of the CD59 negative clones were actually GPI mutants determined by staining with the GPI specific fluorescently labeled bacterial toxin aerolysin (FLAER). The GPI negative cells are most likely caused by mutations in the X-linked pigA gene important in GPI biosynthesis. Small mutations of pigA and CD59 were expected for the alkylating agent EMS and the resulting spectra are significantly different than the large deletions found when analyzing radiation mutants. After analyzing the CD59{sup -} clonal populations we have adjusted the FCMA mutant regions from 1% to 10% of the mean of the CD59 positive peak to include the majority of CD59 mutants.

  17. Use of flow cytometry for the possible identification of radio-induced changes in DNA of animal cells

    International Nuclear Information System (INIS)

    Spano, M.; Leonardi, M.; Cordelli, E.

    1991-01-01

    Since DNA is the main cellular target of ionizing irradiation, methods fit for analyzing DNA alterations should be able to discern irradiated versus control cells. Flow cytometry allows the rapid measurement of DNA content of single chromosomes or cell nuclei at very high resolution on a statistically significant sample. Alterations of chromatine structure can also be analyzed by flow cytometry. Briefly, evaluation of in situ DNA resistance to denaturation can be evaluated by flow cytometric analysis of different staining pattern of single versus double strange regions of DNA. In the present work both approaches were used with the aim to recognize cells derived from an irradiated sample of breast chicken. Although flow cytometry has been demonstrated to be a useful tool to detect DNA alterations and has been widely used to detect damages on DNA induced by several physical and chemical agents, it was unable to detect clastogenic effects induced by electrons on DNA of chicken breast cells. Heavily irradiated nuclei, even if challenged by denaturating treatments that partially collapse chromatine organization, do not present differences from non irradiated samples after flow cytometric DNA content measurement. (16 refs)

  18. An automatic fuzzy-based multi-temporal brain digital subtraction angiography image fusion algorithm using curvelet transform and content selection strategy.

    Science.gov (United States)

    Momeni, Saba; Pourghassem, Hossein

    2014-08-01

    Recently image fusion has prominent role in medical image processing and is useful to diagnose and treat many diseases. Digital subtraction angiography is one of the most applicable imaging to diagnose brain vascular diseases and radiosurgery of brain. This paper proposes an automatic fuzzy-based multi-temporal fusion algorithm for 2-D digital subtraction angiography images. In this algorithm, for blood vessel map extraction, the valuable frames of brain angiography video are automatically determined to form the digital subtraction angiography images based on a novel definition of vessel dispersion generated by injected contrast material. Our proposed fusion scheme contains different fusion methods for high and low frequency contents based on the coefficient characteristic of wrapping second generation of curvelet transform and a novel content selection strategy. Our proposed content selection strategy is defined based on sample correlation of the curvelet transform coefficients. In our proposed fuzzy-based fusion scheme, the selection of curvelet coefficients are optimized by applying weighted averaging and maximum selection rules for the high frequency coefficients. For low frequency coefficients, the maximum selection rule based on local energy criterion is applied to better visual perception. Our proposed fusion algorithm is evaluated on a perfect brain angiography image dataset consisting of one hundred 2-D internal carotid rotational angiography videos. The obtained results demonstrate the effectiveness and efficiency of our proposed fusion algorithm in comparison with common and basic fusion algorithms.

  19. Automatic spectral imaging protocol selection and iterative reconstruction in abdominal CT with reduced contrast agent dose: initial experience.

    Science.gov (United States)

    Lv, Peijie; Liu, Jie; Chai, Yaru; Yan, Xiaopeng; Gao, Jianbo; Dong, Junqiang

    2017-01-01

    To evaluate the feasibility, image quality, and radiation dose of automatic spectral imaging protocol selection (ASIS) and adaptive statistical iterative reconstruction (ASIR) with reduced contrast agent dose in abdominal multiphase CT. One hundred and sixty patients were randomly divided into two scan protocols (n = 80 each; protocol A, 120 kVp/450 mgI/kg, filtered back projection algorithm (FBP); protocol B, spectral CT imaging with ASIS and 40 to 70 keV monochromatic images generated per 300 mgI/kg, ASIR algorithm. Quantitative parameters (image noise and contrast-to-noise ratios [CNRs]) and qualitative visual parameters (image noise, small structures, organ enhancement, and overall image quality) were compared. Monochromatic images at 50 keV and 60 keV provided similar or lower image noise, but higher contrast and overall image quality as compared with 120-kVp images. Despite the higher image noise, 40-keV images showed similar overall image quality compared to 120-kVp images. Radiation dose did not differ between the two protocols, while contrast agent dose in protocol B was reduced by 33 %. Application of ASIR and ASIS to monochromatic imaging from 40 to 60 keV allowed contrast agent dose reduction with adequate image quality and without increasing radiation dose compared to 120 kVp with FBP. • Automatic spectral imaging protocol selection provides appropriate scan protocols. • Abdominal CT is feasible using spectral imaging and 300 mgI/kg contrast agent. • 50-keV monochromatic images with 50 % ASIR provide optimal image quality.

  20. Induction studies with Escherichia coli expressing recombinant interleukin-13 using multi-parameter flow cytometry

    DEFF Research Database (Denmark)

    Shitu, J. O.; Woodley, John; Wnek, R.

    2009-01-01

    The expression of interleukin-13 (IL13) following induction with IPTG in Escherichia coli results in metabolic changes as indicated by multi-parameter flow cytometry and traditional methods of fermentation profiling (O-2 uptake rate, CO2 evolution rate and optical density measurements). Induction...

  1. Preparation and flow cytometry of uniform silica-fluorescent dye microspheres.

    Science.gov (United States)

    Bele, Marjan; Siiman, Olavi; Matijević, Egon

    2002-10-15

    Uniform fluorescent silica-dye microspheres have been prepared by coating preformed monodispersed silica particles with silica layers containing rhodamine 6G or acridine orange. The resulting dispersions exhibit intense fluorescent emission between 500 and 600 nm, over a broad excitation wavelength range of 460 to 550 nm, even with exceedingly small amounts of dyes incorporated into the silica particles (10-30 ppm, expressed as weight of dye relative to weight of dry particles). The fluorescent particles can be prepared in micrometer diameters suitable for analyses using flow cytometry with 488-nm laser excitation.

  2. Metabolic cytometry: capillary electrophoresis with two-color fluorescence detection for the simultaneous study of two glycosphingolipid metabolic pathways in single primary neurons.

    Science.gov (United States)

    Essaka, David C; Prendergast, Jillian; Keithley, Richard B; Palcic, Monica M; Hindsgaul, Ole; Schnaar, Ronald L; Dovichi, Norman J

    2012-03-20

    Metabolic cytometry is a form of chemical cytometry wherein metabolic cascades are monitored in single cells. We report the first example of metabolic cytometry where two different metabolic pathways are simultaneously monitored. Glycolipid catabolism in primary rat cerebella neurons was probed by incubation with tetramethylrhodamine-labeled GM1 (GM1-TMR). Simultaneously, both catabolism and anabolism were probed by coincubation with BODIPY-FL labeled LacCer (LacCer-BODIPY-FL). In a metabolic cytometry experiment, single cells were incubated with substrate, washed, aspirated into a capillary, and lysed. The components were separated by capillary electrophoresis equipped with a two-spectral channel laser-induced fluorescence detector. One channel monitored fluorescence generated by the metabolic products produced from GM1-TMR and the other monitored the metabolic products produced from LacCer-BODIPY-FL. The metabolic products were identified by comparison with the mobility of a set of standards. The detection system produced at least 6 orders of magnitude dynamic range in each spectral channel with negligible spectral crosstalk. Detection limits were 1 zmol for BODIPY-FL and 500 ymol for tetramethylrhodamine standard solutions.

  3. Scalable imaging of scattering organisms with hybrid selective plane illumination microscopy and optoacoustic tomography

    OpenAIRE

    Lin, Hsiao Chun Amy

    2017-01-01

    Biomedical imaging plays a key role in the advancement of medical research. While high-resolution microscopy enables the harvest of molecular and cellular information, a holistic picture on organ level can only be provided by means of macroscopic imaging. Wedged in between Micro-macro, the mesoscopic regime offers important bridging of the information transfer. The focus of the research presented in this thesis centers around the application of selective plane illumination microscopy (SPIM) a...

  4. Fusion set selection with surrogate metric in multi-atlas based image segmentation

    International Nuclear Information System (INIS)

    Zhao, Tingting; Ruan, Dan

    2016-01-01

    Multi-atlas based image segmentation sees unprecedented opportunities but also demanding challenges in the big data era. Relevant atlas selection before label fusion plays a crucial role in reducing potential performance loss from heterogeneous data quality and high computation cost from extensive data. This paper starts with investigating the image similarity metric (termed ‘surrogate’), an alternative to the inaccessible geometric agreement metric (termed ‘oracle’) in atlas relevance assessment, and probes into the problem of how to select the ‘most-relevant’ atlases and how many such atlases to incorporate. We propose an inference model to relate the surrogates and the oracle geometric agreement metrics. Based on this model, we quantify the behavior of the surrogates in mimicking oracle metrics for atlas relevance ordering. Finally, analytical insights on the choice of fusion set size are presented from a probabilistic perspective, with the integrated goal of including the most relevant atlases and excluding the irrelevant ones. Empirical evidence and performance assessment are provided based on prostate and corpus callosum segmentation. (paper)

  5. Feasibility and reliability of digital imaging for estimating food selection and consumption from students' packed lunches.

    Science.gov (United States)

    Taylor, Jennifer C; Sutter, Carolyn; Ontai, Lenna L; Nishina, Adrienne; Zidenberg-Cherr, Sheri

    2018-01-01

    Although increasing attention is placed on the quality of foods in children's packed lunches, few studies have examined the capacity of observational methods to reliably determine both what is selected and consumed from these lunches. The objective of this project was to assess the feasibility and inter-rater reliability of digital imaging for determining selection and consumption from students' packed lunches, by adapting approaches previously applied to school lunches. Study 1 assessed feasibility and reliability of data collection among a sample of packed lunches (n = 155), while Study 2 further examined reliability in a larger sample of packed (n = 386) as well as school (n = 583) lunches. Based on the results from Study 1, it was feasible to collect and code most items in packed lunch images; missing data were most commonly attributed to packaging that limited visibility of contents. Across both studies, there was satisfactory reliability for determining food types selected, quantities selected, and quantities consumed in the eight food categories examined (weighted kappa coefficients 0.68-0.97 for packed lunches, 0.74-0.97 for school lunches), with lowest reliability for estimating condiments and meats/meat alternatives in packed lunches. In extending methods predominately applied to school lunches, these findings demonstrate the capacity of digital imaging for the objective estimation of selection and consumption from both school and packed lunches. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Quantitative Magnetization Transfer Imaging in Human Brain at 3 T via Selective Inversion Recovery

    OpenAIRE

    Dortch, Richard D.; Li, Ke; Gochberg, Daniel F.; Welch, E. Brian; Dula, Adrienne N.; Tamhane, Ashish A.; Gore, John C.; Smith, Seth A.

    2011-01-01

    Quantitative magnetization transfer imaging yields indices describing the interactions between free water protons and immobile, macromolecular protons—including the macromolecular to free pool size ratio (PSR) and the rate of magnetization transfer between pools kmf. This study describes the first implementation of the selective inversion recovery quantitative magnetization transfer method on a clinical 3.0-T scanner in human brain in vivo. Selective inversion recovery data were acquired at 1...

  7. Imaging a Large Sample with Selective Plane Illumination Microscopy Based on Multiple Fluorescent Microsphere Tracking

    Science.gov (United States)

    Ryu, Inkeon; Kim, Daekeun

    2018-04-01

    A typical selective plane illumination microscopy (SPIM) image size is basically limited by the field of view, which is a characteristic of the objective lens. If an image larger than the imaging area of the sample is to be obtained, image stitching, which combines step-scanned images into a single panoramic image, is required. However, accurately registering the step-scanned images is very difficult because the SPIM system uses a customized sample mount where uncertainties for the translational and the rotational motions exist. In this paper, an image registration technique based on multiple fluorescent microsphere tracking is proposed in the view of quantifying the constellations and measuring the distances between at least two fluorescent microspheres embedded in the sample. Image stitching results are demonstrated for optically cleared large tissue with various staining methods. Compensation for the effect of the sample rotation that occurs during the translational motion in the sample mount is also discussed.

  8. The Proteolytically Stable Peptidomimetic Pam-(Lys-ßNSpe)6-NH2 Selectively Inhibits Human Neutrophil Activation via Formyl Peptide Receptor 2

    DEFF Research Database (Denmark)

    Skovbakke, Sarah Line; Heegaard, Peter M. H.; Larsen, Camilla J.

    2015-01-01

    of proteolytically stable HDP mimics consisting of lipidated α-peptide/β-peptoid oligomers was investigated for their effect on neutrophil function. The most promising compound, Pam-(Lys-βNSpe)6-NH2, was shown to inhibit formyl peptide receptor 2 (FPR2) agonist-induced neutrophil granule mobilization and release...... of reactive oxygen species. The potency of Pam-(Lys-βNSpe)6-NH2 was comparable to that of PBP10, the most potent FPR2-selective inhibitor known. The immunomodulatory effects of structural analogues of Pam-(Lys-βNSpe)6-NH2 emphasized the importance of both the lipid and peptidomimetic parts. By using imaging...... flow cytometry in primary neutrophils and FPR-transfected cell lines we found that a fluorescently labelled analogue of Pam-(Lys-βNSpe)6-NH2 interacted selectively with FPR2. Furthermore the interaction between Pam-(Lys-βNSpe)6-NH2 and FPR2 was found to prevent binding of the FPR2-specific activating...

  9. The impact of exercise myocardial perfusion SPECT imaging on the selection of patients for coronary angiography

    International Nuclear Information System (INIS)

    Song Liping; He Zuoxiang; Liu Xiujie; Shi Rongfang; Liu Yunzhong; Tian Yueqin; Zhang Xiaoli; Qin Xuewen; Chen Jilin; Gao Runlin

    2001-01-01

    Objective: Exercise 99 Tc m -MIBI myocardial perfusion SPECT is accurate for the diagnosis of coronary artery disease (CAD). This study assessed the impact of exercise myocardial perfusion imaging on the selection of patients for coronary angiography. Methods: 2188 consecutive patients who underwent exercise myocardial perfusion SPECT in authors' department in 1999 were retrospectively analyzed. Among them, 1807 were men, 381 women (average age: 53.5 +- 7.2 years). Overall, exercise myocardial SPECT was normal in 1731 patients, abnormal in 359 cases, and equivocal in 98 patients. There were 141 patients who underwent CAG within 60 days after myocardial SPECT. Results: Overall, 12% of the patients with abnormal SPECT imaging underwent coronary angiography, but only 5% of the patients with a normal SPECT imaging did (P < 0.001). Among these 141 patients who underwent coronary angiography, significant coronary stenosis was present in 91% of the patients who had had an abnormal SPECT imaging, but only 8% of those who had had a normal SPECT imaging (P < 0.001). In those patients who underwent coronary angiography, revascularization rate was 25% for the patients with abnormal SPECT imaging, but only 1% for the patients with a normal SPECT imaging. Conclusion: The results of exercise myocardial perfusion SPECT have a significant impact on the selection of patients for coronary angiography and revascularization

  10. Picture Yourself Healthy-How Users Select Mediated Images to Shape Health Intentions and Behaviors.

    Science.gov (United States)

    Wilson, Brianna; Knobloch-Westerwick, Silvia; Robinson, Melissa J

    2018-02-20

    Hypotheses on how selective viewing of mediated images may sustain eating habits and aid healthier eating were derived from the Selective Exposure Self- and Affect Management model. The model posits that individuals select to view media to manage their self-concepts-and that this exposure affects subsequent intentions and behaviors. Participants (N = 265) selectively viewed Instagram-like postings featuring healthy or unhealthy food imagery. Beforehand, participants reported habits and perceived expert recommendations regarding food intake. After viewing postings, participants chose gift cards representing healthy or unhealthy food purchases and indicated food intake intentions. Results show that existing eating behavior predicts selective exposure to healthy or unhealthy food imagery, which in turn shapes gift card choices and (both healthy and unhealthy) food intake intentions.

  11. Image registration via optimization over disjoint image regions

    Science.gov (United States)

    Pitts, Todd; Hathaway, Simon; Karelitz, David B.; Sandusky, John; Laine, Mark Richard

    2018-02-06

    Technologies pertaining to registering a target image with a base image are described. In a general embodiment, the base image is selected from a set of images, and the target image is an image in the set of images that is to be registered to the base image. A set of disjoint regions of the target image is selected, and a transform to be applied to the target image is computed based on the optimization of a metric over the selected set of disjoint regions. The transform is applied to the target image so as to register the target image with the base image.

  12. Multiparameter flow cytometry reveals myelodysplasia-related aberrant antigen expression in myelodysplastic/myeloproliferative neoplasms.

    Science.gov (United States)

    Kern, Wolfgang; Bacher, Ulrike; Schnittger, Susanne; Alpermann, Tamara; Haferlach, Claudia; Haferlach, Torsten

    2013-05-01

    Within the myelodysplastic/myeloproliferative neoplasm (MDS/MPN) category of the WHO (2008), only chronic myelomonocytic leukemia was so far evaluated by multiparameter flow cytometry (MFC). To investigate the potential of MFC for MDS/MPNs, unclassifiable (MDS/MPNu), and refractory anemia associated with ring sideroblasts and marked thrombocytosis (RARS-T), we studied 91 patients with these entities (60 males/31 females; 35.3-87.4 years) for MDS-related aberrant immunophenotypes (≥ 2 different cell lineages with ≥ 3 aberrantly expressed antigens). Data were correlated with cytomorphology and cytogenetics. MFC identified MDS-related immunophenotypes in 54/91 (59.3%) of patients. Patients with or without MDS-related immunophenotype did not differ significantly by demographic characteristics, blood values, or median overall survival. MDS-related immunophenotype cases showed a higher number of aberrantly expressed antigens (mean ± SD, 4.9 ± 2.4 vs. 2.0 ± 1.4; P MPNu and RARS-T. MFC therefore may be helpful to separate cases into more "MDS-like" or "MPN-like" subgroups. Copyright © 2012 International Clinical Cytometry Society.

  13. Flow Cytometry-Assisted Cloning of Specific Sequence Motifs from Complex 16S rRNA Gene Libraries

    DEFF Research Database (Denmark)

    Nielsen, Jeppe Lund; Schramm, Andreas; Bernhard, Anne E.

    2004-01-01

    for Systems Biology,3 Seattle, Washington, and Department of Ecological Microbiology, University of Bayreuth, Bayreuth, Germany2 A flow cytometry method was developed for rapid screening and recovery of cloned DNA containing common sequence motifs. This approach, termed fluorescence-activated cell sorting......  FLOW CYTOMETRY-ASSISTED CLONING OF SPECIFIC SEQUENCE MOTIFS FROM COMPLEX 16S RRNA GENE LIBRARIES Jeppe L. Nielsen,1 Andreas Schramm,1,2 Anne E. Bernhard,1 Gerrit J. van den Engh,3 and David A. Stahl1* Department of Civil and Environmental Engineering, University of Washington,1 and Institute......-assisted cloning, was used to recover sequences affiliated with a unique lineage within the Bacteroidetes not abundant in a clone library of environmental 16S rRNA genes.  ...

  14. Utility of peripheral blood immunophenotyping by flow cytometry in the diagnosis of pediatric acute leukemia.

    Science.gov (United States)

    Metrock, Laura K; Summers, Ryan J; Park, Sunita; Gillespie, Scott; Castellino, Sharon; Lew, Glen; Keller, Frank G

    2017-10-01

    Childhood acute leukemia is traditionally diagnosed from a bone marrow aspirate (BMA). New-onset acute leukemia patients do not always have visible circulating blasts in the peripheral blood (PB) at diagnosis. While the role of bone marrow flow cytometry for the diagnosis of acute leukemia is well established, the utility of PB flow cytometry (PBFC) is unknown. We performed a single-institution retrospective analysis to compare PBFC versus BMA in establishing or excluding a diagnosis of childhood acute leukemia. We retrospectively identified 485 PBFC samples with concurrent BMA from 2008 to 2013. Results of four-color flow cytometry for immunophenotypic characterization of leukemic versus nonclonal disease were characterized. Sensitivity and specificity were calculated among patients without a known diagnosis or prior therapy. Among 485 samples eligible for analysis, 120 had negative PBFC and BMA, 359 had positive PBFC and BMA, 3 had negative PBFC and positive BMA, and 3 had positive PBFC and negative BMA. There were small but significant differences in sensitivity (100 vs. 93.8%; P = 0.002) and positive predictive value (100 vs. 93.8%; P = 0.002) favoring BMA over PBFC among those demonstrating absence of circulating morphologic blasts. PBFC has high sensitivity and specificity for the diagnosis of childhood acute leukemia. The predictive value of PBFC remains high for patients without visible circulating blasts and may enhance the diagnostic process for determining the indications for marrow testing. © 2017 Wiley Periodicals, Inc.

  15. Scanning technology selection impacts acceptability and usefulness of image-rich content

    Directory of Open Access Journals (Sweden)

    Kristine M. Alpi

    2016-01-01

    Full Text Available Objective: Clinical and research usefulness of articles can depend on image quality. This study addressed whether scans of figures in black and white (B&W, grayscale, or color, or portable document format (PDF to tagged image file format (TIFF conversions as provided by interlibrary loan or document delivery were viewed as acceptable or useful by radiologists or pathologists. Methods: Residency coordinators selected eighteen figures from studies from radiology, clinical pathology, and anatomic pathology journals.With original PDF controls, each figure was prepared in three or four experimental conditions: PDF conversion to TIFF, and scans from print in B&W, grayscale, and color. Twelve independent observers indicated whether they could identify the features and whether the image quality was acceptable. They also ranked all the experimental conditions of each figure in terms of usefulness. Results: Of 982 assessments of 87 anatomic pathology, 83 clinical pathology, and 77 radiology images, 471 (48% were unidentifiable. Unidentifiability of originals (4% and conversions (10% was low. For scans, unidentifiability ranged from 53% for color, to 74% for grayscale, to 97% for B&W. Of 987 responses about acceptability (n¼405, 41% were said to be unacceptable, 97% of B&W, 66% of grayscale, 41% of color, and 1% of conversions. Hypothesized order (original, conversion, color, grayscale, B&W matched 67% of rankings (n¼215. Conclusions: PDF to TIFF conversion provided acceptable content. Color images are rarely useful in grayscale (12% or B&W (less than 1%. Acceptability of grayscale scans of noncolor originals was 52%. Digital originals are needed for most images. Print images in color or grayscale should be scanned using those modalities.

  16. Screening of Compounds Toxicity against Human Monocytic cell line-THP-1 by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Pick Neora

    2004-01-01

    Full Text Available The worldwide rapid increase in bacterial resistance to numerous antibiotics requires on-going development of new drugs to enter the market. As the development of new antibiotics is lengthy and costly, early monitoring of compound's toxicity is essential in the development of novel agents. Our interest is in a rapid, simple, high throughput screening method to assess cytotoxicity induced by potential agents. Some intracellular pathogens, such as Mycobacterium tuberculosis primary site of infection is human alveolar macrophages. Thus, evaluation of candidate drugs for macrophage toxicity is crucial. Protocols for high throughput drug toxicity screening of macrophages using flow cytometry are lacking in the literature. For this application we modified a preexisting technique, propidium iodide (PI exclusion staining and utilized it for rapid toxicity tests. Samples were prepared in 96 well plates and analyzed by flow cytometry, which allowed for rapid, inexpensive and precise assessment of compound's toxicity associated with cell death.

  17. Detection of Intracellular Factor VIII Protein in Peripheral Blood Mononuclear Cells by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Gouri Shankar Pandey

    2013-01-01

    Full Text Available Flow cytometry is widely used in cancer research for diagnosis, detection of minimal residual disease, as well as immune monitoring and profiling following immunotherapy. Detection of specific host proteins for diagnosis predominantly uses quantitative PCR and western blotting assays. In this study, we optimized a flow cytometry-based detection assay for Factor VIII protein in peripheral blood mononuclear cells (PBMCs. An indirect intracellular staining (ICS method was standardized using monoclonal antibodies to different domains of human Factor VIII protein. The FVIII protein expression level was estimated by calculating the mean and median fluorescence intensities (MFI values for each monoclonal antibody. ICS staining of transiently transfected cell lines supported the method's specificity. Intracellular FVIII protein expression was also detected by the monoclonal antibodies used in the study in PBMCs of five blood donors. In summary, our data suggest that intracellular FVIII detection in PBMCs of hemophilia A patients can be a rapid and reliable method to detect intracellular FVIII levels.

  18. Hessian-based quantitative image analysis of host-pathogen confrontation assays.

    Science.gov (United States)

    Cseresnyes, Zoltan; Kraibooj, Kaswara; Figge, Marc Thilo

    2018-03-01

    Host-fungus interactions have gained a lot of interest in the past few decades, mainly due to an increasing number of fungal infections that are often associated with a high mortality rate in the absence of effective therapies. These interactions can be studied at the genetic level or at the functional level via imaging. Here, we introduce a new image processing method that quantifies the interaction between host cells and fungal invaders, for example, alveolar macrophages and the conidia of Aspergillus fumigatus. The new technique relies on the information content of transmitted light bright field microscopy images, utilizing the Hessian matrix eigenvalues to distinguish between unstained macrophages and the background, as well as between macrophages and fungal conidia. The performance of the new algorithm was measured by comparing the results of our method with that of an alternative approach that was based on fluorescence images from the same dataset. The comparison shows that the new algorithm performs very similarly to the fluorescence-based version. Consequently, the new algorithm is able to segment and characterize unlabeled cells, thus reducing the time and expense that would be spent on the fluorescent labeling in preparation for phagocytosis assays. By extending the proposed method to the label-free segmentation of fungal conidia, we will be able to reduce the need for fluorescence-based imaging even further. Our approach should thus help to minimize the possible side effects of fluorescence labeling on biological functions. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

  19. Improved CT-detection of acute bowel ischemia using frequency selective non-linear image blending.

    Science.gov (United States)

    Schneeweiss, Sven; Esser, Michael; Thaiss, Wolfgang; Boesmueller, Hans; Ditt, Hendrik; Nikolau, Konstantin; Horger, Marius

    2017-07-01

    Computed tomography (CT) as a fast and reliable diagnostic technique is the imaging modality of choice for acute bowel ischemia. However, diagnostic is often difficult mainly due to low attenuation differences between ischemic and perfused segments. To compare the diagnostic efficacy of a new post-processing tool based on frequency selective non-linear blending with that of conventional linear contrast-enhanced CT (CECT) image blending for the detection of bowel ischemia. Twenty-seven consecutive patients (19 women; mean age = 73.7 years, age range = 50-94 years) with acute bowel ischemia were scanned using multidetector CT (120 kV; 100-200 mAs). Pre-contrast and portal venous scans (65-70 s delay) were acquired. All patients underwent surgery for acute bowel ischemia and intraoperative diagnosis as well as histologic evaluation of explanted bowel segments was considered "gold standard." First, two radiologists read the conventional CECT images in which linear blending was adapted for optimal contrast, and second (three weeks later) the frequency selective non-linear blending (F-NLB) image. Attenuation values were compared, both in the involved and non-involved bowel segments creating ratios between unenhanced and CECT. The mean attenuation difference between ischemic and non-ischemic wall in the portal venous scan was 69.54 HU (reader 2 = 69.01 HU) higher for F-NLB compared with conventional CECT. Also, the attenuation ratio between contrast-enhanced and pre-contrast CT data for the non-ischemic walls showed significantly higher values for the F-NLB image (CECT: reader 1 = 2.11 (reader 2 = 3.36), F-NLB: reader 1 = 4.46 (reader 2 = 4.98)]. Sensitivity in detecting ischemic areas increased significantly for both readers using F-NLB (CECT: reader 1/2 = 53%/65% versus F-NLB: reader 1/2 = 62%/75%). Frequency selective non-linear blending improves detection of bowel ischemia compared with conventional CECT by increasing

  20. Block selective redaction for minimizing loss during de-identification of burned in text in irreversibly compressed JPEG medical images.

