Sample records for content image cytometry
-
Image cytometry: nuclear and chromosomal DNA quantification.
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.
-
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.
-
Multi-channel imaging cytometry with a single detector
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.
-
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.
-
Microfluidic Imaging Flow Cytometry by Asymmetric-detection Time-stretch Optical Microscopy (ATOM).
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
-
Optofluidic fluorescent imaging cytometry on a cell phone.
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
-
Analysis of Cellular DNA Content by Flow Cytometry.
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.
-
Hyperchromatic laser scanning cytometry
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.
-
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.
-
BlobFinder, a tool for fluorescence microscopy image cytometry
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 ...
-
Imaging cytometry in a plastic ultra-mobile system
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.
-
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.
-
An open-source solution for advanced imaging flow cytometry data analysis using machine learning.
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
-
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.
-
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.
-
Wide-field fluorescent microscopy and fluorescent imaging flow cytometry on a cell-phone.
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.
-
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)
-
Flow cytometry in diagnostic cytology.
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.
-
CytometryML and other data formats
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
-
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
-
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....
-
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.
-
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.
-
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.
-
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
-
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.
-
Nuclear DNA content of the hybrid plant pathogen Phytophthora andina determined by flow cytometry.
Wang, Jianan; Presser, Jackson W; Goss, Erica M
2016-09-01
Phytophthora andina is a heterothallic plant pathogen of Andean solanaceous hosts and is an interspecific hybrid of P. infestans and an unknown Phytophthora species. The objective of this study was to estimate the nuclear DNA content of isolates in three clonal lineages of P. andina relative to P. infestans Twelve isolates of P. andina and six isolates of P. infestans were measured for nuclear DNA content by propidium iodide-stained flow cytometry. We found that the DNA content of P. andina was similar but slightly smaller, on average, than that of our sample of P. infestans isolates. This is consistent with P. andina being a homoploid hybrid rather than allopolyploid hybrid. Nuclear DNA content was more variable among a smaller sample of P. infestans isolates, including a putative triploid isolate from Mexico, but small differences in nuclear DNA content were also observed among P. andina isolates. Both species appear to be able to tolerate significant variation in genome size. © 2016 by The Mycological Society of America.
-
Microfluidic devices and methods for integrated flow cytometry
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.
-
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.
-
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.
-
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
-
Cutting-edge analysis of extracellular microparticles using ImageStream(X) imaging flow cytometry.
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.
-
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.
-
CytometryML: a markup language for analytical cytology
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.
-
FlowCam: Quantification and Classification of Phytoplankton by Imaging Flow Cytometry.
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.
-
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
-
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...
-
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.
-
Directory of Open Access Journals (Sweden)
Upatham Meesawat
2008-05-01
Full Text Available Nuclear DNA content for the adult plants grown in a greenhouse and in vitro young plantlets of the pigeon orchid (Dendrobium crumenatum Sw. was analyzed using flow cytometry. The resulting 2C DNA values ranged from 2.30±0.14 pgto 2.43±0.06 pg. However, nuclear DNA ploidy levels of long-term in vitro plantlets were found to be triploid and tetraploid.These ploidy levels were confirmed by chromosome counting. Tetraploid individuals (2n = 4x = 76 had approximately two times DNA content than diploid (2n = 2x = 38 individuals. This variation may be due to prolonged cultivation and thepresence of exogenous plant growth regulators.
-
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.
-
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.
-
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.
-
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.
-
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.
-
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}.
-
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
-
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.
-
ggCyto: Next Generation Open-Source Visualization Software for Cytometry.
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/.
-
Image Retargeting by Content-Aware Synthesis
Dong, Weiming; Wu, Fuzhang; Kong, Yan; Mei, Xing; Lee, Tong-Yee; Zhang, Xiaopeng
2014-01-01
Real-world images usually contain vivid contents and rich textural details, which will complicate the manipulation on them. In this paper, we design a new framework based on content-aware synthesis to enhance content-aware image retargeting. By detecting the textural regions in an image, the textural image content can be synthesized rather than simply distorted or cropped. This method enables the manipulation of textural & non-textural regions with different strategy since they have different...
-
An imaging flow cytometry method to assess ricin trafficking in A549 human lung epithelial cells.
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.
-
CytometryML with DICOM and FCS
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.
-
Dictionary-enhanced imaging cytometry
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.
-
Metal-Containing Polystyrene Beads as Standards for Mass Cytometry.
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.
-
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.
-
Analysis of the Budding Yeast Cell Cycle by Flow Cytometry.
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.
-
CytometryML: a data standard which has been designed to interface with other standards
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.
-
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.
-
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: ...
-
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
-
High content analysis of differentiation and cell death in human adipocytes.
Doan-Xuan, Quang Minh; Sarvari, Anitta K; Fischer-Posovszky, Pamela; Wabitsch, Martin; Balajthy, Zoltan; Fesus, Laszlo; Bacso, Zsolt
2013-10-01
Understanding adipocyte biology and its homeostasis is in the focus of current obesity research. We aimed to introduce a high-content analysis procedure for directly visualizing and quantifying adipogenesis and adipoapoptosis by laser scanning cytometry (LSC) in a large population of cell. Slide-based image cytometry and image processing algorithms were used and optimized for high-throughput analysis of differentiating cells and apoptotic processes in cell culture at high confluence. Both preadipocytes and adipocytes were simultaneously scrutinized for lipid accumulation, texture properties, nuclear condensation, and DNA fragmentation. Adipocyte commitment was found after incubation in adipogenic medium for 3 days identified by lipid droplet formation and increased light absorption, while terminal differentiation of adipocytes occurred throughout day 9-14 with characteristic nuclear shrinkage, eccentric nuclei localization, chromatin condensation, and massive lipid deposition. Preadipocytes were shown to be more prone to tumor necrosis factor alpha (TNFα)-induced apoptosis compared to mature adipocytes. Importantly, spontaneous DNA fragmentation was observed at early stage when adipocyte commitment occurs. This DNA damage was independent from either spontaneous or induced apoptosis and probably was part of the differentiation program. © 2013 International Society for Advancement of Cytometry. Copyright © 2013 International Society for Advancement of Cytometry.
-
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
-
CMOS based image cytometry for detection of phytoplankton in ballast water.
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.
-
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.
-
Application of image flow cytometry for the characterization of red blood cell morphology
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.
-
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.
-
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
-
Opto-fluidics based microscopy and flow cytometry on a cell phone for blood analysis.
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
-
Content-Based Image Retrial Based on Hadoop
Directory of Open Access Journals (Sweden)
DongSheng Yin
2013-01-01
Full Text Available Generally, time complexity of algorithms for content-based image retrial is extremely high. In order to retrieve images on large-scale databases efficiently, a new way for retrieving based on Hadoop distributed framework is proposed. Firstly, a database of images features is built by using Speeded Up Robust Features algorithm and Locality-Sensitive Hashing and then perform the search on Hadoop platform in a parallel way specially designed. Considerable experimental results show that it is able to retrieve images based on content on large-scale cluster and image sets effectively.
-
Yang, Xi; Xiao, Xuan; Wu, Wenyan; Shen, Xuemin; Zhou, Zengtong; Liu, Wei; Shi, Linjun
2017-09-01
To quantitatively examine the DNA content and nuclear morphometric status of oral leukoplakia (OL) and investigate its association with the degree of dysplasia in a cytologic study. Oral cytobrush biopsy was carried out to obtain exfoliative epithelial cells from lesions before scalpel biopsy at the same location in a blinded series of 70 patients with OL. Analysis of nuclear morphometry and DNA content status using image cytometry was performed with oral smears stained with the Feulgen-thionin method. Nuclear morphometric analysis revealed significant differences in DNA content amount, DNA index, nuclear area, nuclear radius, nuclear intensity, sphericity, entropy, and fractal dimension (all P content analysis identified 34 patients with OL (48.6%) with DNA content abnormality. Nonhomogeneous lesion (P = .018) and high-grade dysplasia (P = .008) were significantly associated with abnormal DNA content. Importantly, the positive correlation between the degree of oral dysplasia and DNA content status was significant (P = .004, correlation coefficient = 0.342). Cytology analysis of DNA content and nuclear morphometric status using image cytometry may support their use as a screening and monitoring tool for OL progression. Copyright © 2017 Elsevier Inc. All rights reserved.
-
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.
-
Application of content-based image compression to telepathology
Varga, Margaret J.; Ducksbury, Paul G.; Callagy, Grace
2002-05-01
Telepathology is a means of practicing pathology at a distance, viewing images on a computer display rather than directly through a microscope. Without compression, images take too long to transmit to a remote location and are very expensive to store for future examination. However, to date the use of compressed images in pathology remains controversial. This is because commercial image compression algorithms such as JPEG achieve data compression without knowledge of the diagnostic content. Often images are lossily compressed at the expense of corrupting informative content. None of the currently available lossy compression techniques are concerned with what information has been preserved and what data has been discarded. Their sole objective is to compress and transmit the images as fast as possible. By contrast, this paper presents a novel image compression technique, which exploits knowledge of the slide diagnostic content. This 'content based' approach combines visually lossless and lossy compression techniques, judiciously applying each in the appropriate context across an image so as to maintain 'diagnostic' information while still maximising the possible compression. Standard compression algorithms, e.g. wavelets, can still be used, but their use in a context sensitive manner can offer high compression ratios and preservation of diagnostically important information. When compared with lossless compression the novel content-based approach can potentially provide the same degree of information with a smaller amount of data. When compared with lossy compression it can provide more information for a given amount of compression. The precise gain in the compression performance depends on the application (e.g. database archive or second opinion consultation) and the diagnostic content of the images.