    Science.gov (United States)

    Clunie, David A; Gebow, Dan

    2015-01-01

    Deidentification of medical images requires attention to both header information as well as the pixel data itself, in which burned-in text may be present. If the pixel data to be deidentified is stored in a compressed form, traditionally it is decompressed, identifying text is redacted, and if necessary, pixel data are recompressed. Decompression without recompression may result in images of excessive or intractable size. Recompression with an irreversible scheme is undesirable because it may cause additional loss in the diagnostically relevant regions of the images. The irreversible (lossy) JPEG compression scheme works on small blocks of the image independently, hence, redaction can selectively be confined only to those blocks containing identifying text, leaving all other blocks unchanged. An open source implementation of selective redaction and a demonstration of its applicability to multiframe color ultrasound images is described. The process can be applied either to standalone JPEG images or JPEG bit streams encapsulated in other formats, which in the case of medical images, is usually DICOM.

  1. Feature Selection from a Facial Image for Distinction of Sasang Constitution

    Directory of Open Access Journals (Sweden)

    Imhoi Koo

    2009-01-01

    Full Text Available Recently, oriental medicine has received attention for providing personalized medicine through consideration of the unique nature and constitution of individual patients. With the eventual goal of globalization, the current trend in oriental medicine research is the standardization by adopting western scientific methods, which could represent a scientific revolution. The purpose of this study is to establish methods for finding statistically significant features in a facial image with respect to distinguishing constitution and to show the meaning of those features. From facial photo images, facial elements are analyzed in terms of the distance, angle and the distance ratios, for which there are 1225, 61 250 and 749 700 features, respectively. Due to the very large number of facial features, it is quite difficult to determine truly meaningful features. We suggest a process for the efficient analysis of facial features including the removal of outliers, control for missing data to guarantee data confidence and calculation of statistical significance by applying ANOVA. We show the statistical properties of selected features according to different constitutions using the nine distances, 10 angles and 10 rates of distance features that are finally established. Additionally, the Sasang constitutional meaning of the selected features is shown here.

  2. Feature Selection from a Facial Image for Distinction of Sasang Constitution

    Science.gov (United States)

    Koo, Imhoi; Kim, Jong Yeol; Kim, Myoung Geun

    2009-01-01

    Recently, oriental medicine has received attention for providing personalized medicine through consideration of the unique nature and constitution of individual patients. With the eventual goal of globalization, the current trend in oriental medicine research is the standardization by adopting western scientific methods, which could represent a scientific revolution. The purpose of this study is to establish methods for finding statistically significant features in a facial image with respect to distinguishing constitution and to show the meaning of those features. From facial photo images, facial elements are analyzed in terms of the distance, angle and the distance ratios, for which there are 1225, 61 250 and 749 700 features, respectively. Due to the very large number of facial features, it is quite difficult to determine truly meaningful features. We suggest a process for the efficient analysis of facial features including the removal of outliers, control for missing data to guarantee data confidence and calculation of statistical significance by applying ANOVA. We show the statistical properties of selected features according to different constitutions using the nine distances, 10 angles and 10 rates of distance features that are finally established. Additionally, the Sasang constitutional meaning of the selected features is shown here. PMID:19745013

  3. Fast and simple tool for the quantification of biofilm-embedded cells sub-populations from fluorescent microscopic images.

    Directory of Open Access Journals (Sweden)

    Mikhail I Bogachev

    Full Text Available Fluorescent staining is a common tool for both quantitative and qualitative assessment of pro- and eukaryotic cells sub-population fractions by using microscopy and flow cytometry. However, direct cell counting by flow cytometry is often limited, for example when working with cells rigidly adhered either to each other or to external surfaces like bacterial biofilms or adherent cell lines and tissue samples. An alternative approach is provided by using fluorescent microscopy and confocal laser scanning microscopy (CLSM, which enables the evaluation of fractions of cells subpopulations in a given sample. For the quantitative assessment of cell fractions in microphotographs, we suggest a simple two-step algorithm that combines single cells selection and the statistical analysis. To facilitate the first step, we suggest a simple procedure that supports finding the balance between the detection threshold and the typical size of single cells based on objective cell size distribution analysis. Based on a series of experimental measurements performed on bacterial and eukaryotic cells under various conditions, we show explicitly that the suggested approach effectively accounts for the fractions of different cell sub-populations (like the live/dead staining in our samples in all studied cases that are in good agreement with manual cell counting on microphotographs and flow cytometry data. This algorithm is implemented as a simple software tool that includes an intuitive and user-friendly graphical interface for the initial adjustment of algorithm parameters to the microphotographs analysis as well as for the sequential analysis of homogeneous series of similar microscopic images without further user intervention. The software tool entitled BioFilmAnalyzer is freely available online at https://bitbucket.org/rogex/biofilmanalyzer/downloads/.

  4. Quantitative co-localization and pattern analysis of endo-lysosomal cargo in subcellular image cytometry and validation on synthetic image sets

    DEFF Research Database (Denmark)

    Lund, Frederik W.; Wüstner, Daniel

    2017-01-01

    /LYSs. Analysis of endocytic trafficking relies heavily on quantitative fluorescence microscopy, but evaluation of the huge image data sets is challenging and demands computer-assisted statistical tools. Here, we describe how to use SpatTrack (www.sdu.dk/bmb/spattrack), an imaging toolbox, which we developed...... such synthetic vesicle patterns as “ground truth” for validation of two-channel analysis tools in SpatTrack, revealing their high reliability. An improved version of SpatTrack for microscopy-based quantification of cargo transport through the endo-lysosomal system accompanies this protocol....

  5. Molecular Characterization of Gastric Epithelial Cells Using Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Kevin A. Bockerstett

    2018-04-01

    Full Text Available The ability to analyze individual epithelial cells in the gastric mucosa would provide important insight into gastric disease, including chronic gastritis and progression to gastric cancer. However, the successful isolation of viable gastric epithelial cells (parietal cells, neck cells, chief cells, and foveolar cells from gastric glands has been limited due to difficulties in tissue processing. Furthermore, analysis and interpretation of gastric epithelial cell flow cytometry data has been difficult due to the varying sizes and light scatter properties of the different epithelial cells, high levels of autofluorescence, and poor cell viability. These studies were designed to develop a reliable method for isolating viable single cells from the corpus of stomachs and to optimize analyses examining epithelial cells from healthy and diseased stomach tissue by flow cytometry. We performed a two stage enzymatic digestion in which collagenase released individual gastric glands from the stromal tissue of the corpus, followed by a Dispase II digestion that dispersed these glands into greater than 1 × 106 viable single cells per gastric corpus. Single cell suspensions were comprised of all major cell lineages found in the normal gastric glands. A method describing light scatter, size exclusion, doublet discrimination, viability staining, and fluorescently-conjugated antibodies and lectins was used to analyze individual epithelial cells and immune cells. This technique was capable of identifying parietal cells and revealed that gastric epithelial cells in the chronically inflamed mucosa significantly upregulated major histocompatibility complexes (MHC I and II but not CD80 or CD86, which are costimulatory molecules involved in T cell activation. These studies describe a method for isolating viable single cells and a detailed description of flow cytometric analysis of cells from healthy and diseased stomachs. These studies begin to identify effects of

  6. TU-G-201-00: Imaging Equipment Specification and Selection in Radiation Oncology Departments

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2015-06-15

    This session will update therapeutic physicists on technological advancements and radiation oncology features of commercial CT, MRI, and PET/CT imaging systems. Also described are physicists’ roles in every stage of equipment selection, purchasing, and operation, including defining specifications, evaluating vendors, making recommendations, and optimal and safe use of imaging equipment in radiation oncology environment. The first presentation defines important terminology of CT and PET/CT followed by a review of latest innovations, such as metal artifact reduction, statistical iterative reconstruction, radiation dose management, tissue classification by dual energy CT and spectral CT, improvement in spatial resolution and sensitivity in PET, and potentials of PET/MR. We will also discuss important technical specifications and items in CT and PET/CT purchasing quotes and their impacts. The second presentation will focus on key components in the request for proposal for a MRI simulator and how to evaluate vendor proposals. MRI safety issues in radiation Oncology, including MRI scanner Zones (4-zone design), will be discussed. Basic MR terminologies, important functionalities, and advanced features, which are relevant to radiation therapy, will be discussed. In the third presentation, justification of imaging systems for radiation oncology, considerations in room design and construction in a RO department, shared use with diagnostic radiology, staffing needs and training, clinical/research use cases and implementation, will be discussed. The emphasis will be on understanding and bridging the differences between diagnostic and radiation oncology installations, building consensus amongst stakeholders for purchase and use, and integrating imaging technologies into the radiation oncology environment. Learning Objectives: Learn the latest innovations of major imaging systems relevant to radiation therapy Be able to describe important technical specifications of CT, MRI

  7. Multiplexing 32,000 spectra onto 8 detectors: the HARMONI field splitting, image slicing, and wavelength selecting optics

    Science.gov (United States)

    Tecza, Matthias; Thatte, Niranjan; Clarke, Fraser; Freeman, David; Kosmalski, Johan

    2012-09-01

    HARMONI, the High Angular Resolution Monolithic Optical & Near-infrared Integral field spectrograph is one of two first-light instruments for the European Extremely Large Telescope. Over a 256x128 pixel field-of-view HARMONI will simultaneously measure approximately 32,000 spectra. Each spectrum is about 4000 spectral pixels long, and covers a selectable part of the 0.47-2.45 μm wavelength range at resolving powers of either R≍4000, 10000, or 20000. All 32,000 spectra are imaged onto eight HAWAII4RG detectors using a multiplexing scheme that divides the input field into four sub-fields, each imaged onto one image slicer that in turn re-arranges a single sub-field into two long exit slits feeding one spectrograph each. In total we require eight spectrographs, each with one HAWAII4RG detector. A system of articulated and exchangeable fold-mirrors and VPH gratings allows one to select different spectral resolving powers and wavelength ranges of interest while keeping a fixed geometry between the spectrograph collimator and camera avoiding the need for an articulated grating and camera. In this paper we describe both the field splitting and image slicing optics as well as the optics that will be used to select both spectral resolving power and wavelength range.

  8. Selection of Quantum Dot Wavelengths for Biomedical Assays and Imaging

    Directory of Open Access Journals (Sweden)

    Yong Taik Lim

    2003-01-01

    Full Text Available Fluorescent semiconductor nanocrystals (quantum dots [QDs] are hypothesized to be excellent contrast agents for biomedical assays and imaging. A unique property of QDs is that their absorbance increases with increasing separation between excitation and emission wavelengths. Much of the enthusiasm for using QDs in vivo stems from this property, since photon yield should be proportional to the integral of the broadband absorption. In this study, we demonstrate that tissue scatter and absorbance can sometimes offset increasing QD absorption at bluer wavelengths, and counteract this potential advantage. By using a previously validated mathematical model, we explored the effects of tissue absorbance, tissue scatter, wavelength dependence of the scatter, water-to- hemoglobin ratio, and tissue thickness on QD performance. We conclude that when embedded in biological fluids and tissues, QD excitation wavelengths will often be quite constrained, and that excitation and emission wavelengths should be selected carefully based on the particular application. Based on our results, we produced near-infrared QDs optimized for imaging surface vasculature with white light excitation and a silicon CCD camera, and used them to image the coronary vasculature in vivo. Taken together, our data should prove useful in designing fluorescent QD contrast agents optimized for specific biomedical applications.

  9. Selected topics in image science

    International Nuclear Information System (INIS)

    Nalcioglu, O.; Cho, Z.H.

    1984-01-01

    A review of the state of the art in diagnostic imaging via computers. Applications covered include emission tomography, digital radiography, and ultrasound and nuclear magnetic resonance imaging. Contents, abridged: Direct Fourier reconstruction techniques. Radiation detectors for CT instrumentation. Single photon emission computed tomography: potentials and limitations. Matched filtering for digital subtraction angiography

  10. THE SYSTEMATICS OF STRONG LENS MODELING QUANTIFIED: THE EFFECTS OF CONSTRAINT SELECTION AND REDSHIFT INFORMATION ON MAGNIFICATION, MASS, AND MULTIPLE IMAGE PREDICTABILITY

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, Traci L.; Sharon, Keren, E-mail: tljohn@umich.edu [University of Michigan, Department of Astronomy, 1085 South University Avenue, Ann Arbor, MI 48109-1107 (United States)

    2016-11-20

    Until now, systematic errors in strong gravitational lens modeling have been acknowledged but have never been fully quantified. Here, we launch an investigation into the systematics induced by constraint selection. We model the simulated cluster Ares 362 times using random selections of image systems with and without spectroscopic redshifts and quantify the systematics using several diagnostics: image predictability, accuracy of model-predicted redshifts, enclosed mass, and magnification. We find that for models with >15 image systems, the image plane rms does not decrease significantly when more systems are added; however, the rms values quoted in the literature may be misleading as to the ability of a model to predict new multiple images. The mass is well constrained near the Einstein radius in all cases, and systematic error drops to <2% for models using >10 image systems. Magnification errors are smallest along the straight portions of the critical curve, and the value of the magnification is systematically lower near curved portions. For >15 systems, the systematic error on magnification is ∼2%. We report no trend in magnification error with the fraction of spectroscopic image systems when selecting constraints at random; however, when using the same selection of constraints, increasing this fraction up to ∼0.5 will increase model accuracy. The results suggest that the selection of constraints, rather than quantity alone, determines the accuracy of the magnification. We note that spectroscopic follow-up of at least a few image systems is crucial because models without any spectroscopic redshifts are inaccurate across all of our diagnostics.

  11. Sensitivity of hemozoin detection by automated flow cytometry in non- and semi-immune malaria patients

    NARCIS (Netherlands)

    Grobusch, Martin P.; Hänscheid, Thomas; Krämer, Benedikt; Neukammer, Jörg; May, Jürgen; Seybold, Joachim; Kun, Jürgen F. J.; Suttorp, Norbert

    2003-01-01

    BACKGROUND: Cell-Dyn automated blood cell analyzers use laser flow cytometry technology, allowing detection of malaria pigment (hemozoin) in monocytes. We evaluated the value of such an instrument to diagnose malaria in febrile travelers returning to Berlin, Germany, the relation between the

  12. Antimicrobial Activity of Rhoeo discolor Phenolic Rich Extracts Determined by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Rebeca García-Varela

    2015-10-01

    Full Text Available Traditional medicine has led to the discovery of important active substances used in several health-related areas. Phytochemicals in Rhoeo discolor extracts have proven to have important antimicrobial activity. In the present study, our group determined the antimicrobial effects of extracts of Rhoeo discolor, a plant commonly used in Mexico for both medicinal and ornamental purposes. We evaluated the in vitro activity of phenolic rich extracts against specifically chosen microorganisms of human health importance by measuring their susceptibility via agar-disc diffusion assay and flow cytometry: Gram-positive Listeria innocua and Streptococcus mutans, Gram-negative Escherichia coli and Pseudomonas aeruginosa, and lastly a fungal pathogen Candida albicans. Ten different extracts were tested in eight different doses on all the microorganisms. Analytical data revealed a high content of phenolic compounds. Both agar-disc diffusion assay and flow cytometry results demonstrated that Pseudomonas aeruginosa was the least affected by extract exposure. However, low doses of these extracts (predominantly polar, in a range from 1 to 4 μg/mL, did produce a statistically significant bacteriostatic and bactericidal effect on the rest of the microorganisms. These results suggest the addition of certain natural extracts from Rhoeo discolor could act as antibacterial and antimycotic drugs or additives for foods and cosmetics.

  13. SU-E-J-128: Two-Stage Atlas Selection in Multi-Atlas-Based Image Segmentation

    International Nuclear Information System (INIS)

    Zhao, T; Ruan, D

    2015-01-01

    Purpose: In the new era of big data, multi-atlas-based image segmentation is challenged by heterogeneous atlas quality and high computation burden from extensive atlas collection, demanding efficient identification of the most relevant atlases. This study aims to develop a two-stage atlas selection scheme to achieve computational economy with performance guarantee. Methods: We develop a low-cost fusion set selection scheme by introducing a preliminary selection to trim full atlas collection into an augmented subset, alleviating the need for extensive full-fledged registrations. More specifically, fusion set selection is performed in two successive steps: preliminary selection and refinement. An augmented subset is first roughly selected from the whole atlas collection with a simple registration scheme and the corresponding preliminary relevance metric; the augmented subset is further refined into the desired fusion set size, using full-fledged registration and the associated relevance metric. The main novelty of this work is the introduction of an inference model to relate the preliminary and refined relevance metrics, based on which the augmented subset size is rigorously derived to ensure the desired atlases survive the preliminary selection with high probability. Results: The performance and complexity of the proposed two-stage atlas selection method were assessed using a collection of 30 prostate MR images. It achieved comparable segmentation accuracy as the conventional one-stage method with full-fledged registration, but significantly reduced computation time to 1/3 (from 30.82 to 11.04 min per segmentation). Compared with alternative one-stage cost-saving approach, the proposed scheme yielded superior performance with mean and medium DSC of (0.83, 0.85) compared to (0.74, 0.78). Conclusion: This work has developed a model-guided two-stage atlas selection scheme to achieve significant cost reduction while guaranteeing high segmentation accuracy. The benefit

  14. Usefullness of IGH/TCR PCR studies in lymphoproliferative disorders with inconclusive clonality by flow cytometry.

    Science.gov (United States)

    Ribera, Jordi; Zamora, Lurdes; Juncà, Jordi; Rodríguez, Inés; Marcé, Silvia; Cabezón, Marta; Millá, Fuensanta

    2013-07-25

    In up to 5-15% of studies of lymphoproliferative disorders (LPD) flow cytometry (FCM) or immunomorphologic methods cannot discriminate malignant from reactive processes. The aim of this work was to determine the usefulness of PCR for solving these diagnostic uncertainties. We analyzed IGH and TCRγ genes by PCR in 106 samples with inconclusive FCM results. A clonal result was registered in 36/106 studies, with a LPD being confirmed in 27 (75%) of these cases. Specifically, 9/9 IGH clonal and 16/25 TCRγ clonal results were finally diagnosed with LPD. Additionally, 2 clonal TCRγ samples with suspicion of undefined LPD were finally diagnosed with T LPD. Although polyclonal results were obtained in 47 of the cases studied (38 IGH and 9 TCRγ), hematologic neoplasms were diagnosed in 4/38 IGH polyclonal and in 1/9 TCRγ polyclonal studies. There were also 14 PCR polyclonal results (4 IGH, 10 TCRγ), albeit non-conclusive. Of these, 2/4 were eventually diagnosed with B-cell lymphoma and 3/10 with T-cell LPD. In 8 IGH samples the results of PCR techniques were non-informative but in 3/8 cases a B lymphoma was finally confirmed. We concluded that PCR is a useful technique to identify LPD when FCM is inconclusive. A PCR clonal B result is indicative of malignancy but IGH polyclonal and non-conclusive results do not exclude lymphoid neoplasms. Interpretation of T-cell clonality should be based on all the available clinical and analytical data. © 2013 Clinical Cytometry Society. Copyright © 2013 Clinical Cytometry Society.

  15. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods.

    Science.gov (United States)

    Øbro, Nina F; Ryder, Lars P; Madsen, Hans O; Andersen, Mette K; Lausen, Birgitte; Hasle, Henrik; Schmiegelow, Kjeld; Marquart, Hanne V

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and/or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative immunophenotype and antigen modulation) that highlight important methodological pitfalls. These findings demonstrate that with sufficient experience, flow cytometry is reliable for minimal residual disease monitoring in B-cell precursor acute lymphoblastic leukemia, although rare cases require supplementary PCR-based monitoring.

  16. Integrated optical waveguides and inertial focussing microfluidics in silica for microflow cytometry applications

    International Nuclear Information System (INIS)

    Butement, Jonathan T; Rowe, David J; Sessions, Neil P; Hua, Ping; Murugan, G Senthil; Wilkinson, James S; Clark, Owain; Chad, John E; Hunt, Hamish C

    2016-01-01

    A key challenge in the development of a microflow cytometry platform is the integration of the optical components with the fluidics as this requires compatible micro-optical and microfluidic technologies. In this work a microflow cytometry platform is presented comprising monolithically integrated waveguides and deep microfluidics in a rugged silica chip. Integrated waveguides are used to deliver excitation light to an etched microfluidic channel and also collect transmitted light. The fluidics are designed to employ inertial focussing, a particle positioning technique, to reduce signal variation by bringing the flowing particles onto the same plane as the excitation light beam. A fabrication process is described which exploits microelectronics mass production techniques including: sputtering, ICP etching and PECVD. Example devices were fabricated and the effectiveness of inertial focussing of 5.6 µ m fluorescent beads was studied showing lateral and vertical confinement of flowing beads within the microfluidic channel. The fluorescence signals from flowing calibration beads were quantified demonstrating a CV of 26%. Finally the potential of this type of device for measuring the variation in optical transmission from input to output waveguide as beads flowed through the beam was evaluated. (paper)

  17. Neutron Imaging for Selective Laser Melting Inconel Hardware with Internal Passages

    Science.gov (United States)

    Tramel, Terri L.; Norwood, Joseph K.; Bilheux, Hassina

    2014-01-01

    Additive Manufacturing is showing great promise for the development of new innovative designs and large potential life cycle cost reduction for the Aerospace Industry. However, more development work is required to move this technology into space flight hardware production. With selective laser melting (SLM), hardware that once consisted of multiple, carefully machined and inspected pieces, joined together can be made in one part. However standard inspection techniques cannot be used to verify that the internal passages are within dimensional tolerances or surface finish requirements. NASA/MSFC traveled to Oak Ridge National Lab's (ORNL) Spallation Neutron Source to perform some non-destructive, proof of concept imaging measurements to assess the capabilities to understand internal dimensional tolerances and internal passages surface roughness. This presentation will describe 1) the goals of this proof of concept testing, 2) the lessons learned when designing and building these Inconel 718 test specimens to minimize beam time, 3) the neutron imaging test setup and test procedure to get the images, 4) the initial results in images, volume and a video, 4) the assessment of using this imaging technique to gather real data for designing internal flow passages in SLM manufacturing aerospace hardware, and lastly 5) how proper cleaning of the internal passages is critically important. In summary, the initial results are very promising and continued development of a technique to assist in SLM development for aerospace components is desired by both NASA and ORNL. A plan forward that benefits both ORNL and NASA will also be presented, based on the promising initial results. The initial images and volume reconstruction showed that clean, clear images of the internal passages geometry are obtainable. These clear images of the internal passages of simple geometries will be compared to the build model to determine any differences. One surprising result was that a new cleaning

  18. An assessment of software for flow cytometry analysis in banana plants

    Directory of Open Access Journals (Sweden)

    Renata Alves Lara Silva

    2014-02-01

    Full Text Available Flow cytometry is a technique that yields rapid results in analyses of cell properties such as volume, morphological complexity and quantitative DNA content, and it is considered more convenient than other techniques. However, the analysis usually generates histograms marked by variations that can be produced by many factors, including differences between the software packages that capture the data generated by the flow cytometer. The objective of the present work was to evaluate the performance of four software products commonly used in flow cytometry based on quantifications of DNA content and analyses of the coefficients of variation associated with the software outputs. Readings were obtained from 25 ‘NBA’ (AA banana leaf samples using the FACSCalibur (BD flow cytometer, and 25 histograms from each software product (CellQuest™, WinMDI™, FlowJo™ and FCS Express™ were analyzed to obtain the estimated DNA content and the coefficient of variation (CV of the estimates. The values of DNA content obtained from the software did not differ significantly. However, the CV analysis showed that the precision of the WinMDI™ software was low and that the CV values were underestimated, whereas the remaining software showed CV values that were in relatively close agreement with those found in the literature. The CellQuest™ software is recommended because it was developed by the same company that produces the flow cytometer used in the present study.

  19. Label fusion based brain MR image segmentation via a latent selective model

    Science.gov (United States)

    Liu, Gang; Guo, Xiantang; Zhu, Kai; Liao, Hengxu

    2018-04-01

    Multi-atlas segmentation is an effective approach and increasingly popular for automatically labeling objects of interest in medical images. Recently, segmentation methods based on generative models and patch-based techniques have become the two principal branches of label fusion. However, these generative models and patch-based techniques are only loosely related, and the requirement for higher accuracy, faster segmentation, and robustness is always a great challenge. In this paper, we propose novel algorithm that combines the two branches using global weighted fusion strategy based on a patch latent selective model to perform segmentation of specific anatomical structures for human brain magnetic resonance (MR) images. In establishing this probabilistic model of label fusion between the target patch and patch dictionary, we explored the Kronecker delta function in the label prior, which is more suitable than other models, and designed a latent selective model as a membership prior to determine from which training patch the intensity and label of the target patch are generated at each spatial location. Because the image background is an equally important factor for segmentation, it is analyzed in label fusion procedure and we regard it as an isolated label to keep the same privilege between the background and the regions of interest. During label fusion with the global weighted fusion scheme, we use Bayesian inference and expectation maximization algorithm to estimate the labels of the target scan to produce the segmentation map. Experimental results indicate that the proposed algorithm is more accurate and robust than the other segmentation methods.

  20. [Clinical usefulness of urine-formed elements' information obtained from bacteria detection by flow cytometry method that uses nucleic acid staining].

    Science.gov (United States)

    Nakagawa, Hiroko; Yuno, Tomoji; Itho, Kiichi

    2009-03-01

    Recently, specific detection method for Bacteria, by flow cytometry method using nucleic acid staining, was developed as a function of automated urine formed elements analyzer for routine urine testing. Here, we performed a basic study on this bacteria analysis method. In addition, we also have a comparison among urine sediment analysis, urine Gram staining and urine quantitative cultivation, the conventional methods performed up to now. As a result, the bacteria analysis with flow cytometry method that uses nucleic acid staining was excellent in reproducibility, and higher sensitivity compared with microscopic urinary sediment analysis. Based on the ROC curve analysis, which settled urine culture method as standard, cut-off level of 120/microL was defined and its sensitivity = 85.7%, specificity = 88.2%. In the analysis of scattergram, accompanied with urine culture method, among 90% of rod positive samples, 80% of dots were appeared in the area of 30 degrees from axis X. In addition, one case even indicated that analysis of bacteria by flow cytometry and scattergram of time series analysis might be helpful to trace the progress of causative bacteria therefore the information supposed to be clinically significant. Reporting bacteria information with nucleic acid staining flow cytometry method is expected to contribute to a rapid diagnostics and treatment of urinary tract infections. Besides, the contribution to screening examination of microbiology and clinical chemistry, will deliver a more efficient solution to urine analysis.

  1. [Research on K-means clustering segmentation method for MRI brain image based on selecting multi-peaks in gray histogram].

    Science.gov (United States)

    Chen, Zhaoxue; Yu, Haizhong; Chen, Hao

    2013-12-01

    To solve the problem of traditional K-means clustering in which initial clustering centers are selected randomly, we proposed a new K-means segmentation algorithm based on robustly selecting 'peaks' standing for White Matter, Gray Matter and Cerebrospinal Fluid in multi-peaks gray histogram of MRI brain image. The new algorithm takes gray value of selected histogram 'peaks' as the initial K-means clustering center and can segment the MRI brain image into three parts of tissue more effectively, accurately, steadily and successfully. Massive experiments have proved that the proposed algorithm can overcome many shortcomings caused by traditional K-means clustering method such as low efficiency, veracity, robustness and time consuming. The histogram 'peak' selecting idea of the proposed segmentootion method is of more universal availability.

  2. Assessment of ploidy stability of the somatic embryogenesis process in Quercus suber L. using flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Loureiro, J.; Pinto, G.; Lopes, T.; Doležel, Jaroslav; Santos, C.

    2005-01-01

    Roč. 221, - (2005), s. 815-822 ISSN 0032-0935 Institutional research plan: CEZ:AV0Z50380511 Keywords : Flow cytometry * ploidy stability * nuclear DNA content Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.108, year: 2005

  3. Behaviors study of image registration algorithms in image guided radiation therapy

    International Nuclear Information System (INIS)

    Zou Lian; Hou Qing

    2008-01-01

    Objective: Study the behaviors of image registration algorithms, and analyze the elements which influence the performance of image registrations. Methods: Pre-known corresponding coordinates were appointed for reference image and moving image, and then the influence of region of interest (ROI) selection, transformation function initial parameters and coupled parameter spaces on registration results were studied with a software platform developed in home. Results: Region of interest selection had a manifest influence on registration performance. An improperly chosen ROI resulted in a bad registration. Transformation function initial parameters selection based on pre-known information could improve the accuracy of image registration. Coupled parameter spaces would enhance the dependence of image registration algorithm on ROI selection. Conclusions: It is necessary for clinic IGRT to obtain a ROI selection strategy (depending on specific commercial software) correlated to tumor sites. Three suggestions for image registration technique developers are automatic selection of the initial parameters of transformation function based on pre-known information, developing specific image registration algorithm for specific image feature, and assembling real-time image registration algorithms according to tumor sites selected by software user. (authors)

  4. T cell homing to tumors detected by 3D-coordinated positron emission tomography and magnetic resonance imaging

    DEFF Research Database (Denmark)

    Agger, Ralf; Petersen, Mikkel; Petersen, Charlotte Christie

    2007-01-01

    of magnetic resonance imaging with the high sensitivity and spatial accuracy of positron emission tomography. We have used this technique, together with determination of tissue radioactivity, flow cytometry, and microscopy, to characterize and quantitate the specific accumulation of transferred CD8+ T cells...

  5. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods

    DEFF Research Database (Denmark)

    Obro, Nina F; Ryder, Lars P; Madsen, Hans O

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring...... clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and....../or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative...

  6. Assessment of relevant hepatic steatosis in obese adolescents by rapid fat-selective GRE imaging with spatial-spectral excitation: a quantitative comparison with spectroscopic findings

    International Nuclear Information System (INIS)

    Springer, Fabian; Schick, Fritz; Ehehalt, Stefan; Binder, Gerhard; Sommer, Julia; Ballweg, Verena; Machann, Juergen; Claussen, Claus D.

    2011-01-01

    To test the feasibility of fat-selective GRE imaging using a spectral-spatial excitation technique for determination of intrahepatic lipid content (IHL) in obese adolescents. Fat-selective MR imaging (1.5 T) was applied to record a single axial slice through a representative liver region within a single breath-hold. The sequence uses six equidistant slice-selective excitation pulses with binomial amplitude ratios to achieve high selectivity for lipid signals after appropriate shimming. IHL MRI content was quantified using signal intensity of adjacent subcutaneous adipose tissue. As the gold standard for IHL quantification, single-voxel stimulated echo magnetic resonance spectroscopy (MRS) was applied. IHL MRS was quantified using the water peak as a reference. Forty-five MR examinations could be performed, and IHL MRS content ranged from 0.7% to 19.1%. Results from MRS and fat-selective imaging correlated well with Spearman coefficients between r = 0.78 and r = 0.86. There were no relevant regional differences in IHL within the liver parenchyma (p > 0.6359). Fat-selective imaging was able to reliably identify patients with IHL content above 5% with positive/negative likelihood ratio of 11.8 and 0.05, respectively. Fat-selective MR imaging provides both a reliable and a convenient method of rapidly quantifying IHL content in obese adolescents. (orig.)

  7. Advantages of flow cytometry immunophenotyping for the diagnosis of central nervous system non-Hodgkin's lymphoma in AIDS patients.

    Science.gov (United States)

    Subirá, D; Górgolas, M; Castañón, S; Serrano, C; Román, A; Rivas, F; Tomás, J F

    2005-01-01

    Neurological disorders are common in HIV-infected patients. Central nervous system (CNS) lymphoma should always be considered because it is an important cause of morbidity and mortality. To investigate the clinical utility of flow cytometry immunophenotyping (FCI) in diagnosing or discarding leptomeningeal involvement in HIV-infected patients and to compare its sensitivity with that of conventional cytological methods. Fifty-six cerebrospinal fluid (CSF) samples from 29 HIV-infected patients were independently evaluated by flow cytometry and cytology. The description of an aberrant immunophenotype was the criterion used to define the malignant nature of any CSF cell population. FCI and cytology gave concordant results for 48 of the 56 CSF samples studied: 37 were negative for malignancy and 11 had evidence of CNS lymphoma. Discordant results were obtained for eight CSF samples, and the accuracy of the FCI findings could be demonstrated for four CSF samples described as positive for malignancy according to the FCI criteria. A high level of agreement was found between the results obtained using the two methods, but FCI gave at least 25% higher sensitivity than conventional cytomorphological methods for the detection of malignant cells. This advantage suggests that, in case of negative flow cytometry results, disorders other than non-Hodgkin's lymphoma should be strongly considered.

  8. Depth-Based Selective Blurring in Stereo Images Using Accelerated Framework

    Science.gov (United States)

    Mukherjee, Subhayan; Guddeti, Ram Mohana Reddy

    2014-09-01

    We propose a hybrid method for stereo disparity estimation by combining block and region-based stereo matching approaches. It generates dense depth maps from disparity measurements of only 18 % image pixels (left or right). The methodology involves segmenting pixel lightness values using fast K-Means implementation, refining segment boundaries using morphological filtering and connected components analysis; then determining boundaries' disparities using sum of absolute differences (SAD) cost function. Complete disparity maps are reconstructed from boundaries' disparities. We consider an application of our method for depth-based selective blurring of non-interest regions of stereo images, using Gaussian blur to de-focus users' non-interest regions. Experiments on Middlebury dataset demonstrate that our method outperforms traditional disparity estimation approaches using SAD and normalized cross correlation by up to 33.6 % and some recent methods by up to 6.1 %. Further, our method is highly parallelizable using CPU-GPU framework based on Java Thread Pool and APARAPI with speed-up of 5.8 for 250 stereo video frames (4,096 × 2,304).

  9. Noninvasive imaging of multiple myeloma using near infrared fluorescent molecular probe

    Science.gov (United States)

    Hathi, Deep; Zhou, Haiying; Bollerman-Nowlis, Alex; Shokeen, Monica; Akers, Walter J.

    2016-03-01

    Multiple myeloma is a plasma cell malignancy characterized by monoclonal gammopathy and osteolytic bone lesions. Multiple myeloma is most commonly diagnosed in late disease stages, presenting with pathologic fracture. Early diagnosis and monitoring of disease status may improve quality of life and long-term survival for multiple myeloma patients from what is now a devastating and fatal disease. We have developed a near-infrared targeted fluorescent molecular probe with high affinity to the α4β1 integrin receptor (VLA-4)overexpressed by a majority of multiple myeloma cells as a non-radioactive analog to PET/CT tracer currently being developed for human diagnostics. A near-infrared dye that emits about 700 nm was conjugated to a high affinity peptidomimmetic. Binding affinity and specificity for multiple myeloma cells was investigated in vitro by tissue staining and flow cytometry. After demonstration of sensitivity and specificity, preclinical optical imaging studies were performed to evaluate tumor specificity in murine subcutaneous and metastatic multiple myeloma models. The VLA-4-targeted molecular probe showed high affinity for subcutaneous MM tumor xenografts. Importantly, tumor cells specific accumulation in the bone marrow of metastatic multiple myeloma correlated with GFP signal from transfected cells. Ex vivo flow cytometry of tumor tissue and bone marrow further corroborated in vivo imaging data, demonstrating the specificity of the novel agent and potential for quantitative imaging of multiple myeloma burden in these models.

  10. Bis-pyridinium quadrupolar derivatives. High Stokes shift selective probes for bio-imaging

    Science.gov (United States)

    Salice, Patrizio; Versari, Silvia; Bradamante, Silvia; Meinardi, Francesco; Macchi, Giorgio; Pagani, Giorgio A.; Beverina, Luca

    2013-11-01

    We describe the design, synthesis and characterization of five high Stokes shift quadrupolar heteroaryl compounds suitable as fluorescent probes in bio-imaging. In particular, we characterize the photophysical properties and the intracellular localization in Human Umbilical Vein Endothelial Cells (HUVEC) and Human Mesenchymal Stem Cells (HMSCs) for each dye. We show that, amongst all of the investigated derivatives, the 2,5-bis[1-(4-N-methylpyridinium)ethen-2-yl)]- N-methylpyrrole salt is the best candidates as selective mitochondrial tracker. Finally, we recorded the full emission spectrum of the most performing - exclusively mitochondrial selective - fluorescent probe directly from HUVEC stained cells. The emission spectrum collected from the stained mitochondria shows a remarkably more pronounced vibronic structure with respect to the emission of the free fluorophore in solution.

  11. Profiling pleural effusion cells by a diffraction imaging method

    Science.gov (United States)

    Al-Qaysi, Safaa; Hong, Heng; Wen, Yuhua; Lu, Jun Q.; Feng, Yuanming; Hu, Xin-Hua

    2018-02-01

    Assay of cells in pleural effusion (PE) is an important means of disease diagnosis. Conventional cytology of effusion samples, however, has low sensitivity and depends heavily on the expertise of cytopathologists. We applied a polarization diffraction imaging flow cytometry method on effusion cells to investigate their features. Diffraction imaging of the PE cell samples has been performed on 6000 to 12000 cells for each effusion cell sample of three patients. After prescreening to remove images by cellular debris and aggregated non-cellular particles, the image textures were extracted with a gray level co-occurrence matrix (GLCM) algorithm. The distribution of the imaged cells in the GLCM parameters space was analyzed by a Gaussian Mixture Model (GMM) to determine the number of clusters among the effusion cells. These results yield insight on textural features of diffraction images and related cellular morphology in effusion samples and can be used toward the development of a label-free method for effusion cells assay.

  12. Development of an unbiased, semi-automated approach for classifying plasma cell immunophenotype following multicolor flow cytometry of bone marrow aspirates.

    Science.gov (United States)

    Post, Steven R; Post, Ginell R; Nikolic, Dejan; Owens, Rebecca; Insuasti-Beltran, Giovanni

    2018-03-24

    Despite increased usage of multiparameter flow cytometry (MFC) to assess diagnosis, prognosis, and therapeutic efficacy (minimal residual disease, MRD) in plasma cell neoplasms (PCNs), standardization of methodology and data analysis is suboptimal. We investigated the utility of using the mean and median fluorescence intensities (FI) obtained from MFC to objectively describe parameters that distinguish plasma cell (PC) phenotypes. In this retrospective study, flow cytometry results from bone marrow aspirate specimens from 570 patients referred to the Myeloma Institute at UAMS were evaluated. Mean and median FI data were obtained from 8-color MFC of non-neoplastic, malignant, and mixed PC populations using antibodies to CD38, CD138, CD19, CD20, CD27, CD45, CD56, and CD81. Of 570 cases, 252 cases showed only non-neoplastic PCs, 168 showed only malignant PCs, and 150 showed mixed PC populations. Statistical analysis of median FI data for each CD marker showed no difference in expression intensity on non-neoplastic and malignant PCs, between pure and mixed PC populations. ROC analysis of the median FI of CD expression in non-neoplastic and malignant PCs was used to develop an algorithm to convert quantitative FI values to qualitative assessments including "negative," "positive," "dim," and "heterogeneous" expression. FI data derived from 8-color MFC can be used to define marker expression on PCs. Translation of FI data from Infinicyt software to an Excel worksheet streamlines workflow and eliminates transcriptional errors when generating flow reports. © 2018 International Clinical Cytometry Society. © 2018 International Clinical Cytometry Society.

  13. Selecting a separable parametric spatiotemporal covariance structure for longitudinal imaging data.

    Science.gov (United States)

    George, Brandon; Aban, Inmaculada

    2015-01-15

    Longitudinal imaging studies allow great insight into how the structure and function of a subject's internal anatomy changes over time. Unfortunately, the analysis of longitudinal imaging data is complicated by inherent spatial and temporal correlation: the temporal from the repeated measures and the spatial from the outcomes of interest being observed at multiple points in a patient's body. We propose the use of a linear model with a separable parametric spatiotemporal error structure for the analysis of repeated imaging data. The model makes use of spatial (exponential, spherical, and Matérn) and temporal (compound symmetric, autoregressive-1, Toeplitz, and unstructured) parametric correlation functions. A simulation study, inspired by a longitudinal cardiac imaging study on mitral regurgitation patients, compared different information criteria for selecting a particular separable parametric spatiotemporal correlation structure as well as the effects on types I and II error rates for inference on fixed effects when the specified model is incorrect. Information criteria were found to be highly accurate at choosing between separable parametric spatiotemporal correlation structures. Misspecification of the covariance structure was found to have the ability to inflate the type I error or have an overly conservative test size, which corresponded to decreased power. An example with clinical data is given illustrating how the covariance structure procedure can be performed in practice, as well as how covariance structure choice can change inferences about fixed effects. Copyright © 2014 John Wiley & Sons, Ltd.

  14. Flow Cytometry Pulse Width Data Enables Rapid and Sensitive Estimation of Biomass Dry Weight in the Microalgae Chlamydomonas reinhardtii and Chlorella vulgaris

    Science.gov (United States)

    Chioccioli, Maurizio; Hankamer, Ben; Ross, Ian L.

    2014-01-01

    Dry weight biomass is an important parameter in algaculture. Direct measurement requires weighing milligram quantities of dried biomass, which is problematic for small volume systems containing few cells, such as laboratory studies and high throughput assays in microwell plates. In these cases indirect methods must be used, inducing measurement artefacts which vary in severity with the cell type and conditions employed. Here, we utilise flow cytometry pulse width data for the estimation of cell density and biomass, using Chlorella vulgaris and Chlamydomonas reinhardtii as model algae and compare it to optical density methods. Measurement of cell concentration by flow cytometry was shown to be more sensitive than optical density at 750 nm (OD750) for monitoring culture growth. However, neither cell concentration nor optical density correlates well to biomass when growth conditions vary. Compared to the growth of C. vulgaris in TAP (tris-acetate-phosphate) medium, cells grown in TAP + glucose displayed a slowed cell division rate and a 2-fold increased dry biomass accumulation compared to growth without glucose. This was accompanied by increased cellular volume. Laser scattering characteristics during flow cytometry were used to estimate cell diameters and it was shown that an empirical but nonlinear relationship could be shown between flow cytometric pulse width and dry weight biomass per cell. This relationship could be linearised by the use of hypertonic conditions (1 M NaCl) to dehydrate the cells, as shown by density gradient centrifugation. Flow cytometry for biomass estimation is easy to perform, sensitive and offers more comprehensive information than optical density measurements. In addition, periodic flow cytometry measurements can be used to calibrate OD750 measurements for both convenience and accuracy. This approach is particularly useful for small samples and where cellular characteristics, especially cell size, are expected to vary during growth. PMID

  15. Parallel imaging microfluidic cytometer.

    Science.gov (United States)

    Ehrlich, Daniel J; McKenna, Brian K; Evans, James G; Belkina, Anna C; Denis, Gerald V; Sherr, David H; Cheung, Man Ching

    2011-01-01

    By adding an additional degree of freedom from multichannel flow, the parallel microfluidic cytometer (PMC) combines some of the best features of fluorescence-activated flow cytometry (FCM) and microscope-based high-content screening (HCS). The PMC (i) lends itself to fast processing of large numbers of samples, (ii) adds a 1D imaging capability for intracellular localization assays (HCS), (iii) has a high rare-cell sensitivity, and (iv) has an unusual capability for time-synchronized sampling. An inability to practically handle large sample numbers has restricted applications of conventional flow cytometers and microscopes in combinatorial cell assays, network biology, and drug discovery. The PMC promises to relieve a bottleneck in these previously constrained applications. The PMC may also be a powerful tool for finding rare primary cells in the clinic. The multichannel architecture of current PMC prototypes allows 384 unique samples for a cell-based screen to be read out in ∼6-10 min, about 30 times the speed of most current FCM systems. In 1D intracellular imaging, the PMC can obtain protein localization using HCS marker strategies at many times for the sample throughput of charge-coupled device (CCD)-based microscopes or CCD-based single-channel flow cytometers. The PMC also permits the signal integration time to be varied over a larger range than is practical in conventional flow cytometers. The signal-to-noise advantages are useful, for example, in counting rare positive cells in the most difficult early stages of genome-wide screening. We review the status of parallel microfluidic cytometry and discuss some of the directions the new technology may take. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Remarkable coexistence of multiple cytotypes of the Gymnadenia conopsea aggregate (the fragrant orchid): evidence from flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Trávníček, Pavel; Kubátová, B.; Čurn, V.; Rauchová, Jana; Krajníková, E.; Jersáková, J.; Suda, Jan

    2011-01-01

    Roč. 107, č. 1 (2011), s. 77-87 ISSN 0305-7364 Institutional research plan: CEZ:AV0Z6005908 Keywords : coexistence * contact zone * flow cytometry Subject RIV: EF - Botanics Impact factor: 4.030, year: 2011

  17. Assessment of relevant hepatic steatosis in obese adolescents by rapid fat-selective GRE imaging with spatial-spectral excitation: a quantitative comparison with spectroscopic findings

    Energy Technology Data Exchange (ETDEWEB)

    Springer, Fabian; Schick, Fritz [University Hospital Tuebingen, Section on Experimental Radiology, Department of Diagnostic and Interventional Radiology, Tuebingen (Germany); University Hospital Tuebingen, Department of Diagnostic and Interventional Radiology, Tuebingen (Germany); Ehehalt, Stefan; Binder, Gerhard [University Children' s Hospital Tuebingen, Paediatric Endocrinology and Diabetes, Tuebingen (Germany); Sommer, Julia; Ballweg, Verena; Machann, Juergen [University Hospital Tuebingen, Section on Experimental Radiology, Department of Diagnostic and Interventional Radiology, Tuebingen (Germany); Claussen, Claus D. [University Hospital Tuebingen, Department of Diagnostic and Interventional Radiology, Tuebingen (Germany)

    2011-04-15

    To test the feasibility of fat-selective GRE imaging using a spectral-spatial excitation technique for determination of intrahepatic lipid content (IHL) in obese adolescents. Fat-selective MR imaging (1.5 T) was applied to record a single axial slice through a representative liver region within a single breath-hold. The sequence uses six equidistant slice-selective excitation pulses with binomial amplitude ratios to achieve high selectivity for lipid signals after appropriate shimming. IHL{sub MRI} content was quantified using signal intensity of adjacent subcutaneous adipose tissue. As the gold standard for IHL quantification, single-voxel stimulated echo magnetic resonance spectroscopy (MRS) was applied. IHL{sub MRS} was quantified using the water peak as a reference. Forty-five MR examinations could be performed, and IHL{sub MRS} content ranged from 0.7% to 19.1%. Results from MRS and fat-selective imaging correlated well with Spearman coefficients between r = 0.78 and r = 0.86. There were no relevant regional differences in IHL within the liver parenchyma (p > 0.6359). Fat-selective imaging was able to reliably identify patients with IHL content above 5% with positive/negative likelihood ratio of 11.8 and 0.05, respectively. Fat-selective MR imaging provides both a reliable and a convenient method of rapidly quantifying IHL content in obese adolescents. (orig.)

  18. A fast chaos-based image encryption scheme with a dynamic state variables selection mechanism

    Science.gov (United States)

    Chen, Jun-xin; Zhu, Zhi-liang; Fu, Chong; Yu, Hai; Zhang, Li-bo

    2015-03-01

    In recent years, a variety of chaos-based image cryptosystems have been investigated to meet the increasing demand for real-time secure image transmission. Most of them are based on permutation-diffusion architecture, in which permutation and diffusion are two independent procedures with fixed control parameters. This property results in two flaws. (1) At least two chaotic state variables are required for encrypting one plain pixel, in permutation and diffusion stages respectively. Chaotic state variables produced with high computation complexity are not sufficiently used. (2) The key stream solely depends on the secret key, and hence the cryptosystem is vulnerable against known/chosen-plaintext attacks. In this paper, a fast chaos-based image encryption scheme with a dynamic state variables selection mechanism is proposed to enhance the security and promote the efficiency of chaos-based image cryptosystems. Experimental simulations and extensive cryptanalysis have been carried out and the results prove the superior security and high efficiency of the scheme.

  19. Chip-Based Cytometry Illuminated by a Blade-Shape Continuous Light for Multispectral Detection

    Directory of Open Access Journals (Sweden)

    Shi-Wei Lin

    2016-08-01

    Full Text Available A high performance diascopic illumination configuration is presented for the simultaneous detection of cells and particles with different sizes and different fluorescence labels in a microchannel. In the proposed approach, the cells/particles are illuminated by an objective-type dark-field condenser equipped with a low-cost tungsten light source and are then characterized by extracting the side-scatter, absorbance, and fluorescence signals from the spectra obtained by a ultraviolet-visible-near infrared (UV-Vis-NIR spectrometer. A modified computation model is adopted to improve the capability for discriminating more fluorescence dyes simultaneously. The feasibility of the proposed detection configuration is demonstrated by counting and classifying a mixed sample of green, red, and crimson fluorescent-labeled particles and non-labeled particles with various dimensions. The suitability of the proposed system for real-world cytometry applications is then evaluated by classifying a mixed bio-sample comprising of gastric epithelial (AGS cells stained with Trypan-blue and Erythrosin-bluish dye, respectively. The results show that the cytometer enables the efficient detection, identification, and classification of mixed bio-samples without the need for spatial filters or delicate optical components. Consequently, the proposed system has significant potential for high-performance micro-flow cytometry applications.

  20. Super-resolution fluorescence imaging of nanoimprinted polymer patterns by selective fluorophore adsorption combined with redox switching

    KAUST Repository

    Yabiku, Y.

    2013-10-22

    We applied a super-resolution fluorescence imaging based on selective adsorption and redox switching of the fluorescent dye molecules for studying polymer nanostructures. We demonstrate that nano-scale structures of polymer thin films can be visualized with the image resolution better than 80 nm. The method was applied to image 100 nm-wide polymer nanopatterns fabricated by thermal nanoimprinting. The results point to the applicability of the method for evaluating residual polymer thin films and dewetting defect of the polymer resist patterns which are important for the quality control of the fine nanoimprinted patterns. 2013 Author(s).

  1. Super-resolution fluorescence imaging of nanoimprinted polymer patterns by selective fluorophore adsorption combined with redox switching

    Directory of Open Access Journals (Sweden)

    Yu Yabiku

    2013-10-01

    Full Text Available We applied a super-resolution fluorescence imaging based on selective adsorption and redox switching of the fluorescent dye molecules for studying polymer nanostructures. We demonstrate that nano-scale structures of polymer thin films can be visualized with the image resolution better than 80 nm. The method was applied to image 100 nm-wide polymer nanopatterns fabricated by thermal nanoimprinting. The results point to the applicability of the method for evaluating residual polymer thin films and dewetting defect of the polymer resist patterns which are important for the quality control of the fine nanoimprinted patterns.