-
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.
-
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.
-
Content Based Retrieval System for Magnetic Resonance Images
International Nuclear Information System (INIS)
Trojachanets, Katarina
2010-01-01
The amount of medical images is continuously increasing as a consequence of the constant growth and development of techniques for digital image acquisition. Manual annotation and description of each image is impractical, expensive and time consuming approach. Moreover, it is an imprecise and insufficient way for describing all information stored in medical images. This induces the necessity for developing efficient image storage, annotation and retrieval systems. Content based image retrieval (CBIR) emerges as an efficient approach for digital image retrieval from large databases. It includes two phases. In the first phase, the visual content of the image is analyzed and the feature extraction process is performed. An appropriate descriptor, namely, feature vector is then associated with each image. These descriptors are used in the second phase, i.e. the retrieval process. With the aim to improve the efficiency and precision of the content based image retrieval systems, feature extraction and automatic image annotation techniques are subject of continuous researches and development. Including the classification techniques in the retrieval process enables automatic image annotation in an existing CBIR system. It contributes to more efficient and easier image organization in the system.Applying content based retrieval in the field of magnetic resonance is a big challenge. Magnetic resonance imaging is an image based diagnostic technique which is widely used in medical environment. According to this, the number of magnetic resonance images is enormously growing. Magnetic resonance images provide plentiful medical information, high resolution and specific nature. Thus, the capability of CBIR systems for image retrieval from large database is of great importance for efficient analysis of this kind of images. The aim of this thesis is to propose content based retrieval system architecture for magnetic resonance images. To provide the system efficiency, feature
-
A Sensitive Measurement for Estimating Impressions of Image-Contents
Sato, Mie; Matouge, Shingo; Mori, Toshifumi; Suzuki, Noboru; Kasuga, Masao
We have investigated Kansei Content that appeals maker's intention to viewer's kansei. An SD method is a very good way to evaluate subjective impression of image-contents. However, because the SD method is performed after subjects view the image-contents, it is difficult to examine impression of detailed scenes of the image-contents in real time. To measure viewer's impression of the image-contents in real time, we have developed a Taikan sensor. With the Taikan sensor, we investigate relations among the image-contents, the grip strength and the body temperature. We also explore the interface of the Taikan sensor to use it easily. In our experiment, a horror movie is used that largely affects emotion of the subjects. Our results show that there is a possibility that the grip strength increases when the subjects view a strained scene and that it is easy to use the Taikan sensor without its circle base that is originally installed.
-
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
-
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.
-
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.
-
Content Progressive Coding of Limited Bits/pixel Images
DEFF Research Database (Denmark)
Jensen, Ole Riis; Forchhammer, Søren
1999-01-01
A new lossless context based method for content progressive coding of limited bits/pixel images is proposed. Progressive coding is achieved by separating the image into contelnt layers. Digital maps are compressed up to 3 times better than GIF.......A new lossless context based method for content progressive coding of limited bits/pixel images is proposed. Progressive coding is achieved by separating the image into contelnt layers. Digital maps are compressed up to 3 times better than GIF....
-
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.
-
Content-based Image Hiding Method for Secure Network Biometric Verification
Directory of Open Access Journals (Sweden)
Xiangjiu Che
2011-08-01
Full Text Available For secure biometric verification, most existing methods embed biometric information directly into the cover image, but content correlation analysis between the biometric image and the cover image is often ignored. In this paper, we propose a novel biometric image hiding approach based on the content correlation analysis to protect the network-based transmitted image. By using principal component analysis (PCA, the content correlation between the biometric image and the cover image is firstly analyzed. Then based on particle swarm optimization (PSO algorithm, some regions of the cover image are selected to represent the biometric image, in which the cover image can carry partial content of the biometric image. As a result of the correlation analysis, the unrepresented part of the biometric image is embedded into the cover image by using the discrete wavelet transform (DWT. Combined with human visual system (HVS model, this approach makes the hiding result perceptually invisible. The extensive experimental results demonstrate that the proposed hiding approach is robust against some common frequency and geometric attacks; it also provides an effective protection for the secure biometric verification.
-
SCENERY: a web application for (causal) network reconstruction from cytometry data
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/.
-
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.
-
Information content of poisson images
International Nuclear Information System (INIS)
Cederlund, J.
1979-04-01
One major problem when producing images with the aid of Poisson distributed quanta is how best to compromise between spatial and contrast resolution. Increasing the number of image elements improves spatial resolution, but at the cost of fewer quanta per image element, which reduces contrast resolution. Information theory arguments are used to analyse this problem. It is argued that information capacity is a useful concept to describe an important property of the imaging device, but that in order to compute the information content of an image produced by this device some statistical properties (such as the a priori probability of the densities) of the object to be depicted must be taken into account. If these statistical properties are not known one cannot make a correct choice between spatial and contrast resolution. (author)
-
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.
-
Content Based Medical Image Retrieval for Histopathological, CT and MRI Images
Directory of Open Access Journals (Sweden)
Swarnambiga AYYACHAMY
2013-09-01
Full Text Available A content based approach is followed for medical images. The purpose of this study is to access the stability of these methods for medical image retrieval. The methods used in color based retrieval for histopathological images are color co-occurrence matrix (CCM and histogram with meta features. For texture based retrieval GLCM (gray level co-occurrence matrix and local binary pattern (LBP were used. For shape based retrieval canny edge detection and otsu‘s method with multivariable threshold were used. Texture and shape based retrieval were implemented using MRI (magnetic resonance images. The most remarkable characteristics of the article are its content based approach for each medical imaging modality. Our efforts were focused on the initial visual search. From our experiment, histogram with meta features in color based retrieval for histopathological images shows a precision of 60 % and recall of 30 %. Whereas GLCM in texture based retrieval for MRI images shows a precision of 70 % and recall of 20 %. Shape based retrieval for MRI images shows a precision of 50% and recall of 25 %. The retrieval results shows that this simple approach is successful.
-
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.)
-
Content-based image retrieval with ontological ranking
Tsai, Shen-Fu; Tsai, Min-Hsuan; Huang, Thomas S.
2010-02-01
Images are a much more powerful medium of expression than text, as the adage says: "One picture is worth a thousand words." It is because compared with text consisting of an array of words, an image has more degrees of freedom and therefore a more complicated structure. However, the less limited structure of images presents researchers in the computer vision community a tough task of teaching machines to understand and organize images, especially when a limit number of learning examples and background knowledge are given. The advance of internet and web technology in the past decade has changed the way human gain knowledge. People, hence, can exchange knowledge with others by discussing and contributing information on the web. As a result, the web pages in the internet have become a living and growing source of information. One is therefore tempted to wonder whether machines can learn from the web knowledge base as well. Indeed, it is possible to make computer learn from the internet and provide human with more meaningful knowledge. In this work, we explore this novel possibility on image understanding applied to semantic image search. We exploit web resources to obtain links from images to keywords and a semantic ontology constituting human's general knowledge. The former maps visual content to related text in contrast to the traditional way of associating images with surrounding text; the latter provides relations between concepts for machines to understand to what extent and in what sense an image is close to the image search query. With the aid of these two tools, the resulting image search system is thus content-based and moreover, organized. The returned images are ranked and organized such that semantically similar images are grouped together and given a rank based on the semantic closeness to the input query. The novelty of the system is twofold: first, images are retrieved not only based on text cues but their actual contents as well; second, the grouping
-
Information management for high content live cell imaging
Directory of Open Access Journals (Sweden)
White Michael RH
2009-07-01
Full Text Available Abstract Background High content live cell imaging experiments are able to track the cellular localisation of labelled proteins in multiple live cells over a time course. Experiments using high content live cell imaging will generate multiple large datasets that are often stored in an ad-hoc manner. This hinders identification of previously gathered data that may be relevant to current analyses. Whilst solutions exist for managing image data, they are primarily concerned with storage and retrieval of the images themselves and not the data derived from the images. There is therefore a requirement for an information management solution that facilitates the indexing of experimental metadata and results of high content live cell imaging experiments. Results We have designed and implemented a data model and information management solution for the data gathered through high content live cell imaging experiments. Many of the experiments to be stored measure the translocation of fluorescently labelled proteins from cytoplasm to nucleus in individual cells. The functionality of this database has been enhanced by the addition of an algorithm that automatically annotates results of these experiments with the timings of translocations and periods of any oscillatory translocations as they are uploaded to the repository. Testing has shown the algorithm to perform well with a variety of previously unseen data. Conclusion Our repository is a fully functional example of how high throughput imaging data may be effectively indexed and managed to address the requirements of end users. By implementing the automated analysis of experimental results, we have provided a clear impetus for individuals to ensure that their data forms part of that which is stored in the repository. Although focused on imaging, the solution provided is sufficiently generic to be applied to other functional proteomics and genomics experiments. The software is available from: fhttp://code.google.com/p/livecellim/
-
Content dependent selection of image enhancement parameters for mobile displays
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.