  2. Full color emitting fluorescent carbon material as reversible pH sensor with multicolor live cell imaging.

    Science.gov (United States)

    Sharma, Vinay; Kaur, Navpreet; Tiwari, Pranav; Mobin, Shaikh M

    2018-05-01

    Carbon-based nano materials are developed as a cytocompatible alternative to semiconducting quantum dots for bioimaging and fluorescence-based sensing. The green alternatives for the synthesis of carbon materials are imminent. The present study demonstrates microwave based one step quick synthesis of fluorescent carbon material (FCM) having three variants: (i) un-doped fluorescent carbon material (UFCM) (ii) nitrogen doped FCM (N@FCM), and (iii) nitrogen & phosphorus co-doped FCM (N-P@FCM) using sugarcane extract as a carbon source. The N doping was performed using ethylenediamine and phosphoric acid was used for P doping. The heteroatom doped FCM were synthesized due to insolubility of UFCM in water. Unlike, UFCM, the N@FCM and N-P@FCM were found to be highly soluble in water. The N-P@FCM shows highest quantum yield among the three. The N-P@FCM was explored for alkaline pH sensing and it shows a quenching of fluorescence in the pH range 09-14. The sensing behaviour shows reversibility and high selectivity. Further, the sensor was also investigated for their biocompatibility and hence employed as a promising multicolour probe for cancer cell imaging. The generality in cell imaging was investigated by flow cytometry. The hetero-atom doped green carbon-dots may open new avenues for sensing and selective cellular targeting. Copyright © 2018 Elsevier B.V. All rights reserved.

  3. Phase-selective staining of metal salt for scanning electron microscopy imaging of block copolymer film

    Energy Technology Data Exchange (ETDEWEB)

    Li, Jing Ze, E-mail: Lijinge@uestc.edu.cn [State Key Laboratory of Electronic Thin Films and Integrated Devices, School of Microelectronic and Solid-state Electronic, University of Electronic Science and Technology of China, Chengdu 610054 (China); State Key Laboratory of Polymer Materials Engineering (Sichuan University), Chengdu 610054 (China); Xinjiang Key Laboratory of Electronic Information Materials and Devices, Urumuqi 830011 (China); Wang, Ying; Hong Wang, Zhi; Mei, Di; Zou, Wei [State Key Laboratory of Electronic Thin Films and Integrated Devices, School of Microelectronic and Solid-state Electronic, University of Electronic Science and Technology of China, Chengdu 610054 (China); Min Chang, Ai [State Key Laboratory of Polymer Materials Engineering (Sichuan University), Chengdu 610054 (China); Wang, Qi [Xinjiang Key Laboratory of Electronic Information Materials and Devices, Urumuqi 830011 (China); Komura, Motonori; Ito, Kaori [Division of Integrated Molecular Engineering, Chemical Resources Laboratory, Tokyo Institute of Technology, Yokohama 226-8503 (Japan); Iyoda, Tomokazu, E-mail: Iyoda.t.aa@m.titech.ac.jp [Division of Integrated Molecular Engineering, Chemical Resources Laboratory, Tokyo Institute of Technology, Yokohama 226-8503 (Japan)

    2010-09-15

    Three metal salts, i.e., AgNO{sub 3}, HAuCl{sub 4}, and KCl, were proposed as novel staining reagents instead of traditional RuO{sub 4} and OsO{sub 4} labeled with expensive price and extreme toxicity for scanning electron microscopy (SEM) imaging of microphase separated block copolymer film. A simple and costless aqueous solution immersion procedure could ensure selective staining of the metal slat in specific phase of the nanostructured copolymer film, leading to a clear phase contrasted SEM image. The heavy metal salt has better staining effect, demonstrating stable and high signal-to-noise SEM image even at an acceleration voltage as high as 30 kV and magnification up to 250,000 times.

  4. Flow cytometry as a rapid test for detection of penicillin resistance directly in bacterial cells in Enterococcus faecalis and Staphylococcus aureus.

    Science.gov (United States)

    Jarzembowski, T; Wiśniewska, K; Józwik, A; Bryl, E; Witkowski, J

    2008-08-01

    We studied the usefulness of flow cytometry for detection of penicillin resistance in E. faecalis and S. aureus by direct binding of commercially available fluorescent penicillin, Bocillin FL, to cells obtained from culture. There were significantly lower percentages of fluorescent cells and median and mean fluorescence values per particle in penicillin-resistant than in penicillin-sensitive strains of both species observed. The method allows rapid detection of penicillin resistance in S. aureus and E. faecalis. The results encourage further investigations on the detection of antibiotic resistance in bacteria using flow cytometry.

  5. Super-resolution fluorescence imaging of nanoimprinted polymer patterns by selective fluorophore adsorption combined with redox switching

    KAUST Repository

    Yabiku, Y.; Kubo, S.; Nakagawa, M.; Vacha, M.; Habuchi, Satoshi

    2013-01-01

    We applied a super-resolution fluorescence imaging based on selective adsorption and redox switching of the fluorescent dye molecules for studying polymer nanostructures. We demonstrate that nano-scale structures of polymer thin films can

  6. Tracking by flow cytometry antigen-specific follicular helper T cells in wild-type animals after protein vaccination.

    Science.gov (United States)

    Chakarov, Svetoslav; Fazilleau, Nicolas

    2015-01-01

    Flow cytometry is a valuable technology used in immunology to characterize and enumerate the different cell subpopulations specific for a nonself-antigen in the context of an ongoing immune response. Among them, follicular helper T cells are the cognate regulators of B cells in secondary lymphoid tissues. Thus, tracking them is of high interest especially in the context of protein vaccination. For this purpose, transgenic antigen-receptor mouse models have been largely used. It is now clear that transgenic models are not always the best means to study the dynamics of the immune response since they can modify the response. In this chapter, we describe how to track endogenous antigen-specific follicular helper T cells by flow cytometry after protein vaccination in nonmodified wild-type animals, which ultimately provides a comprehensive way to enumerate, characterize, and isolate these particular cells in vivo.

  7. Polyploidy in the Olive Complex (Olea europaea): Evidence from Flow Cytometry and Nuclear Microsatellite Analyses

    Science.gov (United States)

    Besnard, G.; Garcia-Verdugo, C.; Rubio De Casas, R.; Treier, U. A.; Galland, N.; Vargas, P.

    2008-01-01

    Background Phylogenetic and phylogeographic investigations have been previously performed to study the evolution of the olive tree complex (Olea europaea). A particularly high genomic diversity has been found in north-west Africa. However, to date no exhaustive study has been addressed to infer putative polyploidization events and their evolutionary significance in the diversification of the olive tree and its relatives. Methods Representatives of the six olive subspecies were investigated using (a) flow cytometry to estimate genome content, and (b) six highly variable nuclear microsatellites to assess the presence of multiple alleles at co-dominant loci. In addition, nine individuals from a controlled cross between two individuals of O. europaea subsp. maroccana were characterized with microsatellites to check for chromosome inheritance. Key Results Based on flow cytometry and genetic analyses, strong evidence for polyploidy was obtained in subspp. cerasiformis (tetraploid) and maroccana (hexaploid), whereas the other subspecies appeared to be diploids. Agreement between flow cytometry and genetic analyses gives an alternative approach to chromosome counting to determine ploidy level of trees. Lastly, abnormalities in chromosomes inheritance leading to aneuploid formation were revealed using microsatellite analyses in the offspring from the controlled cross in subsp. maroccana. Conclusions This study constitutes the first report for multiple polyploidy in olive tree relatives. Formation of tetraploids and hexaploids may have played a major role in the diversification of the olive complex in north-west Africa. The fact that polyploidy is found in narrow endemic subspecies from Madeira (subsp. cerasiformis) and the Agadir Mountains (subsp. maroccana) suggests that polyploidization has been favoured to overcome inbreeding depression. Lastly, based on previous phylogenetic analyses, we hypothesize that subsp. cerasiformis resulted from hybridization between ancestors

  8. DAFi: A directed recursive data filtering and clustering approach for improving and interpreting data clustering identification of cell populations from polychromatic flow cytometry data.

    Science.gov (United States)

    Lee, Alexandra J; Chang, Ivan; Burel, Julie G; Lindestam Arlehamn, Cecilia S; Mandava, Aishwarya; Weiskopf, Daniela; Peters, Bjoern; Sette, Alessandro; Scheuermann, Richard H; Qian, Yu

    2018-04-17

    Computational methods for identification of cell populations from polychromatic flow cytometry data are changing the paradigm of cytometry bioinformatics. Data clustering is the most common computational approach to unsupervised identification of cell populations from multidimensional cytometry data. However, interpretation of the identified data clusters is labor-intensive. Certain types of user-defined cell populations are also difficult to identify by fully automated data clustering analysis. Both are roadblocks before a cytometry lab can adopt the data clustering approach for cell population identification in routine use. We found that combining recursive data filtering and clustering with constraints converted from the user manual gating strategy can effectively address these two issues. We named this new approach DAFi: Directed Automated Filtering and Identification of cell populations. Design of DAFi preserves the data-driven characteristics of unsupervised clustering for identifying novel cell subsets, but also makes the results interpretable to experimental scientists through mapping and merging the multidimensional data clusters into the user-defined two-dimensional gating hierarchy. The recursive data filtering process in DAFi helped identify small data clusters which are otherwise difficult to resolve by a single run of the data clustering method due to the statistical interference of the irrelevant major clusters. Our experiment results showed that the proportions of the cell populations identified by DAFi, while being consistent with those by expert centralized manual gating, have smaller technical variances across samples than those from individual manual gating analysis and the nonrecursive data clustering analysis. Compared with manual gating segregation, DAFi-identified cell populations avoided the abrupt cut-offs on the boundaries. DAFi has been implemented to be used with multiple data clustering methods including K-means, FLOCK, FlowSOM, and

  9. Analysis of Cellular DNA Content by Flow Cytometry.

    Science.gov (United States)

    Darzynkiewicz, Zbigniew; Huang, Xuan; Zhao, Hong

    2017-11-01

    Cellular DNA content can be measured by flow cytometry with the aim of : (1) revealing cell distribution within the major phases of the cell cycle, (2) estimating frequency of apoptotic cells with fractional DNA content, and/or (3) disclosing DNA ploidy of the measured cell population. In this unit, simple and universally applicable methods for staining fixed cells are presented, as are methods that utilize detergents and/or proteolytic treatment to permeabilize cells and make DNA accessible to fluorochrome. Additionally, supravital cell staining with Hoechst 33342, which is primarily used for sorting live cells based on DNA-content differences for their subsequent culturing, is described. Also presented are methods for staining cell nuclei isolated from paraffin-embedded tissues. Available algorithms are listed for deconvolution of DNA-content-frequency histograms to estimate percentage of cells in major phases of the cell cycle and frequency of apoptotic cells with fractional DNA content. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley and Sons, Inc.

  10. Developing a NIR multispectral imaging for prediction and visualization of peanut protein content using variable selection algorithms

    Science.gov (United States)

    Cheng, Jun-Hu; Jin, Huali; Liu, Zhiwei

    2018-01-01

    The feasibility of developing a multispectral imaging method using important wavelengths from hyperspectral images selected by genetic algorithm (GA), successive projection algorithm (SPA) and regression coefficient (RC) methods for modeling and predicting protein content in peanut kernel was investigated for the first time. Partial least squares regression (PLSR) calibration model was established between the spectral data from the selected optimal wavelengths and the reference measured protein content ranged from 23.46% to 28.43%. The RC-PLSR model established using eight key wavelengths (1153, 1567, 1972, 2143, 2288, 2339, 2389 and 2446 nm) showed the best predictive results with the coefficient of determination of prediction (R2P) of 0.901, and root mean square error of prediction (RMSEP) of 0.108 and residual predictive deviation (RPD) of 2.32. Based on the obtained best model and image processing algorithms, the distribution maps of protein content were generated. The overall results of this study indicated that developing a rapid and online multispectral imaging system using the feature wavelengths and PLSR analysis is potential and feasible for determination of the protein content in peanut kernels.

  11. Identification and Characterization of Plasma Cells in Normal Human Bone Marrow by High-Resolution Flow Cytometry

    NARCIS (Netherlands)

    Terstappen, Leonardus Wendelinus Mathias Marie; Johnsen, Steen; Segers-Nolten, Gezina M.J.; Loken, Michael R.

    1990-01-01

    The low frequency of plasma cells and the lack of specific cell surface markers has been a major obstacle for a detailed characterization of plasma cells in normal human bone marrow. Multiparameter flow cytometry enabled the identification of plasma cells in normal bone marrow aspirates. The plasma

  12. Accuracy of Automated Flow Cytometry-Based Leukocyte Counts To Rule Out Urinary Tract Infection in Febrile Children: a Prospective Cross-Sectional Study

    Science.gov (United States)

    Duong, Hong Phuoc; Wissing, Karl Martin; Tram, Nathalie; Mascart, Georges; Lepage, Philippe

    2016-01-01

    Automated flow cytometry of urine remains an incompletely validated method to rule out urinary tract infection (UTI) in children. This cross-sectional analytical study was performed to compare the predictive values of flow cytometry and a dipstick test as initial diagnostic tests for UTI in febrile children and prospectively included 1,106 children (1,247 episodes). Urine culture was used as the gold standard test for diagnosing UTI. The performance of screening tests to diagnose UTI were established using receiver operating characteristic (ROC) analysis. Among these 1,247 febrile episodes, 221 UTIs were diagnosed (17.7% [95% confidence interval {CI}, 15.6 to 19.8%]). The area under the ROC curve for flow cytometry white blood cell (WBC) counts (0.99 [95% CI, 0.98 to 0.99]) was significantly superior to that for red blood cell (0.74 [95% CI, 0.70 to 0.78]) and bacterial counts (0.89 [95% CI, 0.87 to 0.92]) (P UTI in febrile children. PMID:27682127

  13. Selective interferometric imaging of internal multiples

    KAUST Repository

    Zuberi, M. A H; Alkhalifah, Tariq Ali

    2013-01-01

    are required to construct a Green’s function with all the scattered energy. As an alternative, we develop a three-step procedure, which images the first-order internal scattering using the background Green’s function (from the surface to each image point

  14. Mutant spectra of irradiated CHO AL cells determined with multiple markers analyzed by flow cytometry

    International Nuclear Information System (INIS)

    Ross, Carley D.; French, C. Tenley; Keysar, Stephen B.; Fox, Michael H.

    2007-01-01

    We have previously developed a sensitive and rapid mammalian cell mutation assay which is based on a Chinese hamster ovary cell line that stably incorporates human chromosome 11 (CHO A L ) and uses flow cytometry to measure mutations in CD59. We now show that multiparameter flow cytometry may be used to simultaneously analyze irradiated CHO A L cells for mutations in five CD genes along chromosome 11 (CD59, CD44, CD90, CD98, CD151) and also a GPI-anchor gene. Using this approach, 19 different mutant clones derived from individual sorted mutant cells were analyzed to determine the mutant spectrum induced by ionizing radiation. All clones analyzed were negative for CD59 expression and PCR confirmed that at least CD59 exon 4 was also absent. As expected, ionizing radiation frequently caused large deletions along chromosome 11. This technology can readily be used to rapidly analyze the mutant yield as well as the spectrum of mutations caused by a variety of genotoxic agents and provide greater insight into the mechanisms of mutagenesis

  15. Rapid Flow Cytometry-Based Test for the Diagnosis of Lipopolysaccharide Responsive Beige-Like Anchor (LRBA Deficiency

    Directory of Open Access Journals (Sweden)

    Laura Gámez-Díaz

    2018-04-01

    Full Text Available The diagnosis of lipopolysaccharide-responsive beige-like-anchor-protein (LRBA deficiency currently relies on gene sequencing approaches that do not support a timely diagnosis and clinical management. We developed a rapid and sensitive test for clinical implementation based on the detection of LRBA protein by flow cytometry in peripheral blood cells after stimulation. LRBA protein was assessed in a prospective cohort of 54 healthy donors and 57 patients suspected of LRBA deficiency. Receiver operating characteristics analysis suggested an LRBA:MFI ratio cutoff point of 2.6 to identify LRBA-deficient patients by FACS with 94% sensitivity and 80% specificity and to discriminate them from patients with a similar clinical picture but other disease-causing mutations. This easy flow cytometry-based assay allows a fast screening of patients with suspicion of LRBA deficiency reducing therefore the number of patients requiring LRBA sequencing and accelerating the treatment implementation. Detection of biallelic mutations in LRBA is however required for a definitive diagnosis.

  16. Rapid Flow Cytometry-Based Test for the Diagnosis of Lipopolysaccharide Responsive Beige-Like Anchor (LRBA) Deficiency.

    Science.gov (United States)

    Gámez-Díaz, Laura; Sigmund, Elena C; Reiser, Veronika; Vach, Werner; Jung, Sophie; Grimbacher, Bodo

    2018-01-01

    The diagnosis of lipopolysaccharide-responsive beige-like-anchor-protein (LRBA) deficiency currently relies on gene sequencing approaches that do not support a timely diagnosis and clinical management. We developed a rapid and sensitive test for clinical implementation based on the detection of LRBA protein by flow cytometry in peripheral blood cells after stimulation. LRBA protein was assessed in a prospective cohort of 54 healthy donors and 57 patients suspected of LRBA deficiency. Receiver operating characteristics analysis suggested an LRBA:MFI ratio cutoff point of 2.6 to identify LRBA-deficient patients by FACS with 94% sensitivity and 80% specificity and to discriminate them from patients with a similar clinical picture but other disease-causing mutations. This easy flow cytometry-based assay allows a fast screening of patients with suspicion of LRBA deficiency reducing therefore the number of patients requiring LRBA sequencing and accelerating the treatment implementation. Detection of biallelic mutations in LRBA is however required for a definitive diagnosis.

  17. Time gating for energy selection and scatter rejection: High-energy pulsed neutron imaging at LANSCE

    Science.gov (United States)

    Swift, Alicia; Schirato, Richard; McKigney, Edward; Hunter, James; Temple, Brian

    2015-09-01

    The Los Alamos Neutron Science Center (LANSCE) is a linear accelerator in Los Alamos, New Mexico that accelerates a proton beam to 800 MeV, which then produces spallation neutron beams. Flight path FP15R uses a tungsten target to generate neutrons of energy ranging from several hundred keV to ~600 MeV. The beam structure has micropulses of sub-ns width and period of 1.784 ns, and macropulses of 625 μs width and frequency of either 50 Hz or 100 Hz. This corresponds to 347 micropulses per macropulse, or 1.74 x 104 micropulses per second when operating at 50 Hz. Using a very fast, cooled ICCD camera (Princeton Instruments PI-Max 4), gated images of various objects were obtained on FP15R in January 2015. Objects imaged included blocks of lead and borated polyethylene; a tungsten sphere; and a tungsten, polyethylene, and steel cylinder. Images were obtained in 36 min or less, with some in as little as 6 min. This is novel because the gate widths (some as narrow as 10 ns) were selected to reject scatter and other signal not of interest (e.g. the gamma flash that precedes the neutron pulse), which has not been demonstrated at energies above 14 MeV. This proof-of-principle experiment shows that time gating is possible above 14MeV and is useful for selecting neutron energy and reducing scatter, thus forming clearer images. Future work (simulation and experimental) is being undertaken to improve camera shielding and system design and to precisely determine optical properties of the imaging system.

  18. Identity and Diversity of Human Peripheral Th and T Regulatory Cells Defined by Single-Cell Mass Cytometry.

    Science.gov (United States)

    Kunicki, Matthew A; Amaya Hernandez, Laura C; Davis, Kara L; Bacchetta, Rosa; Roncarolo, Maria-Grazia

    2018-01-01

    Human CD3 + CD4 + Th cells, FOXP3 + T regulatory (Treg) cells, and T regulatory type 1 (Tr1) cells are essential for ensuring peripheral immune response and tolerance, but the diversity of Th, Treg, and Tr1 cell subsets has not been fully characterized. Independent functional characterization of human Th1, Th2, Th17, T follicular helper (Tfh), Treg, and Tr1 cells has helped to define unique surface molecules, transcription factors, and signaling profiles for each subset. However, the adequacy of these markers to recapitulate the whole CD3 + CD4 + T cell compartment remains questionable. In this study, we examined CD3 + CD4 + T cell populations by single-cell mass cytometry. We characterize the CD3 + CD4 + Th, Treg, and Tr1 cell populations simultaneously across 23 memory T cell-associated surface and intracellular molecules. High-dimensional analysis identified several new subsets, in addition to the already defined CD3 + CD4 + Th, Treg, and Tr1 cell populations, for a total of 11 Th cell, 4 Treg, and 1 Tr1 cell subsets. Some of these subsets share markers previously thought to be selective for Treg, Th1, Th2, Th17, and Tfh cells, including CD194 (CCR4) + FOXP3 + Treg and CD183 (CXCR3) + T-bet + Th17 cell subsets. Unsupervised clustering displayed a phenotypic organization of CD3 + CD4 + T cells that confirmed their diversity but showed interrelation between the different subsets, including similarity between Th1-Th2-Tfh cell populations and Th17 cells, as well as similarity of Th2 cells with Treg cells. In conclusion, the use of single-cell mass cytometry provides a systems-level characterization of CD3 + CD4 + T cells in healthy human blood, which represents an important baseline reference to investigate abnormalities of different subsets in immune-mediated pathologies. Copyright © 2017 by The American Association of Immunologists, Inc.

  19. X-ray energy selected imaging with Medipix II

    International Nuclear Information System (INIS)

    Ludwig, J.; Zwerger, A.; Benz, K.-W.; Fiederle, M.; Braml, H.; Fauler, A.; Konrath, J.-P.

    2004-01-01

    Two different X-ray tube accelerating voltages (60 and 70 kV) are used for diagnosis of front teeth and molars. Different energy ranges are necessary as function of tooth thickness to obtain similar contrast for imaging. This technique drives the costs for the X-ray tube up and allows for just two optimized settings. Energy range selection for the detection of the penetrating X-rays would overcome these severe setbacks. The single photon counting chip MEDIPIX2 http://www.cern.ch/medipix exhibits exactly this feature. First simulations and measurements have been carried out using a dental X-ray source. As a demonstrator a real tooth has been used with different cavities and filling materials. Simulations showed in general larger improvements as compared to measurements regarding SNR and contrast: A beneficial factor of 4% wrt SNR and 25% for contrast, measurements showed factors of 2.5 and up to 10%, respectively

  20. X-ray energy selected imaging with Medipix II

    Science.gov (United States)

    Ludwig, J.; Zwerger, A.; Benz, K.-W.; Fiederle, M.; Braml, H.; Fauler, A.; Konrath, J.-P.

    2004-09-01

    Two different X-ray tube accelerating voltages (60 and 70kV) are used for diagnosis of front teeth and molars. Different energy ranges are necessary as function of tooth thickness to obtain similar contrast for imaging. This technique drives the costs for the X-ray tube up and allows for just two optimized settings. Energy range selection for the detection of the penetrating X-rays would overcome these severe setbacks. The single photon counting chip MEDIPIX2 http://www.cern.ch/medipix exhibits exactly this feature.First simulations and measurements have been carried out using a dental X-ray source. As a demonstrator a real tooth has been used with different cavities and filling materials. Simulations showed in general larger improvements as compared to measurements regarding SNR and contrast: A beneficial factor of 4% wrt SNR and 25% for contrast, measurements showed factors of 2.5 and up to 10%, respectively.

  1. CT angiography for planning transcatheter aortic valve replacement using automated tube voltage selection: Image quality and radiation exposure

    Energy Technology Data Exchange (ETDEWEB)

    Mangold, Stefanie [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Diagnostic and Interventional Radiology, Eberhard-Karls University Tuebingen, Tuebingen (Germany); De Cecco, Carlo N. [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Schoepf, U. Joseph, E-mail: schoepf@musc.edu [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Division of Cardiology, Department of Medicine, Medical University of South Carolina, Charleston, SC (United States); Kuhlman, Taylor S.; Varga-Szemes, Akos [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Caruso, Damiano [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Radiological Sciences, Oncology and Pathology, University of Rome “Sapienza”, Rome (Italy); Duguay, Taylor M. [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Tesche, Christian [Department of Cardiology, Heart Centre Munich-Bogenhausen, Munich (Germany); Vogl, Thomas J. [Division of Cardiovascular Imaging, Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Diagnostic and Interventional Radiology, University Hospital Frankfurt, Frankfurt (Germany); Nikolaou, Konstantin [Department of Diagnostic and Interventional Radiology, Eberhard-Karls University Tuebingen, Tuebingen (Germany); and others

    2017-01-15

    Highlights: • TAVR-planning CT was performed with automated tube voltage selection. • Automated tube voltage selection enables individual tube voltage adaptation. • Image quality was diagnostic while radiation exposure was significantly decreased. - Abstract: Purpose: To assess image quality and accuracy of CT angiography (CTA) for transcatheter aortic valve replacement (TAVR) planning performed with 3rd generation dual-source CT (DSCT). Material and methods: We evaluated 125 patients who underwent TAVR-planning CTA on 3rd generation DSCT. A two-part protocol was performed including retrospectively ECG-gated coronary CTA (CCTA) and prospectively ECG-triggered aortoiliac CTA using 60 mL of contrast medium. Automated tube voltage selection and advanced iterative reconstruction were applied. Effective dose (ED), signal-to-noise (SNR) and contrast-to-noise ratios (CNR) were calculated. Five-point scales were used for subjective image quality analysis. In patients who underwent TAVR, sizing parameters were obtained. Results: Image quality was rated good to excellent in 97.6% of CCTA and 100% of aortoiliac CTAs. CTA studies at >100 kV showed decreased objective image quality compared to 70–100 kV (SNR, all p ≤ 0.0459; CNR, all p ≤ 0.0462). Mean ED increased continuously from 70 to >100 kV (CCTA: 4.5 ± 1.7 mSv–13.6 ± 2.9 mSv, all p ≤ 0.0233; aortoiliac CTA: 2.4 ± 0.9 mSv–6.8 ± 2.7 mSv, all p ≤ 0.0414). In 39 patients TAVR was performed and annulus diameter was within the recommended range in all patients. No severe cardiac or vascular complications were noted. Conclusion: 3rd generation DSCT provides diagnostic image quality in TAVR-planning CTA and facilitates reliable assessment of TAVR device and delivery option while reducing radiation dose.