-
Investigating the link between radiologists’ gaze, diagnostic decision, and image content
Tourassi, Georgia; Voisin, Sophie; Paquit, Vincent; Krupinski, Elizabeth
2013-01-01
Objective To investigate machine learning for linking image content, human perception, cognition, and error in the diagnostic interpretation of mammograms. Methods Gaze data and diagnostic decisions were collected from three breast imaging radiologists and three radiology residents who reviewed 20 screening mammograms while wearing a head-mounted eye-tracker. Image analysis was performed in mammographic regions that attracted radiologists’ attention and in all abnormal regions. Machine learning algorithms were investigated to develop predictive models that link: (i) image content with gaze, (ii) image content and gaze with cognition, and (iii) image content, gaze, and cognition with diagnostic error. Both group-based and individualized models were explored. Results By pooling the data from all readers, machine learning produced highly accurate predictive models linking image content, gaze, and cognition. Potential linking of those with diagnostic error was also supported to some extent. Merging readers’ gaze metrics and cognitive opinions with computer-extracted image features identified 59% of the readers’ diagnostic errors while confirming 97.3% of their correct diagnoses. The readers’ individual perceptual and cognitive behaviors could be adequately predicted by modeling the behavior of others. However, personalized tuning was in many cases beneficial for capturing more accurately individual behavior. Conclusions There is clearly an interaction between radiologists’ gaze, diagnostic decision, and image content which can be modeled with machine learning algorithms. PMID:23788627
-
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.
-
The Use of QBIC Content-Based Image Retrieval System
Directory of Open Access Journals (Sweden)
Ching-Yi Wu
2004-03-01
Full Text Available The fast increase in digital images has caught increasing attention on the development of image retrieval technologies. Content-based image retrieval (CBIR has become an important approach in retrieving image data from a large collection. This article reports our results on the use and users study of a CBIR system. Thirty-eight students majored in art and design were invited to use the IBM’s OBIC (Query by Image Content system through the Internet. Data from their information needs, behaviors, and retrieval strategies were collected through an in-depth interview, observation, and self-described think-aloud process. Important conclusions are:（1）There are four types of information needs for image data: implicit, inspirational, ever-changing, and purposive. The types of needs may change during the retrieval process. （2）CBIR is suitable for the example-type query, text retrieval is suitable for the scenario-type query, and image browsing is suitable for the symbolic query. （3）Different from text retrieval, detailed description of the query condition may lead to retrieval failure more easily. （4）CBIR is suitable for the domain-specific image collection, not for the images on the Word-Wide Web.[Article content in Chinese
-
Retinal image quality assessment based on image clarity and content
Abdel-Hamid, Lamiaa; El-Rafei, Ahmed; El-Ramly, Salwa; Michelson, Georg; Hornegger, Joachim
2016-09-01
Retinal image quality assessment (RIQA) is an essential step in automated screening systems to avoid misdiagnosis caused by processing poor quality retinal images. A no-reference transform-based RIQA algorithm is introduced that assesses images based on five clarity and content quality issues: sharpness, illumination, homogeneity, field definition, and content. Transform-based RIQA algorithms have the advantage of considering retinal structures while being computationally inexpensive. Wavelet-based features are proposed to evaluate the sharpness and overall illumination of the images. A retinal saturation channel is designed and used along with wavelet-based features for homogeneity assessment. The presented sharpness and illumination features are utilized to assure adequate field definition, whereas color information is used to exclude nonretinal images. Several publicly available datasets of varying quality grades are utilized to evaluate the feature sets resulting in area under the receiver operating characteristic curve above 0.99 for each of the individual feature sets. The overall quality is assessed by a classifier that uses the collective features as an input vector. The classification results show superior performance of the algorithm in comparison to other methods from literature. Moreover, the algorithm addresses efficiently and comprehensively various quality issues and is suitable for automatic screening systems.
-
An introduction to mass cytometry: fundamentals and applications.
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.
-
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
-
One-dimensional acoustic standing waves in rectangular channels for flow cytometry.
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.
-
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...
-
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....
-
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.
-
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)
-
A content analysis of thinspiration images and text posts on Tumblr.
Wick, Madeline R; Harriger, Jennifer A
2018-03-01
Thinspiration is content advocating extreme weight loss by means of images and/or text posts. While past content analyses have examined thinspiration content on social media and other websites, no research to date has examined thinspiration content on Tumblr. Over the course of a week, 222 images and text posts were collected after entering the keyword 'thinspiration' into the Tumblr search bar. These images were then rated on a variety of characteristics. The majority of thinspiration images included a thin woman adhering to culturally based beauty, often posing in a manner that accentuated her thinness or sexuality. The most common themes for thinspiration text posts included dieting/restraint, weight loss, food guilt, and body guilt. The thinspiration content on Tumblr appears to be consistent with that on other mediums. Future research should utilize experimental methods to examine the potential effects of consuming thinspiration content on Tumblr. Copyright © 2017 Elsevier Ltd. All rights reserved.
-
Investigating the Link Between Radiologists Gaze, Diagnostic Decision, and Image Content
Energy Technology Data Exchange (ETDEWEB)
Tourassi, Georgia [ORNL; Voisin, Sophie [ORNL; Paquit, Vincent C [ORNL; Krupinski, Elizabeth [University of Arizona
2013-01-01
Objective: To investigate machine learning for linking image content, human perception, cognition, and error in the diagnostic interpretation of mammograms. Methods: Gaze data and diagnostic decisions were collected from six radiologists who reviewed 20 screening mammograms while wearing a head-mounted eye-tracker. Texture analysis was performed in mammographic regions that attracted radiologists attention and in all abnormal regions. Machine learning algorithms were investigated to develop predictive models that link: (i) image content with gaze, (ii) image content and gaze with cognition, and (iii) image content, gaze, and cognition with diagnostic error. Both group-based and individualized models were explored. Results: By pooling the data from all radiologists machine learning produced highly accurate predictive models linking image content, gaze, cognition, and error. Merging radiologists gaze metrics and cognitive opinions with computer-extracted image features identified 59% of the radiologists diagnostic errors while confirming 96.2% of their correct diagnoses. The radiologists individual errors could be adequately predicted by modeling the behavior of their peers. However, personalized tuning appears to be beneficial in many cases to capture more accurately individual behavior. Conclusions: Machine learning algorithms combining image features with radiologists gaze data and diagnostic decisions can be effectively developed to recognize cognitive and perceptual errors associated with the diagnostic interpretation of mammograms.
-
Content-Based Image Retrieval Based on Electromagnetism-Like Mechanism
Directory of Open Access Journals (Sweden)
Hamid A. Jalab
2013-01-01
Full Text Available Recently, many researchers in the field of automatic content-based image retrieval have devoted a remarkable amount of research looking for methods to retrieve the best relevant images to the query image. This paper presents a novel algorithm for increasing the precision in content-based image retrieval based on electromagnetism optimization technique. The electromagnetism optimization is a nature-inspired technique that follows the collective attraction-repulsion mechanism by considering each image as an electrical charge. The algorithm is composed of two phases: fitness function measurement and electromagnetism optimization technique. It is implemented on a database with 8,000 images spread across 80 classes with 100 images in each class. Eight thousand queries are fired on the database, and the overall average precision is computed. Experimental results of the proposed approach have shown significant improvement in the retrieval performance in regard to precision.
-
Bread Water Content Measurement Based on Hyperspectral Imaging
DEFF Research Database (Denmark)
Liu, Zhi; Møller, Flemming
2011-01-01
Water content is one of the most important properties of the bread for tasting assesment or store monitoring. Traditional bread water content measurement methods mostly are processed manually, which is destructive and time consuming. This paper proposes an automated water content measurement...... for bread quality based on near-infrared hyperspectral imaging against the conventional manual loss-in-weight method. For this purpose, the hyperspectral components unmixing technology is used for measuring the water content quantitatively. And the definition on bread water content index is presented...
-
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
-
iScreen: Image-Based High-Content RNAi Screening Analysis Tools.
Zhong, Rui; Dong, Xiaonan; Levine, Beth; Xie, Yang; Xiao, Guanghua
2015-09-01
High-throughput RNA interference (RNAi) screening has opened up a path to investigating functional genomics in a genome-wide pattern. However, such studies are often restricted to assays that have a single readout format. Recently, advanced image technologies have been coupled with high-throughput RNAi screening to develop high-content screening, in which one or more cell image(s), instead of a single readout, were generated from each well. This image-based high-content screening technology has led to genome-wide functional annotation in a wider spectrum of biological research studies, as well as in drug and target discovery, so that complex cellular phenotypes can be measured in a multiparametric format. Despite these advances, data analysis and visualization tools are still largely lacking for these types of experiments. Therefore, we developed iScreen (image-Based High-content RNAi Screening Analysis Tool), an R package for the statistical modeling and visualization of image-based high-content RNAi screening. Two case studies were used to demonstrate the capability and efficiency of the iScreen package. iScreen is available for download on CRAN (http://cran.cnr.berkeley.edu/web/packages/iScreen/index.html). The user manual is also available as a supplementary document. © 2014 Society for Laboratory Automation and Screening.
-
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...
-
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.
-
Immuno flow cytometry in marine phytoplankton research
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.
-
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...
-
Interfacing Lab-on-a-Chip Embryo Technology with High-Definition Imaging Cytometry.
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.
-
Automation in high-content flow cytometry screening.
Naumann, U; Wand, M P
2009-09-01
High-content flow cytometric screening (FC-HCS) is a 21st Century technology that combines robotic fluid handling, flow cytometric instrumentation, and bioinformatics software, so that relatively large numbers of flow cytometric samples can be processed and analysed in a short period of time. We revisit a recent application of FC-HCS to the problem of cellular signature definition for acute graft-versus-host-disease. Our focus is on automation of the data processing steps using recent advances in statistical methodology. We demonstrate that effective results, on par with those obtained via manual processing, can be achieved using our automatic techniques. Such automation of FC-HCS has the potential to drastically improve diagnosis and biomarker identification.
-
DNA content in radiation-associated thyroid cancer
International Nuclear Information System (INIS)
Komorowski, R.A.; Deaconson, T.F.; Vetsch, R.; Cerletty, J.M.; Wilson, S.D.
1988-01-01
DNA content has been reported to be of prognostic significance in differentiated thyroid carcinoma. Since malignant tumors with irradiation as an initiator often contain DNA aberrations, the DNA content of well-differentiated thyroid carcinoma in patients with a prior history of low-dose head and neck irradiation was determined and compared with similar nonradiation-associated lesions. The DNA content of thyroid cancers from 53 patients was determined with use of flow cytometry. Sixteen radiation-associated thyroid carcinomas (11 papillary, 3 follicular, and 2 medullary) all were diploid. In a group of 37 nonradiation-associated tumors, 10 were aneuploid (10 of 29 papillary carcinomas and 0 of 2 follicular or 6 medullary carcinomas). This difference in DNA content is significant (p less than 0.02, Fisher's exact test). These findings were unexpected and suggest that if the initiating irradiation causes a DNA aberration, this aberration is not reflected in DNA content as measured by means of flow cytometry
-
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
-
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
-
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
-
A picture tells a thousand words: A content analysis of concussion-related images online.
Ahmed, Osman H; Lee, Hopin; Struik, Laura L
2016-09-01
Recently image-sharing social media platforms have become a popular medium for sharing health-related images and associated information. However within the field of sports medicine, and more specifically sports related concussion, the content of images and meta-data shared through these popular platforms have not been investigated. The aim of this study was to analyse the content of concussion-related images and its accompanying meta-data on image-sharing social media platforms. We retrieved 300 images from Pinterest, Instagram and Flickr by using a standardised search strategy. All images were screened and duplicate images were removed. We excluded images if they were: non-static images; illustrations; animations; or screenshots. The content and characteristics of each image was evaluated using a customised coding scheme to determine major content themes, and images were referenced to the current international concussion management guidelines. From 300 potentially relevant images, 176 images were included for analysis; 70 from Pinterest, 63 from Flickr, and 43 from Instagram. Most images were of another person or a scene (64%), with the primary content depicting injured individuals (39%). The primary purposes of the images were to share a concussion-related incident (33%) and to dispense education (19%). For those images where it could be evaluated, the majority (91%) were found to reflect the Sports Concussion Assessment Tool 3 (SCAT3) guidelines. The ability to rapidly disseminate rich information though photos, images, and infographics to a wide-reaching audience suggests that image-sharing social media platforms could be used as an effective communication tool for sports concussion. Public health strategies could direct educative content to targeted populations via the use of image-sharing platforms. Further research is required to understand how image-sharing platforms can be used to effectively relay evidence-based information to patients and sports medicine
-
A Novel Technique for Shape Feature Extraction Using Content Based Image Retrieval
Directory of Open Access Journals (Sweden)
Dhanoa Jaspreet Singh
2016-01-01
Full Text Available With the advent of technology and multimedia information, digital images are increasing very quickly. Various techniques are being developed to retrieve/search digital information or data contained in the image. Traditional Text Based Image Retrieval System is not plentiful. Since it is time consuming as it require manual image annotation. Also, the image annotation differs with different peoples. An alternate to this is Content Based Image Retrieval (CBIR system. It retrieves/search for image using its contents rather the text, keywords etc. A lot of exploration has been compassed in the range of Content Based Image Retrieval (CBIR with various feature extraction techniques. Shape is a significant image feature as it reflects the human perception. Moreover, Shape is quite simple to use by the user to define object in an image as compared to other features such as Color, texture etc. Over and above, if applied alone, no descriptor will give fruitful results. Further, by combining it with an improved classifier, one can use the positive features of both the descriptor and classifier. So, a tryout will be made to establish an algorithm for accurate feature (Shape extraction in Content Based Image Retrieval (CBIR. The main objectives of this project are: (a To propose an algorithm for shape feature extraction using CBIR, (b To evaluate the performance of proposed algorithm and (c To compare the proposed algorithm with state of art techniques.
-
Content-adaptive Image Enhancement, Based on Sky and Grass Segmentation
Zafarifar, B.; With, de P.H.N.
2009-01-01
Current TV image enhancement functions employ globally controlled settings. A more flexible system can be achieved if the global control is extended to incorporate semantic-level image content information. In this paper, we present a system that extends existing TV image enhancement functions with
-
Content-based histopathology image retrieval using CometCloud.
Qi, Xin; Wang, Daihou; Rodero, Ivan; Diaz-Montes, Javier; Gensure, Rebekah H; Xing, Fuyong; Zhong, Hua; Goodell, Lauri; Parashar, Manish; Foran, David J; Yang, Lin
2014-08-26
The development of digital imaging technology is creating extraordinary levels of accuracy that provide support for improved reliability in different aspects of the image analysis, such as content-based image retrieval, image segmentation, and classification. This has dramatically increased the volume and rate at which data are generated. Together these facts make querying and sharing non-trivial and render centralized solutions unfeasible. Moreover, in many cases this data is often distributed and must be shared across multiple institutions requiring decentralized solutions. In this context, a new generation of data/information driven applications must be developed to take advantage of the national advanced cyber-infrastructure (ACI) which enable investigators to seamlessly and securely interact with information/data which is distributed across geographically disparate resources. This paper presents the development and evaluation of a novel content-based image retrieval (CBIR) framework. The methods were tested extensively using both peripheral blood smears and renal glomeruli specimens. The datasets and performance were evaluated by two pathologists to determine the concordance. The CBIR algorithms that were developed can reliably retrieve the candidate image patches exhibiting intensity and morphological characteristics that are most similar to a given query image. The methods described in this paper are able to reliably discriminate among subtle staining differences and spatial pattern distributions. By integrating a newly developed dual-similarity relevance feedback module into the CBIR framework, the CBIR results were improved substantially. By aggregating the computational power of high performance computing (HPC) and cloud resources, we demonstrated that the method can be successfully executed in minutes on the Cloud compared to weeks using standard computers. In this paper, we present a set of newly developed CBIR algorithms and validate them using two
-
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...
-
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.
-
INTEGRATION OF SPATIAL INFORMATION WITH COLOR FOR CONTENT RETRIEVAL OF REMOTE SENSING IMAGES
Directory of Open Access Journals (Sweden)
Bikesh Kumar Singh
2010-08-01
Full Text Available There is rapid increase in image databases of remote sensing images due to image satellites with high resolution, commercial applications of remote sensing & high available bandwidth in last few years. The problem of content-based image retrieval (CBIR of remotely sensed images presents a major challenge not only because of the surprisingly increasing volume of images acquired from a wide range of sensors but also because of the complexity of images themselves. In this paper, a software system for content-based retrieval of remote sensing images using RGB and HSV color spaces is presented. Further, we also compare our results with spatiogram based content retrieval which integrates spatial information along with color histogram. Experimental results show that the integration of spatial information in color improves the image analysis of remote sensing data. In general, retrievals in HSV color space showed better performance than in RGB color space.
-
Indexing, learning and content-based retrieval for special purpose image databases
M.J. Huiskes (Mark); E.J. Pauwels (Eric)
2005-01-01
textabstractThis chapter deals with content-based image retrieval in special purpose image databases. As image data is amassed ever more effortlessly, building efficient systems for searching and browsing of image databases becomes increasingly urgent. We provide an overview of the current
-
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....