  2. CT angiography for planning transcatheter aortic valve replacement using automated tube voltage selection: Image quality and radiation exposure

    International Nuclear Information System (INIS)

    Mangold, Stefanie; De Cecco, Carlo N.; Schoepf, U. Joseph; Kuhlman, Taylor S.; Varga-Szemes, Akos; Caruso, Damiano; Duguay, Taylor M.; Tesche, Christian; Vogl, Thomas J.; Nikolaou, Konstantin

    2017-01-01

    Highlights: • TAVR-planning CT was performed with automated tube voltage selection. • Automated tube voltage selection enables individual tube voltage adaptation. • Image quality was diagnostic while radiation exposure was significantly decreased. - Abstract: Purpose: To assess image quality and accuracy of CT angiography (CTA) for transcatheter aortic valve replacement (TAVR) planning performed with 3rd generation dual-source CT (DSCT). Material and methods: We evaluated 125 patients who underwent TAVR-planning CTA on 3rd generation DSCT. A two-part protocol was performed including retrospectively ECG-gated coronary CTA (CCTA) and prospectively ECG-triggered aortoiliac CTA using 60 mL of contrast medium. Automated tube voltage selection and advanced iterative reconstruction were applied. Effective dose (ED), signal-to-noise (SNR) and contrast-to-noise ratios (CNR) were calculated. Five-point scales were used for subjective image quality analysis. In patients who underwent TAVR, sizing parameters were obtained. Results: Image quality was rated good to excellent in 97.6% of CCTA and 100% of aortoiliac CTAs. CTA studies at >100 kV showed decreased objective image quality compared to 70–100 kV (SNR, all p ≤ 0.0459; CNR, all p ≤ 0.0462). Mean ED increased continuously from 70 to >100 kV (CCTA: 4.5 ± 1.7 mSv–13.6 ± 2.9 mSv, all p ≤ 0.0233; aortoiliac CTA: 2.4 ± 0.9 mSv–6.8 ± 2.7 mSv, all p ≤ 0.0414). In 39 patients TAVR was performed and annulus diameter was within the recommended range in all patients. No severe cardiac or vascular complications were noted. Conclusion: 3rd generation DSCT provides diagnostic image quality in TAVR-planning CTA and facilitates reliable assessment of TAVR device and delivery option while reducing radiation dose.

  3. A perspective for biomedical data integration: Design of databases for flow cytometry

    Directory of Open Access Journals (Sweden)

    Lakoumentas John

    2008-02-01

    Full Text Available Abstract Background The integration of biomedical information is essential for tackling medical problems. We describe a data model in the domain of flow cytometry (FC allowing for massive management, analysis and integration with other laboratory and clinical information. The paper is concerned with the proper translation of the Flow Cytometry Standard (FCS into a relational database schema, in a way that facilitates end users at either doing research on FC or studying specific cases of patients undergone FC analysis Results The proposed database schema provides integration of data originating from diverse acquisition settings, organized in a way that allows syntactically simple queries that provide results significantly faster than the conventional implementations of the FCS standard. The proposed schema can potentially achieve up to 8 orders of magnitude reduction in query complexity and up to 2 orders of magnitude reduction in response time for data originating from flow cytometers that record 256 colours. This is mainly achieved by managing to maintain an almost constant number of data-mining procedures regardless of the size and complexity of the stored information. Conclusion It is evident that using single-file data storage standards for the design of databases without any structural transformations significantly limits the flexibility of databases. Analysis of the requirements of a specific domain for integration and massive data processing can provide the necessary schema modifications that will unlock the additional functionality of a relational database.

  4. Islet-selectivity of G-protein coupled receptor ligands evaluated for PET imaging of pancreatic {beta}-cell mass

    Energy Technology Data Exchange (ETDEWEB)

    Cline, Gary W., E-mail: gary.cline@yale.edu [Yale University School of Medicine (United States); Zhao, Xiaojian [Yale University School of Medicine (United States); Jakowski, Amy B.; Soeller, Walter C.; Treadway, Judith L. [Pfizer Global Research and Development, Pfizer Inc., Groton CT (United States)

    2011-09-02

    Highlights: {yields} We screened G-protein coupled receptors for imaging pancreatic. {yields} Database mining and immunohistochemistry identified GPCRs enriched in {beta}-cells. {yields} In vitro and in vivo assays were used to determine exocrine vs endocrine specificity. {yields} GPCR candidates for imaging of {beta}-cell mass are Prokineticin-1R, mGluR5, and GLP-1R. -- Abstract: A critical unmet need exists for methods to quantitatively measure endogenous pancreatic {beta}-cell mass (BCM) for the clinical evaluation of therapies to prevent or reverse loss of BCM and diabetes progression. Our objective was to identify G-protein coupled receptors (GPCRs) that are expressed with a high degree of specificity to islet {beta}-cells for receptor-targeted imaging of BCM. GPCRs enriched in pancreatic islets relative to pancreas acinar and hepatic tissue were identified using a database screen. Islet-specific expression was confirmed by human pancreas immunohistochemistry (IHC). In vitro selectivity assessment was determined from the binding and uptake of radiolabeled ligands to the rat insulinoma INS-1 832/13 cell line and isolated rat islets relative to the exocrine pancreas cell-type, PANC-1. Tail-vein injections of radioligands into rats were used to determine favorable image criteria of in vivo biodistribution to the pancreas relative to other internal organs (i.e., liver, spleen, stomach, and lungs). Database and IHC screening identified four candidate receptors for further in vitro and in vivo evaluation for PET imaging of BCM: prokineticin-1 receptor (PK-1R), metabotropic glutamate receptor type-5 (mGluR5), neuropeptide Y-2 receptor (NPY-2R), and glucagon-like peptide 1 receptor (GLP-1R). In vitro specificity ratios gave the following receptor rank order: PK-1R > GLP-1R > NPY-2R > mGluR5. The biodistribution rank order of selectivity to the pancreas was found to be PK-1R > VMAT2 {approx} GLP-1R > mGluR5. Favorable islet selectivity and biodistribution

  5. Islet-selectivity of G-protein coupled receptor ligands evaluated for PET imaging of pancreatic β-cell mass

    International Nuclear Information System (INIS)

    Cline, Gary W.; Zhao, Xiaojian; Jakowski, Amy B.; Soeller, Walter C.; Treadway, Judith L.

    2011-01-01

    Highlights: → We screened G-protein coupled receptors for imaging pancreatic. → Database mining and immunohistochemistry identified GPCRs enriched in β-cells. → In vitro and in vivo assays were used to determine exocrine vs endocrine specificity. → GPCR candidates for imaging of β-cell mass are Prokineticin-1R, mGluR5, and GLP-1R. -- Abstract: A critical unmet need exists for methods to quantitatively measure endogenous pancreatic β-cell mass (BCM) for the clinical evaluation of therapies to prevent or reverse loss of BCM and diabetes progression. Our objective was to identify G-protein coupled receptors (GPCRs) that are expressed with a high degree of specificity to islet β-cells for receptor-targeted imaging of BCM. GPCRs enriched in pancreatic islets relative to pancreas acinar and hepatic tissue were identified using a database screen. Islet-specific expression was confirmed by human pancreas immunohistochemistry (IHC). In vitro selectivity assessment was determined from the binding and uptake of radiolabeled ligands to the rat insulinoma INS-1 832/13 cell line and isolated rat islets relative to the exocrine pancreas cell-type, PANC-1. Tail-vein injections of radioligands into rats were used to determine favorable image criteria of in vivo biodistribution to the pancreas relative to other internal organs (i.e., liver, spleen, stomach, and lungs). Database and IHC screening identified four candidate receptors for further in vitro and in vivo evaluation for PET imaging of BCM: prokineticin-1 receptor (PK-1R), metabotropic glutamate receptor type-5 (mGluR5), neuropeptide Y-2 receptor (NPY-2R), and glucagon-like peptide 1 receptor (GLP-1R). In vitro specificity ratios gave the following receptor rank order: PK-1R > GLP-1R > NPY-2R > mGluR5. The biodistribution rank order of selectivity to the pancreas was found to be PK-1R > VMAT2 ∼ GLP-1R > mGluR5. Favorable islet selectivity and biodistribution characteristics suggest several GPCRs as potential

  6. Selection of optimal multispectral imaging system parameters for small joint arthritis detection

    Science.gov (United States)

    Dolenec, Rok; Laistler, Elmar; Stergar, Jost; Milanic, Matija

    2018-02-01

    Early detection and treatment of arthritis is essential for a successful outcome of the treatment, but it has proven to be very challenging with existing diagnostic methods. Novel methods based on the optical imaging of the affected joints are becoming an attractive alternative. A non-contact multispectral imaging (MSI) system for imaging of small joints of human hands and feet is being developed. In this work, a numerical simulation of the MSI system is presented. The purpose of the simulation is to determine the optimal design parameters. Inflamed and unaffected human joint models were constructed with a realistic geometry and tissue distributions, based on a MRI scan of a human finger with a spatial resolution of 0.2 mm. The light transport simulation is based on a weighted-photon 3D Monte Carlo method utilizing CUDA GPU acceleration. An uniform illumination of the finger within the 400-1100 nm spectral range was simulated and the photons exiting the joint were recorded using different acceptance angles. From the obtained reflectance and transmittance images the spectral and spatial features most indicative of inflammation were identified. Optimal acceptance angle and spectral bands were determined. This study demonstrates that proper selection of MSI system parameters critically affects ability of a MSI system to discriminate the unaffected and inflamed joints. The presented system design optimization approach could be applied to other pathologies.

  7. Flow cytometry for intracellular SPION quantification: specificity and sensitivity in comparison with spectroscopic methods

    Directory of Open Access Journals (Sweden)

    Friedrich RP

    2015-06-01

    Full Text Available Ralf P Friedrich,1 Christina Janko,1 Marina Poettler,1 Philipp Tripal,1 Jan Zaloga,1 Iwona Cicha,1 Stephan Dürr,1,2 Johannes Nowak,3 Stefan Odenbach,3 Ioana Slabu,4 Maik Liebl,4 Lutz Trahms,4 Marcus Stapf,5 Ingrid Hilger,5 Stefan Lyer,1 Christoph Alexiou1 1Department of Otorhinolaryngology, Head and Neck Surgery, Section of Experimental Oncology and Nanomedicine, University hospital Erlangen, 2Department of Otorhinolaryngology, Head and Neck Surgery, Section of Phoniatrics and Pediatric Audiology, University hospital Erlangen, Erlangen, 3Technische Universität Dresden, Chair of Magnetofluiddynamics, Measuring and Automation Technology, Dresden, 4Physikalisch-Technische Bundesanstalt Berlin, Berlin, 5Department of Radiology, Division of Diagnostic and Interventional Radiology, Experimental Radiology, University hospital Jena, Jena, Germany Abstract: Due to their special physicochemical properties, iron nanoparticles offer new promising possibilities for biomedical applications. For bench to bedside translation of superparamagnetic iron oxide nanoparticles (SPIONs, safety issues have to be comprehensively clarified. To understand concentration-dependent nanoparticle-mediated toxicity, the exact quantification of intracellular SPIONs by reliable methods is of great importance. In the present study, we compared three different SPION quantification methods (ultraviolet spectrophotometry, magnetic particle spectroscopy, atomic adsorption spectroscopy and discussed the shortcomings and advantages of each method. Moreover, we used those results to evaluate the possibility to use flow cytometric technique to determine the cellular SPION content. For this purpose, we correlated the side scatter data received from flow cytometry with the actual cellular SPION amount. We showed that flow cytometry provides a rapid and reliable method to assess the cellular SPION content. Our data also demonstrate that internalization of iron oxide nanoparticles in human

  8. An electronic image processing device featuring continuously selectable two-dimensional bipolar filter functions and real-time operation

    International Nuclear Information System (INIS)

    Charleston, B.D.; Beckman, F.H.; Franco, M.J.; Charleston, D.B.

    1981-01-01

    A versatile electronic-analogue image processing system has been developed for use in improving the quality of various types of images with emphasis on those encountered in experimental and diagnostic medicine. The operational principle utilizes spatial filtering which selectively controls the contrast of an image according to the spatial frequency content of relevant and non-relevant features of the image. Noise can be reduced or eliminated by selectively lowering the contrast of information in the high spatial frequency range. Edge sharpness can be enhanced by accentuating the upper midrange spatial frequencies. Both methods of spatial frequency control may be adjusted continuously in the same image to obtain maximum visibility of the features of interest. A precision video camera is used to view medical diagnostic images, either prints, transparencies or CRT displays. The output of the camera provides the analogue input signal for both the electronic processing system and the video display of the unprocessed image. The video signal input to the electronic processing system is processed by a two-dimensional spatial convolution operation. The system employs charged-coupled devices (CCDs), both tapped analogue delay lines (TADs) and serial analogue delay lines (SADs), to store information in the form of analogue potentials which are constantly being updated as new sampled analogue data arrive at the input. This information is convolved with a programmed bipolar radially symmetrical hexagonal function which may be controlled and varied at each radius by the operator in real-time by adjusting a set of front panel controls or by a programmed microprocessor control. Two TV monitors are used, one for processed image display and the other for constant reference to the original image. The working prototype has a full-screen display matrix size of 200 picture elements per horizontal line by 240 lines. The matrix can be expanded vertically and horizontally for the

  9. Attenuation-based automatic kilovolt (kV)-selection in computed tomography of the chest: Effects on radiation exposure and image quality

    Energy Technology Data Exchange (ETDEWEB)

    Eller, Achim; Wuest, Wolfgang; Scharf, Michael; Brand, Michael [Department of Radiology, University Erlangen (Germany); Achenbach, Stephan [Department of Cardiology, University Erlangen (Germany); Uder, Michael [Department of Radiology, University Erlangen (Germany); Imaging Science Institute, Erlangen (Germany); Lell, Michael M., E-mail: Michael.lell@uk-erlangen.de [Department of Radiology, University Erlangen (Germany); Imaging Science Institute, Erlangen (Germany)

    2013-12-01

    Objectives: To evaluate an automated attenuation-based kV-selection in computed tomography of the chest in respect to radiation dose and image quality, compared to a standard 120 kV protocol. Materials and methods: 104 patients were examined using a 128-slice scanner. Fifty examinations (58 ± 15 years, study group) were performed using the automated adaption of tube potential (100–140 kV), based on the attenuation profile of the scout scan, 54 examinations (62 ± 14 years, control group) with fixed 120 kV. Estimated CT dose index (CTDI) of the software-proposed setting was compared with a 120 kV protocol. After the scan CTDI volume (CTDIvol) and dose length product (DLP) were recorded. Image quality was assessed by region of interest (ROI) measurements, subjective image quality by two observers with a 4-point scale (3 – excellent, 0 – not diagnostic). Results: The algorithm selected 100 kV in 78% and 120 kV in 22%. Overall CTDIvol reduction was 26.6% (34% in 100 kV) overall DLP reduction was 22.8% (32.1% in 100 kV) (all p < 0.001). Subjective image quality was excellent in both groups. Conclusion: The attenuation based kV-selection algorithm enables relevant dose reduction (∼27%) in chest-CT while keeping image quality parameters at high levels.

  10. Attenuation-based automatic kilovolt (kV)-selection in computed tomography of the chest: Effects on radiation exposure and image quality

    International Nuclear Information System (INIS)

    Eller, Achim; Wuest, Wolfgang; Scharf, Michael; Brand, Michael; Achenbach, Stephan; Uder, Michael; Lell, Michael M.

    2013-01-01

    Objectives: To evaluate an automated attenuation-based kV-selection in computed tomography of the chest in respect to radiation dose and image quality, compared to a standard 120 kV protocol. Materials and methods: 104 patients were examined using a 128-slice scanner. Fifty examinations (58 ± 15 years, study group) were performed using the automated adaption of tube potential (100–140 kV), based on the attenuation profile of the scout scan, 54 examinations (62 ± 14 years, control group) with fixed 120 kV. Estimated CT dose index (CTDI) of the software-proposed setting was compared with a 120 kV protocol. After the scan CTDI volume (CTDIvol) and dose length product (DLP) were recorded. Image quality was assessed by region of interest (ROI) measurements, subjective image quality by two observers with a 4-point scale (3 – excellent, 0 – not diagnostic). Results: The algorithm selected 100 kV in 78% and 120 kV in 22%. Overall CTDIvol reduction was 26.6% (34% in 100 kV) overall DLP reduction was 22.8% (32.1% in 100 kV) (all p < 0.001). Subjective image quality was excellent in both groups. Conclusion: The attenuation based kV-selection algorithm enables relevant dose reduction (∼27%) in chest-CT while keeping image quality parameters at high levels

  11. Influence of gating phase selection on the image quality of coronary arteries in multidetector row computed tomography

    International Nuclear Information System (INIS)

    Laskowska, K.; Marzec, M.; Serafin, Z.; Nawrocka, E.; Lasek, W.; WWisniewska-Szmyt, J.; Kubica, J.

    2005-01-01

    Motion artifacts caused by cardiac movement disturb the imaging of coronary arteries with multidetector-row spiral computed tomography. The aim of this study was to determine the phase of the heart rate which provides the best quality of coronary artery imaging in retrospective ECG-gated CT. Although 75% is usually the best reconstruction phase, the optimal phase should be established individually for the patient, artery, segment, and type of tomograph for the best imaging quality. Forty-five cardiac CT angiograms of 26 patients were retrospectively evaluated. The examinations were performed with a 4-detector-row tomograph. ECG-gated retrospective reconstructions were relatively delayed at 0%, 12.5%, 25%, 37.5%, 50%, 62.5%, 75%, and 87.5% of the cardiac cycle. Selected coronary arteries of the highest diagnostic quality were estimated in the eight phases of the cardiac cycle. Only arteries of very high image quality were selected for analysis: left coronary artery trunks (44 cases, incl. 37 stented), anterior interventricular branches (36, incl. 3 stented), circumflex branches (16), right coronary rtery branches (23), and posterior interventricular branches (4). The reconstruction phase had a statistically significant impact on the quality of imaging (p < 0.0003). Depending on the case, optimal imaging was noted in various phases, except in the 12.5 % phase. The 75% phase appeared to be the best of all those examined (p < 0.05), both in the group of arteries without stents (p < 0.0006) and in those stented (p < 0.05). In some cases of repeated examinations the best phases differed within the same patient. (author)

  12. Quantification of colour Doppler activity in the wrist in patients with rheumatoid arthritis - the reliability of different methods for image selection and evaluation

    DEFF Research Database (Denmark)

    Ellegaard, K.; Torp-Pedersen, S.; Lund, H.

    2008-01-01

    measurements in the wrist of patients with rheumatoid arthritis (RA) using different selection and quantification methods. Materials and Methods: 14 patients with RA had their wrist scanned twice by the same investigator with an interval of 30 Minutes, The images for analysis were selected either......Purpose: The amount Of colour Doppler activity in the inflamed synovium is used to quantity inflammatory activity. The measurements may vary due to image selection, quantification method, and point in cardiac cycle. This study investigated the test-retest reliability Of ultrasound colour Doppler...... was obtained when the images were selected guided by colour Doppler and the Subsequent quantification was (done in an area defined by anatomical Structures. With this method, the intra-class coefficient ICC (2.1) was 0.95 and the within-subject SD (SW) was 0.017, indicating good reliability. In contrast, poor...

  13. Combined Confocal and Wide-Field High-Resolution Cytometry of Fluorescent In Situ Hybridization-Stained Cells

    Czech Academy of Sciences Publication Activity Database

    Kozubek, Michal; Kozubek, Stanislav; Lukášová, Emilie; Bártová, Eva; Skalníková, M.; Matula, Pa.; Matula, Pe.; Jirsová, Pavla; Cafourková, Alena; Koutná, Irena

    2001-01-01

    Roč. 45, č. 1 (2001), s. 1-12 ISSN 0196-4763 R&D Projects: GA MŠk VS97031; GA ČR GA202/99/P008; GA AV ČR IBS5004010 Institutional research plan: CEZ:AV0Z5004920 Keywords : high-resolution cytometry * fluorescence in situ hybridization * interphase nuclei Subject RIV: BO - Biophysics Impact factor: 2.220, year: 2001

  14. Zinc fixation preserves flow cytometry scatter and fluorescence parameters and allows simultaneous analysis of DNA content and synthesis, and intracellular and surface epitopes

    DEFF Research Database (Denmark)

    Jensen, Uffe Birk; Owens, David; Pedersen, Søren

    2010-01-01

    Zinc salt-based fixation (ZBF) has proved advantageous in histochemical analyses conducted on intact tissues but has not been exploited in flow cytometry procedures that focus on quantitative analysis of individual cells. Here, we show that ZBF performs equally well to paraformaldehyde in the pre......Zinc salt-based fixation (ZBF) has proved advantageous in histochemical analyses conducted on intact tissues but has not been exploited in flow cytometry procedures that focus on quantitative analysis of individual cells. Here, we show that ZBF performs equally well to paraformaldehyde...... allowing subsequent quantitative PCR analysis or labeling for incorporation of the thymidine analog EdU following surface and intracellular epitope staining. Finally, ZBF treatment allows for long-term storage of labeled cells with little change in these parameters. Thus, we present a protocol for zinc...... salt fixation of cells that allows for the simultaneous analysis of DNA and intracellular and cell surface proteins by flow cytometry....

  15. Global phenotypic characterisation of human platelet lysate expanded MSCs by high-throughput flow cytometry.

    Science.gov (United States)

    Reis, Monica; McDonald, David; Nicholson, Lindsay; Godthardt, Kathrin; Knobel, Sebastian; Dickinson, Anne M; Filby, Andrew; Wang, Xiao-Nong

    2018-03-02

    Mesenchymal stromal cells (MSCs) are a promising cell source to develop cell therapy for many diseases. Human platelet lysate (PLT) is increasingly used as an alternative to foetal calf serum (FCS) for clinical-scale MSC production. To date, the global surface protein expression of PLT-expended MSCs (MSC-PLT) is not known. To investigate this, paired MSC-PLT and MSC-FCS were analysed in parallel using high-throughput flow cytometry for the expression of 356 cell surface proteins. MSC-PLT showed differential surface protein expression compared to their MSC-FCS counterpart. Higher percentage of positive cells was observed in MSC-PLT for 48 surface proteins, of which 13 were significantly enriched on MSC-PLT. This finding was validated using multiparameter flow cytometry and further confirmed by quantitative staining intensity analysis. The enriched surface proteins are relevant to increased proliferation and migration capacity, as well as enhanced chondrogenic and osteogenic differentiation properties. In silico network analysis revealed that these enriched surface proteins are involved in three distinct networks that are associated with inflammatory responses, carbohydrate metabolism and cellular motility. This is the first study reporting differential cell surface protein expression between MSC-PLT and MSC-FSC. Further studies are required to uncover the impact of those enriched proteins on biological functions of MSC-PLT.

  16. Metal artifact correction for x-ray computed tomography using kV and selective MV imaging

    International Nuclear Information System (INIS)

    Wu, Meng; Keil, Andreas; Constantin, Dragos; Star-Lack, Josh; Zhu, Lei; Fahrig, Rebecca

    2014-01-01

    Purpose: The overall goal of this work is to improve the computed tomography (CT) image quality for patients with metal implants or fillings by completing the missing kilovoltage (kV) projection data with selectively acquired megavoltage (MV) data that do not suffer from photon starvation. When both of these imaging systems, which are available on current radiotherapy devices, are used, metal streak artifacts are avoided, and the soft-tissue contrast is restored, even for regions in which the kV data cannot contribute any information. Methods: Three image-reconstruction methods, including two filtered back-projection (FBP)-based analytic methods and one iterative method, for combining kV and MV projection data from the two on-board imaging systems of a radiotherapy device are presented in this work. The analytic reconstruction methods modify the MV data based on the information in the projection or image domains and then patch the data onto the kV projections for a FBP reconstruction. In the iterative reconstruction, the authors used dual-energy (DE) penalized weighted least-squares (PWLS) methods to simultaneously combine the kV/MV data and perform the reconstruction. Results: The authors compared kV/MV reconstructions to kV-only reconstructions using a dental phantom with fillings and a hip-implant numerical phantom. Simulation results indicated that dual-energy sinogram patch FBP and the modified dual-energy PWLS method can successfully suppress metal streak artifacts and restore information lost due to photon starvation in the kV projections. The root-mean-square errors of soft-tissue patterns obtained using combined kV/MV data are 10–15 Hounsfield units smaller than those of the kV-only images, and the structural similarity index measure also indicates a 5%–10% improvement in the image quality. The added dose from the MV scan is much less than the dose from the kV scan if a high efficiency MV detector is assumed. Conclusions: The authors have shown that it

  17. Metal artifact correction for x-ray computed tomography using kV and selective MV imaging

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Meng, E-mail: mengwu@stanford.edu [Department of Electrical Engineering, Stanford University, Stanford, California 94305 (United States); Keil, Andreas [microDimensions GmbH, Munich 81379 (Germany); Constantin, Dragos; Star-Lack, Josh [Varian Medical Systems, Inc., Palo Alto, California 94304 (United States); Zhu, Lei [Nuclear and Radiological Engineering and Medical Physics Programs, The George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332 (United States); Fahrig, Rebecca [Department of Radiology, Stanford University, Stanford, California 94305 (United States)

    2014-12-15

    Purpose: The overall goal of this work is to improve the computed tomography (CT) image quality for patients with metal implants or fillings by completing the missing kilovoltage (kV) projection data with selectively acquired megavoltage (MV) data that do not suffer from photon starvation. When both of these imaging systems, which are available on current radiotherapy devices, are used, metal streak artifacts are avoided, and the soft-tissue contrast is restored, even for regions in which the kV data cannot contribute any information. Methods: Three image-reconstruction methods, including two filtered back-projection (FBP)-based analytic methods and one iterative method, for combining kV and MV projection data from the two on-board imaging systems of a radiotherapy device are presented in this work. The analytic reconstruction methods modify the MV data based on the information in the projection or image domains and then patch the data onto the kV projections for a FBP reconstruction. In the iterative reconstruction, the authors used dual-energy (DE) penalized weighted least-squares (PWLS) methods to simultaneously combine the kV/MV data and perform the reconstruction. Results: The authors compared kV/MV reconstructions to kV-only reconstructions using a dental phantom with fillings and a hip-implant numerical phantom. Simulation results indicated that dual-energy sinogram patch FBP and the modified dual-energy PWLS method can successfully suppress metal streak artifacts and restore information lost due to photon starvation in the kV projections. The root-mean-square errors of soft-tissue patterns obtained using combined kV/MV data are 10–15 Hounsfield units smaller than those of the kV-only images, and the structural similarity index measure also indicates a 5%–10% improvement in the image quality. The added dose from the MV scan is much less than the dose from the kV scan if a high efficiency MV detector is assumed. Conclusions: The authors have shown that it

  18. Metal artifact correction for x-ray computed tomography using kV and selective MV imaging.

    Science.gov (United States)

    Wu, Meng; Keil, Andreas; Constantin, Dragos; Star-Lack, Josh; Zhu, Lei; Fahrig, Rebecca

    2014-12-01

    The overall goal of this work is to improve the computed tomography (CT) image quality for patients with metal implants or fillings by completing the missing kilovoltage (kV) projection data with selectively acquired megavoltage (MV) data that do not suffer from photon starvation. When both of these imaging systems, which are available on current radiotherapy devices, are used, metal streak artifacts are avoided, and the soft-tissue contrast is restored, even for regions in which the kV data cannot contribute any information. Three image-reconstruction methods, including two filtered back-projection (FBP)-based analytic methods and one iterative method, for combining kV and MV projection data from the two on-board imaging systems of a radiotherapy device are presented in this work. The analytic reconstruction methods modify the MV data based on the information in the projection or image domains and then patch the data onto the kV projections for a FBP reconstruction. In the iterative reconstruction, the authors used dual-energy (DE) penalized weighted least-squares (PWLS) methods to simultaneously combine the kV/MV data and perform the reconstruction. The authors compared kV/MV reconstructions to kV-only reconstructions using a dental phantom with fillings and a hip-implant numerical phantom. Simulation results indicated that dual-energy sinogram patch FBP and the modified dual-energy PWLS method can successfully suppress metal streak artifacts and restore information lost due to photon starvation in the kV projections. The root-mean-square errors of soft-tissue patterns obtained using combined kV/MV data are 10-15 Hounsfield units smaller than those of the kV-only images, and the structural similarity index measure also indicates a 5%-10% improvement in the image quality. The added dose from the MV scan is much less than the dose from the kV scan if a high efficiency MV detector is assumed. The authors have shown that it is possible to improve the image quality of

  19. Assessment of β-carotene content, cell physiology and morphology of the yellow yeast Rhodotorula glutinis mutant 400A15 using flow cytometry.