-
Visual analytics for semantic queries of TerraSAR-X image content
Espinoza-Molina, Daniela; Alonso, Kevin; Datcu, Mihai
2015-10-01
With the continuous image product acquisition of satellite missions, the size of the image archives is considerably increasing every day as well as the variety and complexity of their content, surpassing the end-user capacity to analyse and exploit them. Advances in the image retrieval field have contributed to the development of tools for interactive exploration and extraction of the images from huge archives using different parameters like metadata, key-words, and basic image descriptors. Even though we count on more powerful tools for automated image retrieval and data analysis, we still face the problem of understanding and analyzing the results. Thus, a systematic computational analysis of these results is required in order to provide to the end-user a summary of the archive content in comprehensible terms. In this context, visual analytics combines automated analysis with interactive visualizations analysis techniques for an effective understanding, reasoning and decision making on the basis of very large and complex datasets. Moreover, currently several researches are focused on associating the content of the images with semantic definitions for describing the data in a format to be easily understood by the end-user. In this paper, we present our approach for computing visual analytics and semantically querying the TerraSAR-X archive. Our approach is mainly composed of four steps: 1) the generation of a data model that explains the information contained in a TerraSAR-X product. The model is formed by primitive descriptors and metadata entries, 2) the storage of this model in a database system, 3) the semantic definition of the image content based on machine learning algorithms and relevance feedback, and 4) querying the image archive using semantic descriptors as query parameters and computing the statistical analysis of the query results. The experimental results shows that with the help of visual analytics and semantic definitions we are able to explain
-
Content-based image retrieval: Color-selection exploited
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
-
Content-Based Image Retrieval: Color-selection exploited
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
-
Detecting content adaptive scaling of images for forensic applications
Fillion, Claude; Sharma, Gaurav
2010-01-01
Content-aware resizing methods have recently been developed, among which, seam-carving has achieved the most widespread use. Seam-carving's versatility enables deliberate object removal and benign image resizing, in which perceptually important content is preserved. Both types of modifications compromise the utility and validity of the modified images as evidence in legal and journalistic applications. It is therefore desirable that image forensic techniques detect the presence of seam-carving. In this paper we address detection of seam-carving for forensic purposes. As in other forensic applications, we pose the problem of seam-carving detection as the problem of classifying a test image in either of two classes: a) seam-carved or b) non-seam-carved. We adopt a pattern recognition approach in which a set of features is extracted from the test image and then a Support Vector Machine based classifier, trained over a set of images, is utilized to estimate which of the two classes the test image lies in. Based on our study of the seam-carving algorithm, we propose a set of intuitively motivated features for the detection of seam-carving. Our methodology for detection of seam-carving is then evaluated over a test database of images. We demonstrate that the proposed method provides the capability for detecting seam-carving with high accuracy. For images which have been reduced 30% by benign seam-carving, our method provides a classification accuracy of 91%.
-
An active, collaborative approach to learning skills in flow cytometry.
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.
-
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
-
Evaluation of moisture content distribution in wood by soft X-ray imaging
International Nuclear Information System (INIS)
Tanaka, T.; Avramidis, S.; Shida, S.
2009-01-01
A technique for nondestructive evaluation of moisture content distribution of Japanese cedar (sugi) during drying using a newly developed soft X-ray digital microscope was investigated. Radial, tangential, and cross-sectional samples measuring 100 x 100 x 10 mm were cut from green sugi wood. Each sample was dried in several steps in an oven and upon completion of each step, the mass was recorded and a soft X-ray image was taken. The relationship between moisture content and the average grayscale value of the soft X-ray image at each step was linear. In addition, the linear regressions overlapped each other regardless of the sample sections. These results showed that soft X-ray images could accurately estimate the moisture content. Applying this relationship to a small section of each sample, the moisture content distribution was estimated from the image differential between the soft X-ray pictures obtained from the sample in question and the same sample in the oven-dried condition. Moisture content profiles for 10-mm-wide parts at the centers of the samples were also obtained. The shapes of the profiles supported the evaluation method used in this study
-
The impact of image-size manipulation and sugar content on children's cereal consumption.
Neyens, E; Aerts, G; Smits, T
2015-12-01
Previous studies have demonstrated that portion sizes and food energy-density influence children's eating behavior. However, the potential effects of front-of-pack image-sizes of serving suggestions and sugar content have not been tested. Using a mixed experimental design among young children, this study examines the effects of image-size manipulation and sugar content on cereal and milk consumption. Children poured and consumed significantly more cereal and drank significantly more milk when exposed to a larger sized image of serving suggestion as compared to a smaller image-size. Sugar content showed no main effects. Nevertheless, cereal consumption only differed significantly between small and large image-sizes when sugar content was low. An advantage of this study was the mundane setting in which the data were collected: a school's dining room instead of an artificial lab. Future studies should include a control condition, with children eating by themselves to reflect an even more natural context. Copyright © 2015 Elsevier Ltd. All rights reserved.
-
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.
-
Supervised learning of tools for content-based search of image databases
Delanoy, Richard L.
1996-03-01
A computer environment, called the Toolkit for Image Mining (TIM), is being developed with the goal of enabling users with diverse interests and varied computer skills to create search tools for content-based image retrieval and other pattern matching tasks. Search tools are generated using a simple paradigm of supervised learning that is based on the user pointing at mistakes of classification made by the current search tool. As mistakes are identified, a learning algorithm uses the identified mistakes to build up a model of the user's intentions, construct a new search tool, apply the search tool to a test image, display the match results as feedback to the user, and accept new inputs from the user. Search tools are constructed in the form of functional templates, which are generalized matched filters capable of knowledge- based image processing. The ability of this system to learn the user's intentions from experience contrasts with other existing approaches to content-based image retrieval that base searches on the characteristics of a single input example or on a predefined and semantically- constrained textual query. Currently, TIM is capable of learning spectral and textural patterns, but should be adaptable to the learning of shapes, as well. Possible applications of TIM include not only content-based image retrieval, but also quantitative image analysis, the generation of metadata for annotating images, data prioritization or data reduction in bandwidth-limited situations, and the construction of components for larger, more complex computer vision algorithms.
-
Image content authentication based on channel coding
Zhang, Fan; Xu, Lei
2008-03-01
The content authentication determines whether an image has been tampered or not, and if necessary, locate malicious alterations made on the image. Authentication on a still image or a video are motivated by recipient's interest, and its principle is that a receiver must be able to identify the source of this document reliably. Several techniques and concepts based on data hiding or steganography designed as a means for the image authentication. This paper presents a color image authentication algorithm based on convolution coding. The high bits of color digital image are coded by the convolution codes for the tamper detection and localization. The authentication messages are hidden in the low bits of image in order to keep the invisibility of authentication. All communications channels are subject to errors introduced because of additive Gaussian noise in their environment. Data perturbations cannot be eliminated but their effect can be minimized by the use of Forward Error Correction (FEC) techniques in the transmitted data stream and decoders in the receiving system that detect and correct bits in error. This paper presents a color image authentication algorithm based on convolution coding. The message of each pixel is convolution encoded with the encoder. After the process of parity check and block interleaving, the redundant bits are embedded in the image offset. The tamper can be detected and restored need not accessing the original image.
-
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.
-
Parallel content-based sub-image retrieval using hierarchical searching.
Yang, Lin; Qi, Xin; Xing, Fuyong; Kurc, Tahsin; Saltz, Joel; Foran, David J
2014-04-01
The capacity to systematically search through large image collections and ensembles and detect regions exhibiting similar morphological characteristics is central to pathology diagnosis. Unfortunately, the primary methods used to search digitized, whole-slide histopathology specimens are slow and prone to inter- and intra-observer variability. The central objective of this research was to design, develop, and evaluate a content-based image retrieval system to assist doctors for quick and reliable content-based comparative search of similar prostate image patches. Given a representative image patch (sub-image), the algorithm will return a ranked ensemble of image patches throughout the entire whole-slide histology section which exhibits the most similar morphologic characteristics. This is accomplished by first performing hierarchical searching based on a newly developed hierarchical annular histogram (HAH). The set of candidates is then further refined in the second stage of processing by computing a color histogram from eight equally divided segments within each square annular bin defined in the original HAH. A demand-driven master-worker parallelization approach is employed to speed up the searching procedure. Using this strategy, the query patch is broadcasted to all worker processes. Each worker process is dynamically assigned an image by the master process to search for and return a ranked list of similar patches in the image. The algorithm was tested using digitized hematoxylin and eosin (H&E) stained prostate cancer specimens. We have achieved an excellent image retrieval performance. The recall rate within the first 40 rank retrieved image patches is ∼90%. Both the testing data and source code can be downloaded from http://pleiad.umdnj.edu/CBII/Bioinformatics/.
-
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
-
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.
-
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.
-
Design Guidelines for a Content-Based Image Retrieval Color-Selection Interface
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
-
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.
-
Basic strategies for valid cytometry using image analysis
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
-
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
-
Highly multiparametric analysis by mass cytometry.
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.
-
Detection of Isoflavones Content in Soybean Based on Hyperspectral Imaging Technology
Directory of Open Access Journals (Sweden)
Tan Kezhu
2014-04-01
Full Text Available Because of many important biological activities, Soybean isoflavones which has great potential for exploitation is significant to practical applications. Due to the conventional methods for determination of soybean isoflavones having long detection period, used too many reagents, couldn’t be detected on-line, and other issues, we propose hyperspectral imaging technology to detect the contents of soybean isoflavones. Based on the 40 varieties of soybeans produced in Heilongjiang province, we get the spectral reflection datum of soybean samples varied from the soybean’s hyperspectral images which are collected by the hyperspectral imaging system, and apply high performance liquid chromatography (HPLC method to determine the true value of the selected samples of isoflavones. The feature wavelengths for isoflavones content prediction (1516, 1572, 1691, 1716 and 1760 nm were selected based on correlation analysis. The prediction model was established by using the method of BP neural network in order to realize the prediction of soybean isoflavones content analysis. The experimental results show that, the ANN model could predict isoflavones content of soybean samples with of 0.9679, the average relative error is 0.8032 %, and the mean square error (MSE is 0.110328, which indicates the effectiveness of the proposed method and provides a theoretical basis for the applications of hyerspectral imaging in non-destructive detection for interior quality of soybean.
-
Nakazato, Munehiro; Manola, L.; Huang, Thomas S.
In content-based image retrieval (CBIR), experimental (trial-and-error) query with relevance feedback is essential for successful retrieval. Unfortunately, the traditional user interfaces are not suitable for trying different combinations of query examples. This is because first, these systems
-
Flow Cytometry Technician | Center for Cancer Research
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
-
Content Based Image Matching for Planetary Science
Deans, M. C.; Meyer, C.
2006-12-01
Planetary missions generate large volumes of data. With the MER rovers still functioning on Mars, PDS contains over 7200 released images from the Microscopic Imagers alone. These data products are only searchable by keys such as the Sol, spacecraft clock, or rover motion counter index, with little connection to the semantic content of the images. We have developed a method for matching images based on the visual textures in images. For every image in a database, a series of filters compute the image response to localized frequencies and orientations. Filter responses are turned into a low dimensional descriptor vector, generating a 37 dimensional fingerprint. For images such as the MER MI, this represents a compression ratio of 99.9965% (the fingerprint is approximately 0.0035% the size of the original image). At query time, fingerprints are quickly matched to find images with similar appearance. Image databases containing several thousand images are preprocessed offline in a matter of hours. Image matches from the database are found in a matter of seconds. We have demonstrated this image matching technique using three sources of data. The first database consists of 7200 images from the MER Microscopic Imager. The second database consists of 3500 images from the Narrow Angle Mars Orbital Camera (MOC-NA), which were cropped into 1024×1024 sub-images for consistency. The third database consists of 7500 scanned archival photos from the Apollo Metric Camera. Example query results from all three data sources are shown. We have also carried out user tests to evaluate matching performance by hand labeling results. User tests verify approximately 20% false positive rate for the top 14 results for MOC NA and MER MI data. This means typically 10 to 12 results out of 14 match the query image sufficiently. This represents a powerful search tool for databases of thousands of images where the a priori match probability for an image might be less than 1%. Qualitatively, correct
-
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
-
FuGEFlow: data model and markup language for flow cytometry.
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
-
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.
-
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.
-
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.
-
Spaceflight Flow Cytometry: Design Challenges and Applications
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.
-
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
-
Learning effective color features for content based image retrieval in dermatology
Bunte, Kerstin; Biehl, Michael; Jonkman, Marcel F.; Petkov, Nicolai
We investigate the extraction of effective color features for a content-based image retrieval (CBIR) application in dermatology. Effectiveness is measured by the rate of correct retrieval of images from four color classes of skin lesions. We employ and compare two different methods to learn
-
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
-
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.
-
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...
-
Ontology of gaps in content-based image retrieval.
Deserno, Thomas M; Antani, Sameer; Long, Rodney
2009-04-01
Content-based image retrieval (CBIR) is a promising technology to enrich the core functionality of picture archiving and communication systems (PACS). CBIR has a potential for making a strong impact in diagnostics, research, and education. Research as reported in the scientific literature, however, has not made significant inroads as medical CBIR applications incorporated into routine clinical medicine or medical research. The cause is often attributed (without supporting analysis) to the inability of these applications in overcoming the "semantic gap." The semantic gap divides the high-level scene understanding and interpretation available with human cognitive capabilities from the low-level pixel analysis of computers, based on mathematical processing and artificial intelligence methods. In this paper, we suggest a more systematic and comprehensive view of the concept of "gaps" in medical CBIR research. In particular, we define an ontology of 14 gaps that addresses the image content and features, as well as system performance and usability. In addition to these gaps, we identify seven system characteristics that impact CBIR applicability and performance. The framework we have created can be used a posteriori to compare medical CBIR systems and approaches for specific biomedical image domains and goals and a priori during the design phase of a medical CBIR application, as the systematic analysis of gaps provides detailed insight in system comparison and helps to direct future research.
-
Ploidy Levels among Species in the ‘Oxalis tuberosa Alliance’ as Inferred by Flow Cytometry
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...
-
Localization-based super-resolution imaging meets high-content screening.
Beghin, Anne; Kechkar, Adel; Butler, Corey; Levet, Florian; Cabillic, Marine; Rossier, Olivier; Giannone, Gregory; Galland, Rémi; Choquet, Daniel; Sibarita, Jean-Baptiste
2017-12-01
Single-molecule localization microscopy techniques have proven to be essential tools for quantitatively monitoring biological processes at unprecedented spatial resolution. However, these techniques are very low throughput and are not yet compatible with fully automated, multiparametric cellular assays. This shortcoming is primarily due to the huge amount of data generated during imaging and the lack of software for automation and dedicated data mining. We describe an automated quantitative single-molecule-based super-resolution methodology that operates in standard multiwell plates and uses analysis based on high-content screening and data-mining software. The workflow is compatible with fixed- and live-cell imaging and allows extraction of quantitative data like fluorophore photophysics, protein clustering or dynamic behavior of biomolecules. We demonstrate that the method is compatible with high-content screening using 3D dSTORM and DNA-PAINT based super-resolution microscopy as well as single-particle tracking.
-
Korsukov, Vladimir N.; Shchukovskaya, Tatyana N.; Kravtsov, Alexander L.; Popov, Youri A.
2002-07-01
Using flow cytometry a low DNA content in inoculated Yersinia pestis EV cells have been shown at the beginning of culture in Hottinger broth pH 7.2. The dependence serotonin action of its concentration on DNA content have been demonstrated. Serotonin accelerated Yersinia pestis culture growth during cultivation in Hottinger broth pH 7.2 both at 28 degrees C and 37 degrees C at concentration of 10-5 M.
-
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.
-
Retrieval Architecture with Classified Query for Content Based Image Recognition
Directory of Open Access Journals (Sweden)
Rik Das
2016-01-01
Full Text Available The consumer behavior has been observed to be largely influenced by image data with increasing familiarity of smart phones and World Wide Web. Traditional technique of browsing through product varieties in the Internet with text keywords has been gradually replaced by the easy accessible image data. The importance of image data has portrayed a steady growth in application orientation for business domain with the advent of different image capturing devices and social media. The paper has described a methodology of feature extraction by image binarization technique for enhancing identification and retrieval of information using content based image recognition. The proposed algorithm was tested on two public datasets, namely, Wang dataset and Oliva and Torralba (OT-Scene dataset with 3688 images on the whole. It has outclassed the state-of-the-art techniques in performance measure and has shown statistical significance.
-
Multiscale Distance Coherence Vector Algorithm for Content-Based Image Retrieval
Jiexian, Zeng; Xiupeng, Liu
2014-01-01
Multiscale distance coherence vector algorithm for content-based image retrieval (CBIR) is proposed due to the same descriptor with different shapes and the shortcomings of antinoise performance of the distance coherence vector algorithm. By this algorithm, the image contour curve is evolved by Gaussian function first, and then the distance coherence vector is, respectively, extracted from the contour of the original image and evolved images. Multiscale distance coherence vector was obtained by reasonable weight distribution of the distance coherence vectors of evolved images contour. This algorithm not only is invariable to translation, rotation, and scaling transformation but also has good performance of antinoise. The experiment results show us that the algorithm has a higher recall rate and precision rate for the retrieval of images polluted by noise. PMID:24883416
-
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.
-
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).
-
Determination of fat and total protein content in milk using conventional digital imaging.
Kucheryavskiy, Sergey; Melenteva, Anastasiia; Bogomolov, Andrey
2014-04-01
The applicability of conventional digital imaging to quantitative determination of fat and total protein in cow's milk, based on the phenomenon of light scatter, has been proved. A new algorithm for extracting features from digital images of milk samples has been developed. The algorithm takes into account spatial distribution of light, diffusely transmitted through a sample. The proposed method has been tested on two sample sets prepared from industrial raw milk standards, with variable fat and protein content. Partial Least-Squares (PLS) regression on the features calculated from images of monochromatically illuminated milk samples resulted in models with high prediction performance when analysed the sets separately (best models with cross-validated R(2)=0.974 for protein and R(2)=0.973 for fat content). However when analysed the sets jointly with the obtained results were significantly worse (best models with cross-validated R(2)=0.890 for fat content and R(2)=0.720 for protein content). The results have been compared with previously published Vis/SW-NIR spectroscopic study of similar samples. Copyright © 2013 Elsevier B.V. All rights reserved.