    Science.gov (United States)

    Cutzu, Raffaela; Clemente, Ana; Reis, Alberto; Nobre, Beatriz; Mannazzu, Ilaria; Roseiro, José; Lopes da Silva, Teresa

    2013-08-01

    Flow cytometry was used to assess β-carotene content, cell membrane permeability, cell size and granularity in Rhodotorula glutinis mutant 400A15 grown under different oxygen transfer coefficients (k L a) and carbon to nitrogen ratios (C/N). A Doehlert distribution was used in order to select the best conditions that induced the highest carotenoids production. The highest β-carotene content (0.79 mg g(-1) DCW) at the lowest k L a and C/N (5 × 10(-3) s(-1) and 11.3 respectively). Under these conditions, the biomass concentration attained 18.60 g L(-1). The highest ratio of cells with permeabilised membranes (2.6 %), and the highest cell size and granularity were also obtained under these conditions. It was observed that C/N showed a stronger influence than the k L a on the measured cell parameters.

  20. Monitoring Immune Responses in Organ Recipients by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Al-Mukhalafi Zuha

    2001-01-01

    Full Text Available Allograft rejection remains a major barrier to successful organ transplan-tation. Cellular and humoral immune responses play a critical role in mediating graft rejection. During the last few years, monoclonal antibodies have been used as a new specific therapeutic approach in the prevention of allograft rejection. Recently, the technology of flow cytometry has become a useful tool for monitoring immunological responses in transplant recipients. The application of this valuable tool in clinical transplantation at the present time is aimed at, i determining the extent of immuno-suppressive therapy through T-cell receptor analysis of cellular components, ii monitoring levels of alloreactive antibodies to identify high-risk recipients (sensitized patients in the pre-operative period and iii to predict rejection by monitoring their development post-operatively. In future, further development of this technology may demonstrate greater benefit to the field of organ transplantation.

  1. Correlation between flow cytometry and histologic findings: ten year experience in the investigation of lymphoproliferative diseases

    Directory of Open Access Journals (Sweden)

    Alanna Mara Pinheiro Sobreira Bezerra

    2011-06-01

    Full Text Available Objective: To demonstrate the advantages of correlatingflow cytometry immunophenotyping with the pathology/immunohistochemistry of lymph nodes or nodules in the diagnosisof lymphoproliferative diseases. Methods: A retrospective studywas carried out of 157 biopsy or fine-needle aspiration lymph nodes/nodule specimens taken from 142 patients, from 1999 and 2009.The specimens were simultaneously studied with flow cytometryand pathology at Hospital Israelita Albert Einstein. The specimenswere prepared in hematoxylin/eosin, Giemsa, or monoclonal antibodystained slides for detecting specific antibodies for the purposesof pathology/immunohistochemical analysis. The samples werehemolyzed and marked with different monoclonal antibody panels fordifferent antigens in flow cytometry immunophenotyping. Results:The diagnostic results of pathology/immunohistochemical studiesand flow cytometry immunophenotyping agreed in 115 patients(81%, corresponding to 127 specimens, as follows according tothe pathologic diagnosis: 63 patients with non-Hodgkin’s B-celllymphoma; 26 patients with reactive lymphoid hyperplasia; 5 patientswith non-Hodgkin’s T-cell lymphoma; 4 patients with atypical lymphoidproliferation; 5 patients with a chronic granulomatous inflammatoryprocess; 5 patients with a non-hematologic diagnosis; 2 patientswith granulocytic sarcoma; 2 patients with thymoma; 1 patientwith byphenotypic leukemia; 1 patient with kappa plasmocytoma;1 patient with Hodgkin’s lymphoma. Subtypes of lymphomas couldbe classified by associating the two techniques: 19 patients withfollicular lymphoma; 15 patients with diffuse large B-cell lymphoma; 7patients with small lymphocytic B-cell lymphoma/chronic lymphocyticleukemia; 3 patients with mantle cell lymphoma; 1 patient withBurkitt’s lymphoma; 1 patient with MALT type lymphoma; 1 patientwith post-transplant lymphoproliferative disease; 2 patients with highgrade non-Hodgkin’s B-cell lymphoma; 1 patient with low grade

  2. New detection systems of bacteria using highly selective media designed by SMART: selective medium-design algorithm restricted by two constraints.

    Directory of Open Access Journals (Sweden)

    Takeshi Kawanishi

    Full Text Available Culturing is an indispensable technique in microbiological research, and culturing with selective media has played a crucial role in the detection of pathogenic microorganisms and the isolation of commercially useful microorganisms from environmental samples. Although numerous selective media have been developed in empirical studies, unintended microorganisms often grow on such media probably due to the enormous numbers of microorganisms in the environment. Here, we present a novel strategy for designing highly selective media based on two selective agents, a carbon source and antimicrobials. We named our strategy SMART for highly Selective Medium-design Algorithm Restricted by Two constraints. To test whether the SMART method is applicable to a wide range of microorganisms, we developed selective media for Burkholderia glumae, Acidovorax avenae, Pectobacterium carotovorum, Ralstonia solanacearum, and Xanthomonas campestris. The series of media developed by SMART specifically allowed growth of the targeted bacteria. Because these selective media exhibited high specificity for growth of the target bacteria compared to established selective media, we applied three notable detection technologies: paper-based, flow cytometry-based, and color change-based detection systems for target bacteria species. SMART facilitates not only the development of novel techniques for detecting specific bacteria, but also our understanding of the ecology and epidemiology of the targeted bacteria.

  3. High throughput imaging cytometer with acoustic focussing.

    Science.gov (United States)

    Zmijan, Robert; Jonnalagadda, Umesh S; Carugo, Dario; Kochi, Yu; Lemm, Elizabeth; Packham, Graham; Hill, Martyn; Glynne-Jones, Peter

    2015-10-31

    We demonstrate an imaging flow cytometer that uses acoustic levitation to assemble cells and other particles into a sheet structure. This technique enables a high resolution, low noise CMOS camera to capture images of thousands of cells with each frame. While ultrasonic focussing has previously been demonstrated for 1D cytometry systems, extending the technology to a planar, much higher throughput format and integrating imaging is non-trivial, and represents a significant jump forward in capability, leading to diagnostic possibilities not achievable with current systems. A galvo mirror is used to track the images of the moving cells permitting exposure times of 10 ms at frame rates of 50 fps with motion blur of only a few pixels. At 80 fps, we demonstrate a throughput of 208 000 beads per second. We investigate the factors affecting motion blur and throughput, and demonstrate the system with fluorescent beads, leukaemia cells and a chondrocyte cell line. Cells require more time to reach the acoustic focus than beads, resulting in lower throughputs; however a longer device would remove this constraint.

  4. Automatic spectral imaging protocol selection and iterative reconstruction in abdominal CT with reduced contrast agent dose: initial experience

    Energy Technology Data Exchange (ETDEWEB)

    Lv, Peijie; Liu, Jie; Chai, Yaru; Yan, Xiaopeng; Gao, Jianbo; Dong, Junqiang [The First Affiliated Hospital of Zhengzhou University, Department of Radiology, Zhengzhou, Henan Province (China)

    2017-01-15

    To evaluate the feasibility, image quality, and radiation dose of automatic spectral imaging protocol selection (ASIS) and adaptive statistical iterative reconstruction (ASIR) with reduced contrast agent dose in abdominal multiphase CT. One hundred and sixty patients were randomly divided into two scan protocols (n = 80) each; protocol A, 120 kVp/450 mgI/kg, filtered back projection algorithm (FBP); protocol B, spectral CT imaging with ASIS and 40 to 70 keV monochromatic images generated per 300 mgI/kg, ASIR algorithm. Quantitative parameters (image noise and contrast-to-noise ratios [CNRs]) and qualitative visual parameters (image noise, small structures, organ enhancement, and overall image quality) were compared. Monochromatic images at 50 keV and 60 keV provided similar or lower image noise, but higher contrast and overall image quality as compared with 120-kVp images. Despite the higher image noise, 40-keV images showed similar overall image quality compared to 120-kVp images. Radiation dose did not differ between the two protocols, while contrast agent dose in protocol B was reduced by 33 %. Application of ASIR and ASIS to monochromatic imaging from 40 to 60 keV allowed contrast agent dose reduction with adequate image quality and without increasing radiation dose compared to 120 kVp with FBP. (orig.)

  5. Automatic spectral imaging protocol selection and iterative reconstruction in abdominal CT with reduced contrast agent dose: initial experience

    International Nuclear Information System (INIS)

    Lv, Peijie; Liu, Jie; Chai, Yaru; Yan, Xiaopeng; Gao, Jianbo; Dong, Junqiang

    2017-01-01

    To evaluate the feasibility, image quality, and radiation dose of automatic spectral imaging protocol selection (ASIS) and adaptive statistical iterative reconstruction (ASIR) with reduced contrast agent dose in abdominal multiphase CT. One hundred and sixty patients were randomly divided into two scan protocols (n = 80) each; protocol A, 120 kVp/450 mgI/kg, filtered back projection algorithm (FBP); protocol B, spectral CT imaging with ASIS and 40 to 70 keV monochromatic images generated per 300 mgI/kg, ASIR algorithm. Quantitative parameters (image noise and contrast-to-noise ratios [CNRs]) and qualitative visual parameters (image noise, small structures, organ enhancement, and overall image quality) were compared. Monochromatic images at 50 keV and 60 keV provided similar or lower image noise, but higher contrast and overall image quality as compared with 120-kVp images. Despite the higher image noise, 40-keV images showed similar overall image quality compared to 120-kVp images. Radiation dose did not differ between the two protocols, while contrast agent dose in protocol B was reduced by 33 %. Application of ASIR and ASIS to monochromatic imaging from 40 to 60 keV allowed contrast agent dose reduction with adequate image quality and without increasing radiation dose compared to 120 kVp with FBP. (orig.)

  6. In vivo Raman flow cytometry for real-time detection of carbon nanotube kinetics in lymph, blood, and tissues

    Science.gov (United States)

    Biris, Alexandru S.; Galanzha, Ekaterina I.; Li, Zhongrui; Mahmood, Meena; Xu, Yang; Zharov, Vladimir P.

    2009-03-01

    Nanoparticles are intensively being explored as contrast agents for medical diagnostics and therapies using various optical methods. We present the first demonstration of the use of time-resolved Raman spectroscopy for in vivo real-time detection of circulating carbon nanotubes (CNTs) or cancer cells labeled with CNTs in the lymph, blood, and tissues of live animals with fast spectral acquisition times of down to few milliseconds. After intravenously administering CNTs in the tail vein of the rat, this technique provides the ability to detect the circulation of CNTs in the blood microvessels of the intact rat ear. The capability of Raman spectroscopy is also demonstrated to monitor, identify, and image the CNTs during their transportation by lymphatics in the rat ear and mesentery. The strong and specific Raman scattering properties of CNTs make it possible to detect in vitro and in vivo single cancer cells (HeLa) tagged with CNTs. In vivo Raman flow cytometry opens a new avenue for multiparameter analysis of circulating nanoparticles with strong Raman scattering properties and their pharmokinetics in blood and lymph systems. Moreover, this technology has the potential for molecular detection and identification of circulating tumor cells, and infections labeled with CNTs.

  7. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry

    Directory of Open Access Journals (Sweden)

    L.P.S. Alves

    Full Text Available The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i cell permeabilization, ii Nile red staining, and iii analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99 compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots.

  8. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry.

    Science.gov (United States)

    Alves, L P S; Almeida, A T; Cruz, L M; Pedrosa, F O; de Souza, E M; Chubatsu, L S; Müller-Santos, M; Valdameri, G

    2017-01-16

    The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots.

  9. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry

    Science.gov (United States)

    Alves, L.P.S.; Almeida, A.T.; Cruz, L.M.; Pedrosa, F.O.; de Souza, E.M.; Chubatsu, L.S.; Müller-Santos, M.; Valdameri, G.

    2017-01-01

    The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots. PMID:28099582

  10. Recombinant carcinoembryonic antigen as a reporter gene for molecular imaging

    International Nuclear Information System (INIS)

    Kenanova, Vania; Barat, Bhaswati; Olafsen, Tove; Chatziioannou, Arion; Herschman, Harvey R.; Wu, Anna M.; Braun, Jonathan

    2009-01-01

    Reporter genes can provide a way of noninvasively assessing gene activity in vivo. However, current reporter gene strategies may be limited by the immunogenicity of foreign reporter proteins, endogenous expression, or unwanted biological activity. We have developed a reporter gene based on carcinoembryonic antigen (CEA), a human protein with limited normal tissue expression. To construct a CEA reporter gene for PET, a CEA minigene (N-A3) was fused to the extracellular and transmembrane domains of the human FcγRIIb receptor. The NA3-FcγRIIb recombinant gene, driven by a CMV promoter, was transfected in Jurkat (human T cell leukemia) cells. Expression was analyzed by flow cytometry, immunohistochemistry (IHC), and microPET imaging. Flow cytometry identified Jurkat clones stably expressing NA3-FcγRIIb at low, medium, and high levels. High and medium NA3-FcγRIIb expression could also be detected by Western blot. Reporter gene positive and negative Jurkat cells were used to establish xenografts in athymic mice. IHC showed staining of the tumor with high reporter gene expression; medium and low N-A3 expression was not detected. MicroPET imaging, using an anti-CEA 124 I-labeled single-chain Fv-Fc antibody fragment, demonstrated that only high N-A3 expression could be detected. Specific accumulation of activity was visualized at the N-A3 positive tumor as early as 4 h. MicroPET image quantitation showed tumor activity of 1.8 ± 0.2, 15.2 ± 1.3, and 4.6 ± 1.2 percent injected dose per gram (%ID/g) at 4, 20, and 48 h, respectively. Biodistribution at 48 h demonstrated tumor uptake of 4.8 ± 0.8%ID/g. The CEA N-A3 minigene has the potential to be used as a reporter gene for imaging cells in vivo. (orig.)

  11. Flow cytometry: design, development and experimental validation

    International Nuclear Information System (INIS)

    Seigneur, Alain

    1987-01-01

    The flow cytometry techniques allow the analysis and sorting of living biologic cells at rates above five to ten thousand events per second. After a short review, we present in this report the design and development of a 'high-tech' apparatus intended for research laboratories and the experimental results. The first part deals with the physical principles allowing morphologic and functional analysis of cells or cellular components. The measured parameters are as follows: electrical resistance pulse sizing, light scattering and fluorescence. Hydrodynamic centering is used, and in the same way, the division of a water-stream into droplets leading to electrostatic sorting of particles. The second part deals with the apparatus designed by the 'Commissariat a l'Energie Atomique' (C.E.A.) and industrialised by 'ODAM' (ATC 3000). The last part of this thesis work is the performance evaluations of this cyto-meter. The difference between the two size measurement methods are analyzed: electrical resistance pulse sizing versus small-angle light scattering. By an original optics design, high sensitivity has been reached in the fluorescence measurement: the equivalent noise corresponds to six hundred fluorescein isothiocyanate (FITC) molecules. The sorting performances have also been analyzed and the cell viability proven. (author) [fr

  12. Flow cytometry, microsatellites and niche models reveal the origins and geographical structure of Alnus glutinosa populations in Europe

    Czech Academy of Sciences Publication Activity Database

    Mandák, Bohumil; Vít, Petr; Krak, Karol; Trávníček, Pavel; Havrdová, Alena; Hadincová, Věroslava; Zákravský, Petr; Jarolímová, Vlasta; Bacles, C. F. E.; Douda, Jan

    2016-01-01

    Roč. 117, č. 1 (2016), s. 107-120 ISSN 0305-7364 R&D Projects: GA ČR(CZ) GAP504/11/0402 Institutional support: RVO:67985939 Keywords : flow cytometry * microsatellites * glacial refugia Subject RIV: EF - Botanics Impact factor: 4.041, year: 2016

  13. Size selectivity of magnetite core- (Ag/Au) shell nanoparticles for multimodal imaging applications

    Science.gov (United States)

    Singh, Pinki; Upadhyay, Chandan

    2017-10-01

    The magnetic and optical properties of nanomaterials play a significant role in the selection of the materials to be used for contrast enhancement in different biological and cell imaging techniques. These nanoparticles can also be used as drug delivery agents. The calculation of absorption efficiency and surface plasmon resonance wavelength has been performed using Mie theory and MATLAB programs. The study of spectrum calculated indicates the dependence of several optical properties such as resonance and absorption efficiency peak on ratio of core radius to the thickness of shell. A systematic study on the effect of nanoparticle dimension has been presented which clearly indicates that the optical properties can be modulated across the visible and near infrared range by changing these parameters. These properties of nanomaterials make them suitable for their application as multimodal imaging agents as they are also magnetically active and biocompatible.

  14. B-ALL minimal residual disease flow cytometry: an application of a novel method for optimization of a single-tube model.

    Science.gov (United States)

    Shaver, Aaron C; Greig, Bruce W; Mosse, Claudio A; Seegmiller, Adam C

    2015-05-01

    Optimizing a clinical flow cytometry panel can be a subjective process dependent on experience. We develop a quantitative method to make this process more rigorous and apply it to B lymphoblastic leukemia/lymphoma (B-ALL) minimal residual disease (MRD) testing. We retrospectively analyzed our existing three-tube, seven-color B-ALL MRD panel and used our novel method to develop an optimized one-tube, eight-color panel, which was tested prospectively. The optimized one-tube, eight-color panel resulted in greater efficiency of time and resources with no loss in diagnostic power. Constructing a flow cytometry panel using a rigorous, objective, quantitative method permits optimization and avoids problems of interdependence and redundancy in a large, multiantigen panel. Copyright© by the American Society for Clinical Pathology.

  15. Multiattribute selection of acute stroke imaging software platform for Extending the Time for Thrombolysis in Emergency Neurological Deficits (EXTEND) clinical trial.

    Science.gov (United States)

    Churilov, Leonid; Liu, Daniel; Ma, Henry; Christensen, Soren; Nagakane, Yoshinari; Campbell, Bruce; Parsons, Mark W; Levi, Christopher R; Davis, Stephen M; Donnan, Geoffrey A

    2013-04-01

    The appropriateness of a software platform for rapid MRI assessment of the amount of salvageable brain tissue after stroke is critical for both the validity of the Extending the Time for Thrombolysis in Emergency Neurological Deficits (EXTEND) Clinical Trial of stroke thrombolysis beyond 4.5 hours and for stroke patient care outcomes. The objective of this research is to develop and implement a methodology for selecting the acute stroke imaging software platform most appropriate for the setting of a multi-centre clinical trial. A multi-disciplinary decision making panel formulated the set of preferentially independent evaluation attributes. Alternative Multi-Attribute Value Measurement methods were used to identify the best imaging software platform followed by sensitivity analysis to ensure the validity and robustness of the proposed solution. Four alternative imaging software platforms were identified. RApid processing of PerfusIon and Diffusion (RAPID) software was selected as the most appropriate for the needs of the EXTEND trial. A theoretically grounded generic multi-attribute selection methodology for imaging software was developed and implemented. The developed methodology assured both a high quality decision outcome and a rational and transparent decision process. This development contributes to stroke literature in the area of comprehensive evaluation of MRI clinical software. At the time of evaluation, RAPID software presented the most appropriate imaging software platform for use in the EXTEND clinical trial. The proposed multi-attribute imaging software evaluation methodology is based on sound theoretical foundations of multiple criteria decision analysis and can be successfully used for choosing the most appropriate imaging software while ensuring both robust decision process and outcomes. © 2012 The Authors. International Journal of Stroke © 2012 World Stroke Organization.

  16. CymeR: cytometry analysis using KNIME, docker and R.

    Science.gov (United States)

    Muchmore, B; Alarcón-Riquelme, M E

    2017-03-01

    Here we present open-source software for the analysis of high-dimensional cytometry data using state of the art algorithms. Importantly, use of the software requires no programming ability, and output files can either be interrogated directly in CymeR or they can be used downstream with any other cytometric data analysis platform. Also, because we use Docker to integrate the multitude of components that form the basis of CymeR, we have additionally developed a proof-of-concept of how future open-source bioinformatic programs with graphical user interfaces could be developed. CymeR is open-source software that ties several components into a single program that is perhaps best thought of as a self-contained data analysis operating system. Please see https://github.com/bmuchmore/CymeR/wiki for detailed installation instructions. brian.muchmore@genyo.es or marta.alarcon@genyo.es. © The Author 2016. Published by Oxford University Press.

  17. Quantitative testing of the methodology for genome size estimation in plants using flow cytometry: a case study of the Primulina genus

    Directory of Open Access Journals (Sweden)

    Jing eWang

    2015-05-01

    Full Text Available Flow cytometry (FCM is a commonly used method for estimating genome size in many organisms. The use of flow cytometry in plants is influenced by endogenous fluorescence inhibitors and may cause an inaccurate estimation of genome size; thus, falsifying the relationship between genome size and phenotypic traits/ecological performance. Quantitative optimization of FCM methodology minimizes such errors, yet there are few studies detailing this methodology. We selected the genus Primulina, one of the most representative and diverse genera of the Old World Gesneriaceae, to evaluate the methodology effect on determining genome size. Our results showed that buffer choice significantly affected genome size estimation in six out of the eight species examined and altered the 2C-value (DNA content by as much as 21.4%. The staining duration and propidium iodide (PI concentration slightly affected the 2C-value. Our experiments showed better histogram quality when the samples were stained for 40 minutes at a PI concentration of 100 µg ml-1. The quality of the estimates was not improved by one-day incubation in the dark at 4 °C or by centrifugation. Thus, our study determined an optimum protocol for genome size measurement in Primulina: LB01 buffer supplemented with 100 µg ml-1 PI and stained for 40 minutes. This protocol also demonstrated a high universality in other Gesneriaceae genera. We report the genome size of nine Gesneriaceae species for the first time. The results showed substantial genome size variation both within and among the species, with the 2C-value ranging between 1.62 and 2.71 pg. Our study highlights the necessity of optimizing the FCM methodology prior to obtaining reliable genome size estimates in a given taxon.

  18. An unsupervised technique for optimal feature selection in attribute profiles for spectral-spatial classification of hyperspectral images

    Science.gov (United States)

    Bhardwaj, Kaushal; Patra, Swarnajyoti

    2018-04-01

    Inclusion of spatial information along with spectral features play a significant role in classification of remote sensing images. Attribute profiles have already proved their ability to represent spatial information. In order to incorporate proper spatial information, multiple attributes are required and for each attribute large profiles need to be constructed by varying the filter parameter values within a wide range. Thus, the constructed profiles that represent spectral-spatial information of an hyperspectral image have huge dimension which leads to Hughes phenomenon and increases computational burden. To mitigate these problems, this work presents an unsupervised feature selection technique that selects a subset of filtered image from the constructed high dimensional multi-attribute profile which are sufficiently informative to discriminate well among classes. In this regard the proposed technique exploits genetic algorithms (GAs). The fitness function of GAs are defined in an unsupervised way with the help of mutual information. The effectiveness of the proposed technique is assessed using one-against-all support vector machine classifier. The experiments conducted on three hyperspectral data sets show the robustness of the proposed method in terms of computation time and classification accuracy.

  19. Flow cytometry analysis of inflammatory cells isolated from the sciatic nerve and DRG after chronic constriction injury in mice.

    Science.gov (United States)

    Liu, Liping; Yin, Yan; Li, Fei; Malhotra, Charvi; Cheng, Jianguo

    2017-06-01

    Cellular responses to nerve injury play a central role in the pathogenesis of neuropathic pain. However, the analysis of site specific cellular responses to nerve injury and neuropathic pain is limited to immunohistochemistry staining with numerous limitations. We proposed to apply flow cytometry to overcome some of the limitations and developed two protocols for isolation of cells from small specimens of the sciatic nerve and dorsal root ganglion (DRG) in mice. RESULTS AND COMPARASION WITH EXISTING: methods We found that both the non-enzymatic and enzymatic approaches were highly effective in harvesting a sufficient number of cells for flow cytometry analysis in normal and pathological conditions. The total number of cells in the injury site of the sciatic and its DRGs increased significantly 14days after chronic constriction injury (CCI) of the sciatic nerve, compared to sham surgery control or the contralateral control. The enzymatic approach yielded a significantly higher total number of cells and CD45 negative cells, suggesting that this approach allows for harvest of more resident cells, compared to the non-enzymatic method. The percentage of CD45 + /CD11b + cells was significantly increased in the sciatic nerve but not in the DRG. These results were consistent with both protocols. We thus offer two simple and effective protocols that allow for application of flow cytometry to the investigation of cellular and molecular mechanisms of neuropathic pain. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Immunophenotype Discovery, Hierarchical Organization, and Template-based Classification of Flow Cytometry Samples

    Directory of Open Access Journals (Sweden)

    Ariful Azad

    2016-08-01

    Full Text Available We describe algorithms for discovering immunophenotypes from large collections of flow cytometry (FC samples, and using them to organize the samples into a hierarchy based on phenotypic similarity. The hierarchical organization is helpful for effective and robust cytometry data mining, including the creation of collections of cell populations characteristic of different classes of samples, robust classification, and anomaly detection. We summarize a set of samples belonging to a biological class or category with a statistically derived template for the class. Whereas individual samples are represented in terms of their cell populations (clusters, a template consists of generic meta-populations (a group of homogeneous cell populations obtained from the samples in a class that describe key phenotypes shared among all those samples. We organize an FC data collection in a hierarchical data structure that supports the identification of immunophenotypes relevant to clinical diagnosis. A robust template-based classification scheme is also developed, but our primary focus is in the discovery of phenotypic signatures and inter-sample relationships in an FC data collection. This collective analysis approach is more efficient and robust since templates describe phenotypic signatures common to cell populations in several samples, while ignoring noise and small sample-specific variations.We have applied the template-base scheme to analyze several data setsincluding one representing a healthy immune system, and one of Acute Myeloid Leukemia (AMLsamples. The last task is challenging due to the phenotypic heterogeneity of the severalsubtypes of AML. However, we identified thirteen immunophenotypes corresponding to subtypes of AML, and were able to distinguish Acute Promyelocytic Leukemia from other subtypes of AML.

  1. Attenuation-based automatic kilovolt (kV)-selection in computed tomography of the chest: effects on radiation exposure and image quality.

    Science.gov (United States)

    Eller, Achim; Wuest, Wolfgang; Scharf, Michael; Brand, Michael; Achenbach, Stephan; Uder, Michael; Lell, Michael M

    2013-12-01

    To evaluate an automated attenuation-based kV-selection in computed tomography of the chest in respect to radiation dose and image quality, compared to a standard 120 kV protocol. 104 patients were examined using a 128-slice scanner. Fifty examinations (58 ± 15 years, study group) were performed using the automated adaption of tube potential (100-140 kV), based on the attenuation profile of the scout scan, 54 examinations (62 ± 14 years, control group) with fixed 120 kV. Estimated CT dose index (CTDI) of the software-proposed setting was compared with a 120 kV protocol. After the scan CTDI volume (CTDIvol) and dose length product (DLP) were recorded. Image quality was assessed by region of interest (ROI) measurements, subjective image quality by two observers with a 4-point scale (3--excellent, 0--not diagnostic). The algorithm selected 100 kV in 78% and 120 kV in 22%. Overall CTDIvol reduction was 26.6% (34% in 100 kV) overall DLP reduction was 22.8% (32.1% in 100 kV) (all pimage quality was excellent in both groups. The attenuation based kV-selection algorithm enables relevant dose reduction (~27%) in chest-CT while keeping image quality parameters at high levels. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  2. Computed tomography with selectable image resolution

    International Nuclear Information System (INIS)

    Dibianca, F.A.; Dallapiazza, D.G.

    1981-01-01

    A computed tomography system x-ray detector has a central group of half-width detector elements and groups of full-width elements on each side of the central group. To obtain x-ray attenuation data for whole body layers, the half-width elements are switched effectively into paralleled pairs so all elements act like full-width elements and an image of normal resolution is obtained. For narrower head layers, the elements in the central group are used as half-width elements so resolution which is twice as great as normal is obtained. The central group is also used in the half-width mode and the outside groups are used in the full-width mode to obtain a high resolution image of a body zone within a full body layer. In one embodiment data signals from the detector are switched by electronic multiplexing and in another embodiment a processor chooses the signals for the various kinds of images that are to be reconstructed. (author)

  3. A flow cytometry-based workflow for detection and quantification of anti-plasmodial antibodies in vaccinated and naturally exposed individuals

    DEFF Research Database (Denmark)

    Ajua, Anthony; Engleitner, Thomas; Esen, Meral

    2012-01-01

    information about natural exposure and vaccine immunogenicity. A novel, cytometry-based workflow for the quantitative detection of anti-plasmodial antibodies in human serum is presented. METHODS: Fixed red blood cells (RBCs), infected with late stages of P. falciparum were utilized to detect malaria...... vaccine trials in semiimmune adults and pre-school children residing in a malaria endemic area. RESULTS: Fixation, permeabilization, and staining of infected RBCs were adapted for best operation in flow cytometry. As asexual vaccine candidates are designed to induce antibody patterns similar to semi...... with those obtained by manual gating (r between 0.79 and 0.99) and outperformed other model-driven gating methods. Bland-Altman plots confirmed the agreement of manual gating and OSA derived results. A-1.33 fold increase (p=0.003) in the number of positive cells after vaccination in a subgroup of preschool...

  4. Unilateral uveitis masquerade syndrome caused by diffuse large B-cell lymphoma diagnosed using multiparametric flow cytometry of the aqueous humor.

    Science.gov (United States)

    Monsalvo, Silvia; Serrano, Cristina; Prieto, Elena; Fernández-Sanz, Guillermo; Puente, Maria-Camino; Rodriguez-Pinilla, Maria; Garcia Raso, Aranzazu; Llamas, Pilar; Cordoba, Raul

    2017-07-01

    The uveitis masquerade syndromes (UMS) are a group of ocular diseases that may mimic chronic intraocular inflammation. Many malignant entities such as non-Hodgkin's lymphomas may masquerade as uveitis. We report a case of an HIV-positive patient with masquerade syndrome presenting unilateral uveitis. 45-year-old Caucasian man with a diagnosis of diffuse large B-cell lymphoma (DLBCL). The patient was diagnosed by a biopsy of an abdominal mass which showed fragments of gastric mucosa with diffuse growth of neoplastic cells. At diagnosis, the patient suffered from unilateral blurring of vision and a sudden decrease of left-eye visual acuity. A slit-lamp examination of the left eye revealed a diagnosis of anterior uveitis. The patient exhibited no signs of posterior uveitis. An anterior-chamber paracentesis was performed and analyzed by multiparameter flow cytometry (MFC), showing cells CD45, CD19, CD20, CD22, and CD38 positives, and moderate expression of CD10 with kappa light chain restriction, showing a monoclonal B-cell population. The patient received CHOP-R with intrathecal methotrexate followed by consolidation high dose methotrexate obtaining a complete response which is ongoing. Differential diagnosis between chronic uveitis and ocular lymphoma may be challenging. We advocate anterior-chamber paracentesis in cases of refractory uveitis in patients with hematologic malignancies. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

  5. Prognostic value of flow cytometry in surgically treated primary gastric lymphoma Valor pronóstico de la citometría de flujo en el linfoma gástrico

    Directory of Open Access Journals (Sweden)

    F. Fernández

    2006-11-01

    Full Text Available Aim: to investigate whether flow cytometry could help to define the optimal therapeutic strategy of primary gastric lymphomas. Material and method: retrospective study of 46 patients having primary gastric lymphoma -according to Dawson criteria- in Ann Arbor stage I E and II E, who were surgically treated. From selected paraffin-embedded tissue blocks of the tumor, DNA content was studied by flow cytometry (FC. Other pathological tumor features were analysed by hematoxiline-eosine and Giemsa stains as well as immunohistochemical study; any possible influence on postoperative survival was investigated through statistical analysis. Results: the DNA ploidy pattern was diploid in 40 cases (87% and aneuploid (hyperdiploid in 6 (13%. Postoperative survival probability (PSP was 62.7% at 5 years. Statistical analysis showed significant prognostic value for Ann Arbor classification -with higher PSP for stage I E (p = 0.009- and FC parameters: diploid tumors had higher PSP than aneuploid tumors. Also tumors having S-phase (p = 0.044 or G2-M phase values (p = 0.023 under the respective mean values had higher PSP. No influence on PSP was found for wall invasion, Helicobacter pylori infection, Isaacson's histologic type or resection margin involvement. No significant relationship was appreciated between Isaacson's histologic type and DNA ploidy patterns. Conclusion: FC could be useful in assessing gastric lymphoma prognosis.

  6. Image quality (IQ) guided multispectral image compression

    Science.gov (United States)

    Zheng, Yufeng; Chen, Genshe; Wang, Zhonghai; Blasch, Erik

    2016-05-01

    Image compression is necessary for data transportation, which saves both transferring time and storage space. In this paper, we focus on our discussion on lossy compression. There are many standard image formats and corresponding compression algorithms, for examples, JPEG (DCT -- discrete cosine transform), JPEG 2000 (DWT -- discrete wavelet transform), BPG (better portable graphics) and TIFF (LZW -- Lempel-Ziv-Welch). The image quality (IQ) of decompressed image will be measured by numerical metrics such as root mean square error (RMSE), peak signal-to-noise ratio (PSNR), and structural Similarity (SSIM) Index. Given an image and a specified IQ, we will investigate how to select a compression method and its parameters to achieve an expected compression. Our scenario consists of 3 steps. The first step is to compress a set of interested images by varying parameters and compute their IQs for each compression method. The second step is to create several regression models per compression method after analyzing the IQ-measurement versus compression-parameter from a number of compressed images. The third step is to compress the given image with the specified IQ using the selected compression method (JPEG, JPEG2000, BPG, or TIFF) according to the regressed models. The IQ may be specified by a compression ratio (e.g., 100), then we will select the compression method of the highest IQ (SSIM, or PSNR). Or the IQ may be specified by a IQ metric (e.g., SSIM = 0.8, or PSNR = 50), then we will select the compression method of the highest compression ratio. Our experiments tested on thermal (long-wave infrared) images (in gray scales) showed very promising results.

  7. Feature-size dependent selective edge enhancement of x-ray images

    International Nuclear Information System (INIS)

    Herman, S.

    1988-01-01

    Morphological filters are nonlinear signal transformations that operate on a picture directly in the space domain. Such filters are based on the theory of mathematical morphology previously formulated. The filt4er being presented here features a ''mask'' operator (called a ''structuring element'' in some of the literature) which is a function of the two spatial coordinates x and y. The two basic mathematical operations are called ''masked erosion'' and ''masked dilation''. In the case of masked erosion the mask is passed over the input image in a raster pattern. At each position of the mask, the pixel values under the mask are multiplied by the mask pixel values. Then the output pixel value, located at the center position of the mask,is set equal to the minimum of the product of the mask and input values. Similarity, for masked dilation, the output pixel value is the maximum of the product of the input and the mask pixel values. The two basic processes of dilation and erosion can be used to construct the next level of operations the ''positive sieve'' (also called ''opening'') and the ''negative sieve'' (''closing''). The positive sieve modifies the peaks in the image whereas the negative sieve works on image valleys. The positive sieve is implemented by passing the output of the masked erosion step through the masked dilation function. The negative sieve reverses this procedure, using a dilation followed by an erosion. Each such sifting operator is characterized by a ''hole size''. It will be shown that the choice of hole size will select the range of pixel detail sizes which are to be enhanced. The shape of the mask will govern the shape of the enhancement. Finally positive sifting is used to enhance positive-going (peak) features, whereas negative enhances the negative-going (valley) landmarks

  8. Robust selectivity to two-object images in human visual cortex

    Science.gov (United States)

    Agam, Yigal; Liu, Hesheng; Papanastassiou, Alexander; Buia, Calin; Golby, Alexandra J.; Madsen, Joseph R.; Kreiman, Gabriel

    2010-01-01

    SUMMARY We can recognize objects in a fraction of a second in spite of the presence of other objects [1–3]. The responses in macaque areas V4 and inferior temporal cortex [4–15] to a neuron’s preferred stimuli are typically suppressed by the addition of a second object within the receptive field (see however [16, 17]). How can this suppression be reconciled with rapid visual recognition in complex scenes? One option is that certain “special categories” are unaffected by other objects [18] but this leaves the problem unsolved for other categories. Another possibility is that serial attentional shifts help ameliorate the problem of distractor objects [19–21]. Yet, psychophysical studies [1–3], scalp recordings [1] and neurophysiological recordings [14, 16, 22–24], suggest that the initial sweep of visual processing contains a significant amount of information. We recorded intracranial field potentials in human visual cortex during presentation of flashes of two-object images. Visual selectivity from temporal cortex during the initial ~200 ms was largely robust to the presence of other objects. We could train linear decoders on the responses to isolated objects and decode information in two-object images. These observations are compatible with parallel, hierarchical and feed-forward theories of rapid visual recognition [25] and may provide a neural substrate to begin to unravel rapid recognition in natural scenes. PMID:20417105

  9. Organizing the Cellular and Molecular Heterogeneity in High-Grade Serous Ovarian Cancer by Mass Cytometry

    Science.gov (United States)

    2013-10-01

    Acknowledgements The authors wish to thank Drs Scott Tanner, Olga Ornatsky, Dmitry Bandura , Mitch Winnik and Mark Nitz for their critical reading of this...Quality assurance for polychromatic flow cytometry using a suite of calibration beads. Nat Protoc 2012, 7:2067-2079. 23. Baranov VI, Quinn Z, Bandura ... Bandura DR, Tanner SD, Dick J: Multiple cellular antigen detection by ICP-MS. J Immunol Methods 2006, 308:68-76. 25. Ornatsky OI, Kinach R, Bandura DR

  10. Immunological Tools: Engaging Students in the Use and Analysis of Flow Cytometry and Enzyme-linked Immunosorbent Assay (ELISA)

    Science.gov (United States)

    Ott, Laura E.; Carson, Susan

    2014-01-01

    Flow cytometry and enzyme-linked immunosorbent assay (ELISA) are commonly used techniques associated with clinical and research applications within the immunology and medical fields. The use of these techniques is becoming increasingly valuable in many life science and engineering disciplines as well. Herein, we report the development and…

  11. Traction cytometry: regularization in the Fourier approach and comparisons with finite element method.

    Science.gov (United States)

    Kulkarni, Ankur H; Ghosh, Prasenjit; Seetharaman, Ashwin; Kondaiah, Paturu; Gundiah, Namrata

    2018-05-09

    Traction forces exerted by adherent cells are quantified using displacements of embedded markers on polyacrylamide substrates due to cell contractility. Fourier Transform Traction Cytometry (FTTC) is widely used to calculate tractions but has inherent limitations due to errors in the displacement fields; these are mitigated through a regularization parameter (γ) in the Reg-FTTC method. An alternate finite element (FE) approach computes tractions on a domain using known boundary conditions. Robust verification and recovery studies are lacking but essential in assessing the accuracy and noise sensitivity of the traction solutions from the different methods. We implemented the L2 regularization method and defined a maximum curvature point in the traction with γ plot as the optimal regularization parameter (γ*) in the Reg-FTTC approach. Traction reconstructions using γ* yield accurate values of low and maximum tractions (Tmax) in the presence of up to 5% noise. Reg-FTTC is hence a clear improvement over the FTTC method but is inadequate to reconstruct low stresses such as those at nascent focal adhesions. FE, implemented using a node-by-node comparison, showed an intermediate reconstruction compared to Reg-FTTC. We performed experiments using mouse embryonic fibroblast (MEF) and compared results between these approaches. Tractions from FTTC and FE showed differences of ∼92% and 22% as compared to Reg-FTTC. Selection of an optimum value of γ for each cell reduced variability in the computed tractions as compared to using a single value of γ for all the MEF cells in this study.

  12. Magnetite/CdTe magnetic-fluorescent composite nanosystem for magnetic separation and bio-imaging

    International Nuclear Information System (INIS)

    Kale, Anup; Yadav, Prasad; Gholap, Haribhau; Jog, J P; Ogale, Satishchandra; Kale, Sonia; Shastry, Padma; Pasricha, Renu; Lefez, Benoit; Hannoyer, Beatrice

    2011-01-01

    A new synthesis protocol is described to obtain a CdTe decorated magnetite bifunctional nanosystem via dodecylamine (DDA) as cross linker. High resolution transmission electron microscopy (HRTEM), energy-dispersive x-ray spectroscopy (EDAX), vibrating sample magnetometry (VSM), Fourier transform infrared spectroscopy (FTIR), diffuse reflectance spectroscopy (DRS) and fluorescence microscopy are used to characterize the constitution, size, composition and physical properties of these superparamagnetic-fluorescent nanoparticles. These CdTe decorated magnetite nanoparticles were then functionalized with anti-epidermal growth factor receptor (EGFR) antibody to specifically target cells expressing this receptor. The EGFR is a transmembrane glycoprotein and is expressed on tumor cells from different tissue origins including human leukemic cell line Molt-4 cells. The magnetite-CdTe composite nanosystem is shown to perform excellently for specific selection, magnetic separation and fluorescent detection of EGFR positive Molt-4 cells from a mixed population. Flow cytometry and confocal laser scanning microscopy results show that this composite nanosystem has great potential in antibody functionalized magnetic separation and imaging of cells using cell surface receptor antibody.

  13. Flow cytometry in the diagnosis of myelodysplastic syndromes (MDS) and the value of myeloid nuclear differentiation antigen (MNDA).

    Science.gov (United States)

    Bellos, Frauke; Kern, Wolfgang

    2014-09-25

    Background: Confirming diagnosis of myelodysplastic syndromes (MDS) is often challenging. Standard diagnostic methods are cytomorphology (CM) and cytogenetics (CG). Multiparameter flow cytometry (MFC) is upcoming in MDS diagnostic work up, comparability and investigator experience are critical. Myeloid nuclear differentiation antigen (MNDA) in myelomonocytic cells might be expressed more weakly in patients with MDS. The analysis of MNDA may thus improve diagnostic capabilities of MFC in MDS. Methods: Staining methods and antibody combinations for MFC in MDS are outlined, giving details for interpretation of results in regard to dyspoiesis. MFC results are correlated with CM and CG and with survival data. Use of myeloid nuclear differentiation antigen (MNDA) in MDS diagnostics was evaluated in 239 patients with MDS, AML, other cytopenic conditions and in 30 negative controls. Results: Strong correlation between findings in CM and MFC was found; MFC results correlated well with those of CG. Patients with higher grades of dysplasia in MFC had shorter overall survival. Percentages of granulocytes and monocytes with diminished MNDA expression (%dimG, %dimM) were higher in patients with MDS and AML. Mean fluorescence intensity (MFI) of MNDA in monocytes was lower in MDS and AML. Cut-off values for %dimG (12%) and %dimM (22%) as well as for MFI in monocytes (72) were defined discriminating between MDS and non-MDS. Conclusion: MFC adds significant information on dyspoiesis in the diagnostic work up for MDS and provides prognostic information. MNDA expression can be assessed by MFC and may facilitate evaluation of dyspoiesis when added to MDS MFC panels. © 2014 Clinical Cytometry Society. Copyright © 2014 Clinical Cytometry Society.

  14. Filter and slice thickness selection in SPECT image reconstruction

    International Nuclear Information System (INIS)

    Ivanovic, M.; Weber, D.A.; Wilson, G.A.; O'Mara, R.E.

    1985-01-01

    The choice of filter and slice thickness in SPECT image reconstruction as function of activity and linear and angular sampling were investigated in phantom and patient imaging studies. Reconstructed transverse and longitudinal spatial resolution of the system were measured using a line source in a water filled phantom. Phantom studies included measurements of the Data Spectrum phantom; clinical studies included tomographic procedures in 40 patients undergoing imaging of the temporomandibular joint. Slices of the phantom and patient images were evaluated for spatial of the phantom and patient images were evaluated for spatial resolution, noise, and image quality. Major findings include; spatial resolution and image quality improve with increasing linear sampling frequencies over the range of 4-8 mm/p in the phantom images, best spatial resolution and image quality in clinical images were observed at a linear sampling frequency of 6mm/p, Shepp and Logan filter gives the best spatial resolution for phantom studies at the lowest linear sampling frequency; smoothed Shepp and Logan filter provides best quality images without loss of resolution at higher frequencies and, spatial resolution and image quality improve with increased angular sampling frequency in the phantom at 40 c/p but appear to be independent of angular sampling frequency at 400 c/p

  15. Detection of microparticles from human red blood cells by multiparametric flow cytometry

    Science.gov (United States)

    Grisendi, Giulia; Finetti, Elena; Manganaro, Daniele; Cordova, Nicoletta; Montagnani, Giuliano; Spano, Carlotta; Prapa, Malvina; Guarneri, Valentina; Otsuru, Satoru; Horwitz, Edwin M.; Mari, Giorgio; Dominici, Massimo

    2015-01-01

    Background During storage, red blood cells (RBC) undergo chemical and biochemical changes referred to as “storage lesions”. These events determine the loss of RBC integrity, resulting in lysis and release of microparticles. There is growing evidence of the clinical importance of microparticles and their role in blood transfusion-related side effects and pathogen transmission. Flow cytometry is currently one of the most common techniques used to quantify and characterise microparticles. Here we propose multiparametric staining to monitor and quantify the dynamic release of microparticles by stored human RBC. Material and methods RBC units (n=10) were stored under blood bank conditions for up to 42 days. Samples were tested at different time points to detect microparticles and determine the haemolysis rate (HR%). Microparticles were identified by flow cytometry combining carboxyfluorescein diacetate succinimidyl ester (CFSE) dye, annexin V and anti-glycophorin A antibody. Results We demonstrated that CFSE can be successfully used to label closed vesicles with an intact membrane. The combination of CFSE and glycophorin A antibody was effective for monitoring and quantifying the dynamic release of microparticles from RBC during storage. Double staining with CFSE/glycophorin A was a more precise approach, increasing vesicle detection up to 4.7-fold vs the use of glycophorin A/annexin V alone. Moreover, at all the time points tested, we found a robust correlation (R=0.625; p=0.0001) between HR% and number of microparticles detected. Discussion Multiparametric staining, based on a combination of CFSE, glycophorin A antibody and annexin V, was able to detect, characterise and monitor the release of microparticles from RBC units during storage, providing a sensitive approach to labelling and identifying microparticles for transfusion medicine and, more broadly, for cell-based therapies. PMID:25369588

  16. Reproducible diagnosis of chronic lymphocytic leukemia by flow cytometry: An European Research Initiative on CLL (ERIC) & European Society for Clinical Cell Analysis (ESCCA) Harmonisation project.

    Science.gov (United States)

    Rawstron, Andy C; Kreuzer, Karl-Anton; Soosapilla, Asha; Spacek, Martin; Stehlikova, Olga; Gambell, Peter; McIver-Brown, Neil; Villamor, Neus; Psarra, Katherina; Arroz, Maria; Milani, Raffaella; de la Serna, Javier; Cedena, M Teresa; Jaksic, Ozren; Nomdedeu, Josep; Moreno, Carol; Rigolin, Gian Matteo; Cuneo, Antonio; Johansen, Preben; Johnsen, Hans E; Rosenquist, Richard; Niemann, Carsten Utoft; Kern, Wolfgang; Westerman, David; Trneny, Marek; Mulligan, Stephen; Doubek, Michael; Pospisilova, Sarka; Hillmen, Peter; Oscier, David; Hallek, Michael; Ghia, Paolo; Montserrat, Emili

    2018-01-01

    The diagnostic criteria for CLL rely on morphology and immunophenotype. Current approaches have limitations affecting reproducibility and there is no consensus on the role of new markers. The aim of this project was to identify reproducible criteria and consensus on markers recommended for the diagnosis of CLL. ERIC/ESCCA members classified 14 of 35 potential markers as "required" or "recommended" for CLL diagnosis, consensus being defined as >75% and >50% agreement, respectively. An approach to validate "required" markers using normal peripheral blood was developed. Responses were received from 150 participants with a diagnostic workload >20 CLL cases per week in 23/150 (15%), 5-20 in 82/150 (55%), and 97% concordance with current approaches. A pilot study to validate staining quality was completed in 11 centers. Markers considered as "required" for the diagnosis of CLL by the participants in this study (CD19, CD5, CD20, CD23, Kappa, and Lambda) are consistent with current diagnostic criteria and practice. Importantly, a reproducible approach to validate and apply these markers in individual laboratories has been identified. Finally, a consensus "recommended" panel of markers to refine diagnosis in borderline cases (CD43, CD79b, CD81, CD200, CD10, and ROR1) has been defined and will be prospectively evaluated. © 2017 International Clinical Cytometry Society. © 2017 The Authors. Cytometry Part B: Clinical Cytometry published by Wiley Periodicals, Inc. on behalf of International Clinical Cytometry Society.

  17. Generic maximum likely scale selection

    DEFF Research Database (Denmark)

    Pedersen, Kim Steenstrup; Loog, Marco; Markussen, Bo

    2007-01-01

    in this work is on applying this selection principle under a Brownian image model. This image model provides a simple scale invariant prior for natural images and we provide illustrative examples of the behavior of our scale estimation on such images. In these illustrative examples, estimation is based......The fundamental problem of local scale selection is addressed by means of a novel principle, which is based on maximum likelihood estimation. The principle is generally applicable to a broad variety of image models and descriptors, and provides a generic scale estimation methodology. The focus...

  18. Diffusion and Perfusion MR Imaging in Acute Stroke: Clinical Utility and Potential Limitations for Treatment Selection

    DEFF Research Database (Denmark)

    Bateman, Mathew; Slater, Lee-Anne; Leslie-Mazwi, Thabele M

    2017-01-01

    Magnetic resonance (MR) diffusion-weighted imaging (DWI) and perfusion-weighted imaging (PWI) offer unique insight into acute ischemic stroke pathophysiology. These techniques may offer the ability to apply pathophysiology to accurately individualize acute stroke reperfusion treatment, including ...... to be investigated in ongoing randomized controlled trials, and continued research into these techniques will help achieve the goal of tissue-based decision making and individualized acute stroke treatment....... extending the opportunity of reperfusion treatment to well beyond the current time-based treatment windows. This review examines the use of DWI and PWI in the major stroke trials, their current clinical utility, and potential limitations for reperfusion treatment selection. DWI and PWI continue...

  19. Cyanine 5.5 conjugated nanobubbles as a tumor selective contrast agent for dual ultrasound-fluorescence imaging in a mouse model.

    Directory of Open Access Journals (Sweden)

    Liyi Mai

    Full Text Available Nanobubbles and microbubbles are non-invasive ultrasound imaging contrast agents that may potentially enhance diagnosis of tumors. However, to date, both nanobubbles and microbubbles display poor in vivo tumor-selectivity over non-targeted organs such as liver. We report here cyanine 5.5 conjugated nanobubbles (cy5.5-nanobubbles of a biocompatible chitosan-vitamin C lipid system as a dual ultrasound-fluorescence contrast agent that achieved tumor-selective imaging in a mouse tumor model. Cy5.5-nanobubble suspension contained single bubble spheres and clusters of bubble spheres with the size ranging between 400-800 nm. In the in vivo mouse study, enhancement of ultrasound signals at tumor site was found to persist over 2 h while tumor-selective fluorescence emission was persistently observed over 24 h with intravenous injection of cy5.5-nanobubbles. In vitro cell study indicated that cy5.5-flurescence dye was able to accumulate in cancer cells due to the unique conjugated nanobubble structure. Further in vivo fluorescence study suggested that cy5.5-nanobubbles were mainly located at tumor site and in the bladder of mice. Subsequent analysis confirmed that accumulation of high fluorescence was present at the intact subcutaneous tumor site and in isolated tumor tissue but not in liver tissue post intravenous injection of cy5.5-nanobubbles. All these results led to the conclusion that cy5.5-nanobubbles with unique crosslinked chitosan-vitamin C lipid system have achieved tumor-selective imaging in vivo.