-
Phase-image-based content-addressable holographic data storage
John, Renu; Joseph, Joby; Singh, Kehar
2004-03-01
We propose and demonstrate the use of phase images for content-addressable holographic data storage. Use of binary phase-based data pages with 0 and π phase changes, produces uniform spectral distribution at the Fourier plane. The absence of strong DC component at the Fourier plane and more intensity of higher order spatial frequencies facilitate better recording of higher spatial frequencies, and improves the discrimination capability of the content-addressable memory. This improves the results of the associative recall in a holographic memory system, and can give low number of false hits even for small search arguments. The phase-modulated pixels also provide an opportunity of subtraction among data pixels leading to better discrimination between similar data pages.
-
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
-
Growth regulators, DNA content and anatomy in vitro -cultivated ...
African Journals Online (AJOL)
Growth regulators, DNA content and anatomy in vitro -cultivated Curcuma longa ... Shoots were inoculated in MS culture medium with the addition of 30 g/L of sucrose ... flow cytometry, utilizing two reference standards, green pea, and tomato.
-
Magnetic resonance imaging and quantitative analysis of contents of epidermoid and dermoid cysts
Energy Technology Data Exchange (ETDEWEB)
Takeshita, Mikihiko; Kubo, Osami; Hiyama, Hirofumi; Tajika, Yasuhiko; Izawa, Masahiro; Kagawa, Mizuo; Takakura, Kintomo; Kobayashi, Naotoshi; Toyoda, Masako [Tokyo Women' s Medical Coll. (Japan)
1994-07-01
The intracapsular cholesterol protein, and calcium contents of epidermoid and dermoid cysts from seven patients were compared with the signal intensities on T[sub 1]-weighted spin-echo magnetic resonance (MR) images. All specimens had a paste-like consistency when resected. Epidermoid and dermoid cysts demonstrated a wide range of cholesterol and calcium contents, and epidermoid cysts were not always rich in cholesterol. Five patients had cysts with lower signal intensity than white matter, which contained more than 18.3 mg/g wet weight of protein. One of these patients had the highest cholesterol content of all seven patients (22.25 mg/g wet weight) and another had the highest calcium content (0.75 mg/g wet weight). Two patients had cysts with higher signal intensity than white matter, with protein contents of lower than 4.3 mg/g wet weight. High protein content (>18.3 mg/g wet weight) may decrease signal intensity on T[sub 1]-weighted MR images, while low protein content (<4.3 mg/g wet weight) may increase signal intensity in epidermoid and dermoid cysts with high viscosity (paste-like consistency) contents. (author).
-
Hyperspectral imaging detection of decayed honey peaches based on their chlorophyll content.
Sun, Ye; Wang, Yihang; Xiao, Hui; Gu, Xinzhe; Pan, Leiqing; Tu, Kang
2017-11-15
Honey peach is a very common but highly perishable market fruit. When pathogens infect fruit, chlorophyll as one of the important components related to fruit quality, decreased significantly. Here, the feasibility of hyperspectral imaging to determine the chlorophyll content thus distinguishing diseased peaches was investigated. Three optimal wavelengths (617nm, 675nm, and 818nm) were selected according to chlorophyll content via successive projections algorithm. Partial least square regression models were established to determine chlorophyll content. Three band ratios were obtained using these optimal wavelengths, which improved spatial details, but also integrates the information of chemical composition from spectral characteristics. The band ratio values were suitable to classify the diseased peaches with 98.75% accuracy and clearly show the spatial distribution of diseased parts. This study provides a new perspective for the selection of optimal wavelengths of hyperspectral imaging via chlorophyll content, thus enabling the detection of fungal diseases in peaches. Copyright © 2017 Elsevier Ltd. All rights reserved.
-
Hessian-based quantitative image analysis of host-pathogen confrontation assays.
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.
-
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
-
Content Based Radiographic Images Indexing and Retrieval Using Pattern Orientation Histogram
Directory of Open Access Journals (Sweden)
Abolfazl Lakdashti
2008-06-01
Full Text Available Introduction: Content Based Image Retrieval (CBIR is a method of image searching and retrieval in a database. In medical applications, CBIR is a tool used by physicians to compare the previous and current medical images associated with patients pathological conditions. As the volume of pictorial information stored in medical image databases is in progress, efficient image indexing and retrieval is increasingly becoming a necessity. Materials and Methods: This paper presents a new content based radiographic image retrieval approach based on histogram of pattern orientations, namely pattern orientation histogram (POH. POH represents the spatial distribution of five different pattern orientations: vertical, horizontal, diagonal down/left, diagonal down/right and non-orientation. In this method, a given image is first divided into image-blocks and the frequency of each type of pattern is determined in each image-block. Then, local pattern histograms for each of these image-blocks are computed. Results: The method was compared to two well known texture-based image retrieval methods: Tamura and Edge Histogram Descriptors (EHD in MPEG-7 standard. Experimental results based on 10000 IRMA radiography image dataset, demonstrate that POH provides better precision and recall rates compared to Tamura and EHD. For some images, the recall and precision rates obtained by POH are, respectively, 48% and 18% better than the best of the two above mentioned methods. Discussion and Conclusion: Since we exploit the absolute location of the pattern in the image as well as its global composition, the proposed matching method can retrieve semantically similar medical images.
-
Plant leaf chlorophyll content retrieval based on a field imaging spectroscopy system.
Liu, Bo; Yue, Yue-Min; Li, Ru; Shen, Wen-Jing; Wang, Ke-Lin
2014-10-23
A field imaging spectrometer system (FISS; 380-870 nm and 344 bands) was designed for agriculture applications. In this study, FISS was used to gather spectral information from soybean leaves. The chlorophyll content was retrieved using a multiple linear regression (MLR), partial least squares (PLS) regression and support vector machine (SVM) regression. Our objective was to verify the performance of FISS in a quantitative spectral analysis through the estimation of chlorophyll content and to determine a proper quantitative spectral analysis method for processing FISS data. The results revealed that the derivative reflectance was a more sensitive indicator of chlorophyll content and could extract content information more efficiently than the spectral reflectance, which is more significant for FISS data compared to ASD (analytical spectral devices) data, reducing the corresponding RMSE (root mean squared error) by 3.3%-35.6%. Compared with the spectral features, the regression methods had smaller effects on the retrieval accuracy. A multivariate linear model could be the ideal model to retrieve chlorophyll information with a small number of significant wavelengths used. The smallest RMSE of the chlorophyll content retrieved using FISS data was 0.201 mg/g, a relative reduction of more than 30% compared with the RMSE based on a non-imaging ASD spectrometer, which represents a high estimation accuracy compared with the mean chlorophyll content of the sampled leaves (4.05 mg/g). Our study indicates that FISS could obtain both spectral and spatial detailed information of high quality. Its image-spectrum-in-one merit promotes the good performance of FISS in quantitative spectral analyses, and it can potentially be widely used in the agricultural sector.
-
Filtering adult image content with topic models
Lienhart, Rainer (Prof. Dr.); Hauke, Rudolf
2009-01-01
Protecting children from exposure to adult content has become a serious problem in the real world. Current statistics show that, for instance, the average age of first Internet exposure to pornography is 11 years, that the largest consumer group of Internet pornography is the age group of 12-to-17-year-olds and that 90% of the 8-to-16-year-olds have viewed porn online. To protect our children, effective algorithms for detecting adult images are needed. In this research we evaluate the use of ...
-
Cytobank: providing an analytics platform for community cytometry data analysis and collaboration.
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.
-
Shedding Light on Filovirus Infection with High-Content Imaging
Directory of Open Access Journals (Sweden)
Rekha G. Panchal
2012-08-01
Full Text Available Microscopy has been instrumental in the discovery and characterization of microorganisms. Major advances in high-throughput fluorescence microscopy and automated, high-content image analysis tools are paving the way to the systematic and quantitative study of the molecular properties of cellular systems, both at the population and at the single-cell level. High-Content Imaging (HCI has been used to characterize host-virus interactions in genome-wide reverse genetic screens and to identify novel cellular factors implicated in the binding, entry, replication and egress of several pathogenic viruses. Here we present an overview of the most significant applications of HCI in the context of the cell biology of filovirus infection. HCI assays have been recently implemented to quantitatively study filoviruses in cell culture, employing either infectious viruses in a BSL-4 environment or surrogate genetic systems in a BSL-2 environment. These assays are becoming instrumental for small molecule and siRNA screens aimed at the discovery of both cellular therapeutic targets and of compounds with anti-viral properties. We discuss the current practical constraints limiting the implementation of high-throughput biology in a BSL-4 environment, and propose possible solutions to safely perform high-content, high-throughput filovirus infection assays. Finally, we discuss possible novel applications of HCI in the context of filovirus research with particular emphasis on the identification of possible cellular biomarkers of virus infection.