  20. PET imaging with [18F]fluoroethoxybenzovesamicol ([18F]FEOBV) following selective lesion of cholinergic pedunculopontine tegmental neurons in rat

    International Nuclear Information System (INIS)

    Cyr, Marilyn; Parent, Maxime J.; Mechawar, Naguib; Rosa-Neto, Pedro; Soucy, Jean-Paul; Aliaga, Antonio; Kostikov, Alexey; Maclaren, Duncan A.A.; Clark, Stewart D.; Bedard, Marc-Andre

    2014-01-01

    Introduction: [ 18 F]fluoroethoxybenzovesamicol ([ 18 F]FEOBV) is a PET radiotracer with high selectivity and specificity to the vesicular acetylcholine transporter (VAChT). It has been shown to be a sensitive in vivo measurement of changes of cholinergic innervation densities following lesion of the nucleus basalis of Meynert (NBM) in rat. The current study used [ 18 F]FEOBV with PET imaging to detect the effect of a highly selective lesion of the pedunculopontine (PPTg) nucleus in rat. Methods: After bilateral and selective lesions of the PPTg cholinergic neurons, rats were scanned using [ 18 F]FEOBV, then sacrificed, and their brain tissues collected for immunostaining and quantification of the VAChT. Results: Comparisons with control rats revealed that cholinergic losses can be detected in the brainstem, lateral thalamus, and pallidum by using both in vivo imaging methods with [ 18 F]FEOBV, and ex vivo measurements. In the brainstem PPTg area, significant correlations were observed between in vivo and ex vivo measurements, while this was not the case in the thalamic and pallidal projection sites. Conclusions: These findings support PET imaging with [ 18 F]FEOBV as a reliable in vivo method for the detection of neuronal terminal losses resulting from lesion of the PPTg. Useful applications can be found in the study of neurodegenerative diseases in human, such as Parkinson’s disease, multiple system atrophy, progressive supranuclear palsy, or dementia with Lewy bodies

  1. Preconception sex selection for non-medical and intermediate reasons: ethical reflections.

    Science.gov (United States)

    de Wert, G; Dondorp, W

    2010-01-01

    Sex selection for non-medical reasons is forbidden in many countries. Focusing on preconception sex selection, the authors first observe that it is unclear what should count as a 'medical reason' in this context and argue for the existence of 'intermediate reasons' that do not fit well within the rigid distinction between 'medical'and 'non-medical'. The article further provides a critical review of the arguments for the prohibition of sex selection for non-medical reasons and finds that none of these are conclusive. The authors conclude that the ban should be reconsidered, but also that existing-- societal concerns about possible harmful effects should be taken seriously. Measures to this effect may include limiting the practice to couples who already have at least one child of the sex opposite to that which they now want to select ('family balancing'). Finally, a difficult set of questions is raised by concerns about the reliability and unproven (long-term) safety of the only technology (flow cytometry) proven to work.

  2. 3D-imaging of selective laser melting defects in a Co–Cr–Mo alloy by synchrotron radiation micro-CT

    International Nuclear Information System (INIS)

    Zhou, Xin; Wang, Dianzheng; Liu, Xihe; Zhang, DanDan; Qu, Shilian; Ma, Jing; London, Gary; Shen, Zhijian; Liu, Wei

    2015-01-01

    Microstructure defects set the mechanical property limits for solid Co–Cr–Mo alloy prepared by selective laser melting (SLM). Previous studies were mainly based on 2D SEM images and thus not able to provide information of the 3D morphologies of the complex defects. In this paper, the remaining porosities in Co–Cr–Mo alloy parts prepared by selective laser melting were presented in relation to the laser processing parameters. In order to understand the defect forming mechanism, accurate 3D images of defects inside SLM fabricated Co–Cr–Mo samples were provided by synchrotron radiation micro-CT imaging of 300 μm thick slices cut from a 10 mm cube. With 3D reconstructed images distinctive morphologies of SLM defects spanning across the consolidated powder layers were generated. The faults can be classified as single layer or multi-layers defects. The accidental single layer defects form as gaps between adjacent laser melt tracks or melt track discontinuousness caused by inherent fluid instability under various disturbances. The first formed single layer defect generates often a multi-layer defect spanning for 2–3 subsequent powder layers. By stabilizing the melt pool flow and by reducing the surface roughness through adjusting processing parameters it appears possible to reduce the defect concentrations

  3. Development of an Immunomagnetic Separation Method for Viable Salmonella Typhimurium Detected by Flow Cytometry

    DEFF Research Database (Denmark)

    Ahmed, Shakil; Rubahn, Horst-Günter; Erdmann, Helmut

    2016-01-01

    for detection of food-related bacteria. In this study, a flow cytometry based immunomagnetic separation (IMS) method for the isolation and enrichment of Salmonella Typhimurium from liquid samples was developed and optimized. Both polyclonal and monoclonal antibodies have been used to couple with 1 micron sized...... and bacteria, immunocapture time, staining and buffering conditions for the viability assays were optimized. The capture efficiency of IMS was>98% for a range of Salmonella Typhimurium cell concentrations from 103 to 105/mL using 108/mL bead concentration. The method proved to have high (98%) specificity...

  4. Cell proliferation and apoptosis in the primary enamel knot measured by flow cytometry of laser microdissected samples

    Czech Academy of Sciences Publication Activity Database

    Matalová, Eva; Dubská, L.; Fleischmannová, Jana; Chlastáková, Ivana; Janečková, Eva; Tucker, A. S.

    2010-01-01

    Roč. 55, č. 8 (2010), s. 570-575 ISSN 0003-9969 R&D Projects: GA AV ČR KJB500450802; GA AV ČR IAA600450904; GA ČR GA203/08/1680 Institutional research plan: CEZ:AV0Z50450515 Keywords : Laser capture microdissection * Flow cytometry * Primary enamel knot Subject RIV: EA - Cell Biology Impact factor: 1.463, year: 2010

  5. Asymmetric-detection time-stretch optical microscopy (ATOM) for ultrafast high-contrast cellular imaging in flow

    Science.gov (United States)

    Wong, Terence T. W.; Lau, Andy K. S.; Ho, Kenneth K. Y.; Tang, Matthew Y. H.; Robles, Joseph D. F.; Wei, Xiaoming; Chan, Antony C. S.; Tang, Anson H. L.; Lam, Edmund Y.; Wong, Kenneth K. Y.; Chan, Godfrey C. F.; Shum, Ho Cheung; Tsia, Kevin K.

    2014-01-01

    Accelerating imaging speed in optical microscopy is often realized at the expense of image contrast, image resolution, and detection sensitivity – a common predicament for advancing high-speed and high-throughput cellular imaging. We here demonstrate a new imaging approach, called asymmetric-detection time-stretch optical microscopy (ATOM), which can deliver ultrafast label-free high-contrast flow imaging with well delineated cellular morphological resolution and in-line optical image amplification to overcome the compromised imaging sensitivity at high speed. We show that ATOM can separately reveal the enhanced phase-gradient and absorption contrast in microfluidic live-cell imaging at a flow speed as high as ~10 m/s, corresponding to an imaging throughput of ~100,000 cells/sec. ATOM could thus be the enabling platform to meet the pressing need for intercalating optical microscopy in cellular assay, e.g. imaging flow cytometry – permitting high-throughput access to the morphological information of the individual cells simultaneously with a multitude of parameters obtained in the standard assay. PMID:24413677

  6. Selecting optimal monochromatic level with spectral CT imaging for improving imaging quality in hepatic venography

    International Nuclear Information System (INIS)

    Sun Jun; Luo Xianfu; Wang Shou'an; Wang Jun; Sun Jiquan; Wang Zhijun; Wu Jingtao

    2013-01-01

    Objective: To investigate the effect of spectral CT monochromatic images for improving imaging quality in hepatic venography. Methods: Thirty patients underwent spectral CT examination on a GE Discovery CT 750 HD scanner. During portal phase, 1.25 mm slice thickness polychromatic images and optimal monochromatic images were obtained, and volume rendering and maximum intensity projection were created to show the hepatic veins respectively. The overall imaging quality was evaluated on a five-point scale by two radiologists. Inter-observer agreement in subjective image quality grading was assessed by Kappa statistics. Paired-sample t test were used to compare hepatic vein attenuation, hepatic parenchyma attenuation, CT value difference between the hepatic vein and the liver parenchyma, image noise, vein-to-liver contrast-to-noise ratio (CNR), the image quality score of hepatic venography between the two image data sets. Results: The monochromatic images at 50 keV were found to demonstrate the best CNR for hepatic vein.The hepatic vein attenuation [(329 ± 47) HU], hepatic parenchyma attenuation [(178 ± 33) HU], CT value difference between the hepatic vein and the liver parenchyma [(151 ± 33) HU], image noise (17.33 ± 4.18), CNR (9.13 ± 2.65), the image quality score (4.2 ± 0.6) of optimal monochromatic images were significantly higher than those of polychromatic images [(149 ± 18) HU], [(107 ± 14) HU], [(43 ±11) HU], 12.55 ± 3.02, 3.53 ± 1.03, 3.1 ± 0.8 (t values were 24.79, 13.95, 18.85, 9.07, 13.25 and 12.04, respectively, P < 0.01). In the comparison of image quality, Kappa value was 0.81 with optimal monochromatic images and 0.69 with polychromatic images. Conclusion: Monochromatic images of spectral CT could improve CNR for displaying hepatic vein and improve the image quality compared to the conventional polychromatic images. (authors)

  7. BMC Ecology image competition 2014: the winning images.

    Science.gov (United States)

    Harold, Simon; Henderson, Caspar; Baguette, Michel; Bonsall, Michael B; Hughes, David; Settele, Josef

    2014-08-29

    BMC Ecology showcases the winning entries from its second Ecology Image Competition. More than 300 individual images were submitted from an international array of research scientists, depicting life on every continent on earth. The journal's Editorial Board and guest judge Caspar Henderson outline why their winning selections demonstrated high levels of technical skill and aesthetic sense in depicting the science of ecology, and we also highlight a small selection of highly commended images that we simply couldn't let you miss out on.

  8. MR imaging of osteonecrosis using frequency selective chemical shift sequences; Neue Aspekte in der MR-Diagnostik der Osteonekrose: Selektive Fett/Wasser-Bildgebung

    Energy Technology Data Exchange (ETDEWEB)

    Duda, S H [Abt. fuer Radiologische Diagnostik, Tuebingen Univ. (Germany); Laniado, M [Abt. fuer Radiologische Diagnostik, Tuebingen Univ. (Germany); Schick, F [Inst. fuer Physik, Tuebingen Univ. (Germany)

    1994-12-31

    The MR appearance of osteonecrosis was assessed on selective fat- and water images to further evaluate the nature of double-line sign. Conventional T1- and T2-weighted SE and frequency selective chemical shift images of eight patients with avascular necrosis of the femoral head and three patients with bone infarcts were retrospectively reviewed. Eight of 11 patients showed a double-line sign on T2-weighted SE images. In these cases, correlation with selective water images revealed that a chemical shift artifact contributed to appearance and location of the hyperintense line. The authors conclude that chemical shift imaging improves our understanding of the nature of the double-line sign. (orig.) [Deutsch] Das MR-tomographische Erscheinungsbild der Osteonekrose auf selektiven Fett- und Wasserbildern wurde analysiert, um das in der Literatur beschriebene Doppellinienzeichen naeher zu untersuchen. Hierfuer wurden sowohl die herkoemmlichen T1- und T2-gewichteten Spin-Echo-Sequenzen herangezogen, als auch frequenzselektive Bilder, die aufgrund chemischer Verschiebung gewonnen wurden (1,5 T). Es wurden die Untersuchungen von acht Patienten mit avaskulaerer Hueftkopfnekrose und von drei Patienten mit Knocheninfarkten retrospektiv ausgewertet. Acht von 11 Patienten zeigten ein Doppellinienzeichen auf den T2-gewichteten Bildern. Die Korrelation mit den selektiven Wasserbildern ergab, dass durch chemische Verschiebung bedingte Artefakte das Erscheinungsbild und den Ort der hyperintensen Linie beeinflussten. Die Bildgebung mit Hilfe der chemischen Verschiebung verbessert unser Verstaendnis der MRT-Charakteristika der Osteonekrose. (orig.)

  9. Quantitative analysis of gold and carbon nanoparticles in mammalian cells by flow cytometry light scattering

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Gang [Nanjing University, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences (China); Liu, Naicheng; Wang, Zhenheng [Nanjing University, Department of Orthopedics, Jinling Hospital, School of Medicine (China); Shi, Tongguo; Gan, Jingjing; Wang, Zhenzhen; Zhang, Junfeng, E-mail: jfzhang@nju.edu.cn [Nanjing University, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences (China)

    2017-02-15

    Nanoparticle-based applications for diagnostics and therapeutics have been extensively studied. These applications require a profound understanding of the fate of nanoparticles (NPs) in cellular environments. However, until now, few analytical methods are available and most of them rely on fluorescent properties or special elements of NPs; therefore, for NPs without observable optical properties or special elements, the existing methods are hardly applicable. In this study, we introduce a flow cytometry light scattering (FCLS)-based approach that quantifies in situ NPs accurately in mammalian cells. Continuous cells of heterogeneous human epithelial colorectal adenocarcinoma (Caco-2 cells), mouse peritoneal macrophages (MPM), and human adenocarcinomic alveolar basal epithelia (A549 cells) were cultured with NPs with certain concentrations and size. The intensity of the flow cytometric side scattered light, which indicates the quantity of NPs in the cells, was analyzed. The result shows an accurate size- and dose-dependent uptake of Au NPs (5, 30, 250 nm) in Caco-2 cells. The size- and dose- dependence of Au NPs (5, 30, 250 nm) and carbon NPs (50, 500 nm) in cells was validated by transmission electron microscope (TEM). This paper demonstrates the great potential of flow cytometry light scattering in the quantitative study of the size and dose effect on in situ metallic or non-metallic NPs in mammalian cells.

  10. Quantitative analysis of gold and carbon nanoparticles in mammalian cells by flow cytometry light scattering

    Science.gov (United States)

    Zhou, Gang; Liu, Naicheng; Wang, Zhenheng; Shi, Tongguo; Gan, Jingjing; Wang, Zhenzhen; Zhang, Junfeng

    2017-02-01

    Nanoparticle-based applications for diagnostics and therapeutics have been extensively studied. These applications require a profound understanding of the fate of nanoparticles (NPs) in cellular environments. However, until now, few analytical methods are available and most of them rely on fluorescent properties or special elements of NPs; therefore, for NPs without observable optical properties or special elements, the existing methods are hardly applicable. In this study, we introduce a flow cytometry light scattering (FCLS)-based approach that quantifies in situ NPs accurately in mammalian cells. Continuous cells of heterogeneous human epithelial colorectal adenocarcinoma (Caco-2 cells), mouse peritoneal macrophages (MPM), and human adenocarcinomic alveolar basal epithelia (A549 cells) were cultured with NPs with certain concentrations and size. The intensity of the flow cytometric side scattered light, which indicates the quantity of NPs in the cells, was analyzed. The result shows an accurate size- and dose-dependent uptake of Au NPs (5, 30, 250 nm) in Caco-2 cells. The size- and dose- dependence of Au NPs (5, 30, 250 nm) and carbon NPs (50, 500 nm) in cells was validated by transmission electron microscope (TEM). This paper demonstrates the great potential of flow cytometry light scattering in the quantitative study of the size and dose effect on in situ metallic or non-metallic NPs in mammalian cells.

  11. Image Gallery

    Science.gov (United States)

    ... R S T U V W X Y Z Image Gallery Share: The Image Gallery contains high-quality digital photographs available from ... Select a category below to view additional thumbnail images. Images are available for direct download in 2 ...

  12. Quantitative PET Imaging with Novel HER3-Targeted Peptides Selected by Phage Display to Predict Androgen-Independent Prostate Cancer Progression

    Science.gov (United States)

    2017-12-01

    Independent Prostate Cancer Progression PRINCIPAL INVESTIGATOR: Benjamin Larimer, PhD CONTRACTING ORGANIZATION: Massachusetts General Hospital Boston...TYPE Final 3. DATES COVERED 1 Aug 2016 – 19 August 2017 Selected by Phage Display to Predict Androgen-Independent Prostate Cancer Progression 5a...highly specific peptide that targets HER3 for prostate cancer imaging. The peptide was labeled with a PET imaging radionuclide and injected into mice

  13. Evaluation of selective arterial embolization effect by chitosan micro-hydrogels in hindlimb sarcoma rodent models using various imaging modalities

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Tai Kyoung; Kwon, Jeong Il; Na, Kyung Sook [Chonbuk National University Medical School and Hospital, Jeonju (Korea, Republic of); and others

    2015-09-15

    Embolization is mainly used to reduce the size of locally advanced tumors. In this study, selective arterial catheterization with chitosan micro-hydrogels (CMH) into the femoral artery was performed and the therapeutic effect was validated using different imaging methods. Male SD rats (n = 18, 6 weeks old) were randomly assigned into three groups: Group 1 as control, Group 2 without any ligation of distal femoral artery, and Group 3 with temporary ligation of the distal femoral artery. RR1022 sarcoma cell lines were inoculated into thigh muscle. After 1 week, CMH was injected into the proximal femoral artery. Different imaging modalities were performed during a 3-week follow-up. The tumor size was significantly (P < 0.001) decreased in both Group 2 and Group 3 (P < 0.001) after selective arterial embolization therapy. 18F-FDG-PET/CT revealed decreased intensity of 18F-FDG uptake in tumors. The accumulation status of 125I-CMH near the tumor was verified by gamma camera. Appropriate selective arterial embolization therapy with CMH was.

  14. Analysis of basophil activation by flow cytometry in pediatric house dust mite allergy.

    Science.gov (United States)

    González-Muñoz, Miguel; Villota, Julian; Moneo, Ignacio

    2008-06-01

    Detection of allergen-induced basophil activation by flow cytometry has been shown to be a useful tool for allergy diagnosis. The aim of this study was to assess the potential of this technique for the diagnosis of pediatric house dust mite allergy. Quantification of total and specific IgE and basophil activation test were performed to evaluate mite allergic (n = 24), atopic (n = 23), and non-allergic children (n = 9). Allergen-induced basophil activation was detected as a CD63-upregulation. Receiver operating characteristics (ROC) curve analysis was performed to calculate the optimal cut-off value of activated basophils discriminating mite allergic and non-allergic children. ROC curve analysis yielded a threshold value of 18% activated basophils when mite-sensitized and atopic children were studied [area under the curve (AUC) = 0.99, 95% confidence interval (CI) = 0.97-1.01, p 43 kU/l) values for Dermatophagoides pteronyssinus allergen. They also showed positive prick (wheal diameter >1.0 cm) and basophil activation (>87%) tests and high specific IgE (>100 kU/l) with shrimp allergen. Shrimp sensitization was demonstrated by high levels of Pen a 1-specific IgE (>100 kU/l). Cross-reactivity between mite and shrimp was confirmed by fluorescence enzyme immunoassay (FEIA-CAP) inhibition study in these two cases. This study demonstrated that the analysis of allergen-induced CD63 upregulation by flow cytometry is a reliable tool for diagnosis of mite allergy in pediatric patients, with sensitivity similar to routine diagnostic tests and a higher specificity. Furthermore, this method can provide additional information in case of disagreement between in vivo and in vitro test results.

  15. Measuring cell cycle progression kinetics with metabolic labeling and flow cytometry.

    Science.gov (United States)

    Fleisig, Helen; Wong, Judy

    2012-05-22

    Precise control of the initiation and subsequent progression through the various phases of the cell cycle are of paramount importance in proliferating cells. Cell cycle division is an integral part of growth and reproduction and deregulation of key cell cycle components have been implicated in the precipitating events of carcinogenesis. Molecular agents in anti-cancer therapies frequently target biological pathways responsible for the regulation and coordination of cell cycle division. Although cell cycle kinetics tend to vary according to cell type, the distribution of cells amongst the four stages of the cell cycle is rather consistent within a particular cell line due to the consistent pattern of mitogen and growth factor expression. Genotoxic events and other cellular stressors can result in a temporary block of cell cycle progression, resulting in arrest or a temporary pause in a particular cell cycle phase to allow for instigation of the appropriate response mechanism. The ability to experimentally observe the behavior of a cell population with reference to their cell cycle progression stage is an important advance in cell biology. Common procedures such as mitotic shake off, differential centrifugation or flow cytometry-based sorting are used to isolate cells at specific stages of the cell cycle. These fractionated, cell cycle phase-enriched populations are then subjected to experimental treatments. Yield, purity and viability of the separated fractions can often be compromised using these physical separation methods. As well, the time lapse between separation of the cell populations and the start of experimental treatment, whereby the fractionated cells can progress from the selected cell cycle stage, can pose significant challenges in the successful implementation and interpretation of these experiments. Other approaches to study cell cycle stages include the use of chemicals to synchronize cells. Treatment of cells with chemical inhibitors of key

  16. Effect of acetic acid on Saccharomyces carlsbergensis ATCC 6269 batch ethanol production monitored by flow cytometry.

    Science.gov (United States)

    Freitas, Cláudia; Neves, Elisabete; Reis, Alberto; Passarinho, Paula C; da Silva, Teresa Lopes

    2012-11-01

    Bioethanol produced from lignocellulosic materials has been considered a sustainable alternative fuel. Such type of raw materials have a huge potential, but their hydrolysis into mono-sugars releases toxic compounds such as weak acids, which affect the microorganisms' physiology, inhibiting the growth and ethanol production. Acetic acid (HAc) is the most abundant weak acid in the lignocellulosic materials hydrolysates. In order to understand the physiological changes of Saccharomyces carlsbergensis when fermenting in the presence of different acetic acid (HAc) concentrations, the yeast growth was monitored by multi-parameter flow cytometry at same time that the ethanol production was assessed. The membrane potential stain DiOC(6)(3) fluorescence intensity decreased as the HAc concentration increased, which was attributed to the plasmic membrane potential reduction as a result of the toxic effect of the HAc undissociated form. Nevertheless, the proportion of cells with permeabilized membrane did not increase with the HAc concentration increase. Fermentations ending at lower external pH and higher ethanol concentrations depicted the highest proportions of permeabilized cells and cells with increased reactive oxygen species levels. Flow cytometry allowed monitoring, near real time (at-line), the physiological states of the yeast during the fermentations. The information obtained can be used to optimize culture conditions to improve bioethanol production.

  17. Ploidy Levels among Species in the ‘Oxalis tuberosa Alliance’ as Inferred by Flow Cytometry

    OpenAIRE

    EMSHWILLER, EVE

    2002-01-01

    The ‘Oxalis tuberosa alliance’ is a group of Andean Oxalis species allied to the Andean tuber crop O. tuberosa Molina (Oxalidaceae), commonly known as ‘oca’. As part of a larger project studying the origins of polyploidy and domestication of cultivated oca, flow cytometry was used to survey DNA ploidy levels among Bolivian and Peruvian accessions of alliance members. In addition, this study provided a first assessment of C‐values in the alliance by estimating nuclear DNA contents of these acc...

  18. The evolution of phase holographic imaging from a research idea to publicly traded company

    Science.gov (United States)

    Egelberg, Peter

    2018-02-01

    Recognizing the value and unmet need for label-free kinetic cell analysis, Phase Holograhic Imaging defines its market segment as automated, easy to use and affordable time-lapse cytometry. The process of developing new technology, meeting customer expectations, sources of corporate funding and R&D adjustments prompted by field experience will be reviewed. Additionally, it is discussed how relevant biological information can be extracted from a sequence of quantitative phase images, with negligible user assistance and parameter tweaking, to simultaneously provide cell culture characteristics such as cell growth rate, viability, division rate, mitosis duration, phagocytosis rate, migration, motility and cell-cell adherence without requiring any artificial cell manipulation.

  19. Autologous Pure Platelet-Rich Plasma Dermal Injections for Facial Skin Rejuvenation: Clinical, Instrumental, and Flow Cytometry Assessment.

    Science.gov (United States)

    Cameli, Norma; Mariano, Maria; Cordone, Iole; Abril, Elva; Masi, Serena; Foddai, Maria Laura

    2017-06-01

    Platelet-rich plasma (PRP) is an emerging treatment in dermatology recently proposed for skin rejuvenation. To evaluate the efficacy and safety of autologous pure PRP dermal injections on facial skin rejuvenation, investigating the cellularity of PRP samples. Twelve patients underwent 3 sessions of PRP injection at 1-month intervals. The clinical and instrumental outcomes were evaluated before (T0) and 1 month (T1) after the end of treatment by means of transepidermal water loss, corneometry, Cutometer, Visioscan, and Visioface. A flow cytometry characterization on PRP and peripheral blood (PB) samples was performed. Clinical and patient evaluation showed improvement of skin texture. Skin gross elasticity, skin smoothness parameters, skin barrier function, and capacitance were significantly improved. No difference between PRP and PB lymphocyte immunological asset was observed. A leukocyte population (mainly CD3) and neutrophils depletion were documented in all the PRP samples. This instrumental study demonstrated that PRP poor in leukocytes can provide objective improvements in skin biostimulation. Flow cytometry showed no variability among the PRP samples using a reproducible separation system and a low content in proinflammatory cells. Although a pilot study, it may be helpful for future investigations on PRP cellularity.

  20. Automated imaging of cellular spheroids with selective plane illumination microscopy on a chip (Conference Presentation)

    Science.gov (United States)

    Paiè, Petra; Bassi, Andrea; Bragheri, Francesca; Osellame, Roberto

    2017-02-01

    Selective plane illumination microscopy (SPIM) is an optical sectioning technique that allows imaging of biological samples at high spatio-temporal resolution. Standard SPIM devices require dedicated set-ups, complex sample preparation and accurate system alignment, thus limiting the automation of the technique, its accessibility and throughput. We present a millimeter-scaled optofluidic device that incorporates selective plane illumination and fully automatic sample delivery and scanning. To this end an integrated cylindrical lens and a three-dimensional fluidic network were fabricated by femtosecond laser micromachining into a single glass chip. This device can upgrade any standard fluorescence microscope to a SPIM system. We used SPIM on a CHIP to automatically scan biological samples under a conventional microscope, without the need of any motorized stage: tissue spheroids expressing fluorescent proteins were flowed in the microchannel at constant speed and their sections were acquired while passing through the light sheet. We demonstrate high-throughput imaging of the entire sample volume (with a rate of 30 samples/min), segmentation and quantification in thick (100-300 μm diameter) cellular spheroids. This optofluidic device gives access to SPIM analyses to non-expert end-users, opening the way to automatic and fast screening of a high number of samples at subcellular resolution.