-
Ontology of Gaps in Content-Based Image Retrieval
Deserno, Thomas M.; Antani, Sameer; Long, Rodney
2008-01-01
Content-based image retrieval (CBIR) is a promising technology to enrich the core functionality of picture archiving and communication systems (PACS). CBIR has a potential for making a strong impact in diagnostics, research, and education. Research as reported in the scientific literature, however, has not made significant inroads as medical CBIR applications incorporated into routine clinical medicine or medical research. The cause is often attributed (without supporting analysis) to the ina...
-
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)
-
A Novel Optimization-Based Approach for Content-Based Image Retrieval
Directory of Open Access Journals (Sweden)
Manyu Xiao
2013-01-01
Full Text Available Content-based image retrieval is nowadays one of the possible and promising solutions to manage image databases effectively. However, with the large number of images, there still exists a great discrepancy between the users’ expectations (accuracy and efficiency and the real performance in image retrieval. In this work, new optimization strategies are proposed on vocabulary tree building, retrieval, and matching methods. More precisely, a new clustering strategy combining classification and conventional K-Means method is firstly redefined. Then a new matching technique is built to eliminate the error caused by large-scaled scale-invariant feature transform (SIFT. Additionally, a new unit mechanism is proposed to reduce the cost of indexing time. Finally, the numerical results show that excellent performances are obtained in both accuracy and efficiency based on the proposed improvements for image retrieval.
-
Estimation of water content in the leaves of fruit trees using infra-red images
International Nuclear Information System (INIS)
Muramatsu, N.; Hiraoka, K.
2006-01-01
A method was developed to evaluate water contents of fruit trees using infra-red photography. The irrigation of potted satsuma mandarin trees and grapevines was suppressed to induce water stress. During the drought treatment the leaf edges of basal parts of the shoots of grapevines became necrotic and the area of necrosis extended as the duration of stress increased. Necrosis was clearly distinguished from the viable areas on infra-red images. In satsuma mandarin, an abscission layer formed at the basal part of the petiole, then the leaves fell. Thus, detailed analysis was indispensable for detecting of the leaf water content. After obtaining infra-red images of satsuma mandarin leaves with or without water stress, a background treatment (subtraction of the background image) was performed on the images, then the average brightness of the leaf was determined using image analyzing software (Image Pro-plus). Coefficient correlation between the water status index using the infra-red camera and water content determined from dry weight and fresh weight of leaves was significant (r = 0.917 for adaxial surface data and r = 0.880 for abaxial surface data). These data indicate that infra-red photography is useful for detecting the degree of plant water stress
-
Flow cytometry, fluorescent probes, and flashing bacteria
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,
-
Design and Application of Sensors for Chemical Cytometry.
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.
-
Subnuclear foci quantification using high-throughput 3D image cytometry
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.
-
Ploidy levels among species in the 'Oxalis tuberosa alliance' as inferred by flow cytometry.
Emshwiller, Eve
2002-06-01
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 accessions using chicken erythrocytes as internal standard. Ten Bolivian accessions of cultivated O. tuberosa were confirmed to be octoploid, with a mean nuclear DNA content of approx. 3.6 pg/2C. Two Peruvian wild Oxalis species, O. phaeotricha and O. picchensis, were inferred to be tetraploid (both with approx. 1.67 pg/2C), the latter being one of the putative progenitors of O. tuberosa identified by chloroplast-expressed glutamine synthetase data in prior work. The remaining accessions (from 78 populations provisionally identified as 35 species) were DNA diploid, with nuclear DNA contents varying from 0.79 to 1.34 pg/2C.
-
Nanoparticulate NaA zeolite composites for MRI: Effect of iron oxide content on image contrast
Gharehaghaji, Nahideh; Divband, Baharak; Zareei, Loghman
2018-06-01
In the current study, Fe3O4/NaA nanocomposites with various amounts of Fe3O4 (3.4, 6.8 & 10.2 wt%) were synthesized and characterized to study the effect of nano iron oxide content on the magnetic resonance (MR) image contrast. The cell viability of the nanocomposites was investigated by MTT assay method. T2 values as well as r2 relaxivities were determined with a 1.5 T MRI scanner. The results of the MTT assay confirmed the nanocomposites cytocompatibility up to 6.8% of the iron oxide content. Although the magnetization saturations and susceptibility values of the nanocomposites were increased as a function of the iron oxide content, their relaxivity was decreased from 921.78 mM-1 s-1 for the nanocomposite with the lowest iron oxide content to 380.16 mM-1 s-1 for the highest one. Therefore, Fe3O4/NaA nanocomposite with 3.4% iron oxide content led to the best MR image contrast. Nano iron oxide content and dispersion in the nanocomposites structure have important role in the nanocomposite r2 relaxivity and the MR image contrast. Aggregation of the iron oxide nanoparticles is a limiting factor in using of the high iron oxide content nanocomposites.
-
Quantitative image of bone mineral content
International Nuclear Information System (INIS)
Katoh, Tsuguhisa
1990-01-01
A dual energy subtraction system was constructed on an experimental basis for the quantitative image of bone mineral content. The system consists of a radiographing system and an image processor. Two radiograms were taken with dual x-ray energy in a single exposure using an x-ray beam dichromized by a tin filter. In this system, a film cassette was used where a low speed film-screen system, a copper filter and a high speed film-screen system were layered on top of each other. The images were read by a microdensitometer and processed by a personal computer. The image processing included the corrections of the film characteristics and heterogeneity in the x-ray field, and the dual energy subtraction in which the effect of the high energy component of the dichromized beam on the tube side image was corrected. In order to determine the accuracy of the system, experiments using wedge phantoms made of mixtures of epoxy resin and bone mineral-equivalent materials in various fractions were performed for various tube potentials and film processing conditions. The results indicated that the relative precision of the system was within ±4% and that the propagation of the film noise was within ±11 mg/cm 2 for the 0.2 mm pixels. The results also indicated that the system response was independent of the tube potential and the film processing condition. The bone mineral weight in each phalanx of the freshly dissected hand of a rhesus monkey was measured by this system and compared with the ash weight. The results showed an error of ±10%, slightly larger than that of phantom experiments, which is probably due to the effect of fat and the variation of focus-object distance. The air kerma in free air at the object was approximately 0.5 mGy for one exposure. The results indicate that this system is applicable to clinical use and provides useful information for evaluating a time-course of localized bone disease. (author)
-
Preibisch, Stephan; Rohlfing, Torsten; Hasak, Michael P.; Tomancak, Pavel
2008-03-01
Single Plane Illumination Microscopy (SPIM; Huisken et al., Nature 305(5686):1007-1009, 2004) is an emerging microscopic technique that enables live imaging of large biological specimens in their entirety. By imaging the living biological sample from multiple angles SPIM has the potential to achieve isotropic resolution throughout even relatively large biological specimens. For every angle, however, only a relatively shallow section of the specimen is imaged with high resolution, whereas deeper regions appear increasingly blurred. In order to produce a single, uniformly high resolution image, we propose here an image mosaicing algorithm that combines state of the art groupwise image registration for alignment with content-based image fusion to prevent degrading of the fused image due to regional blurring of the input images. For the registration stage, we introduce an application-specific groupwise transformation model that incorporates per-image as well as groupwise transformation parameters. We also propose a new fusion algorithm based on Gaussian filters, which is substantially faster than fusion based on local image entropy. We demonstrate the performance of our mosaicing method on data acquired from living embryos of the fruit fly, Drosophila, using four and eight angle acquisitions.
-
Development of automatic image analysis methods for high-throughput and high-content screening
Di, Zi
2013-01-01
This thesis focuses on the development of image analysis methods for ultra-high content analysis of high-throughput screens where cellular phenotype responses to various genetic or chemical perturbations that are under investigation. Our primary goal is to deliver efficient and robust image analysis
-
An investigation of content and media images in gay men's magazines.
Saucier, Jason A; Caron, Sandra L
2008-01-01
This study provides an analysis of gay men's magazines, examining both the content and advertisements. Four magazine titles were selected, including The Advocate, Genre, Instinct, and Out, each targeting gay men as its target audience. These magazines were coded for both article content and advertisement content. In the advertisement analysis, both the type of advertisement and characteristics of the men depicted within the advertisement when present. The results mirror previous research findings relating to the portrayal of women, including the objectification of specific body parts and the high community standards set by the images depicted. These findings were reinforced by both the advertisements and content analyzed to include a high degree of importance being placed on having the right body type. Implications for further research are discussed.
-
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
-
DIMOND II: Measures for optimising radiological information content and dose in digital imaging
International Nuclear Information System (INIS)
Dowling, A.; Malone, J.; Marsh, D.
2001-01-01
The European Commission concerted action on 'Digital Imaging: Measures for Optimising Radiological Information Content and Dose', DIMOND II, was conducted by 12 European partners over the period January 1997 to June 1999. The objective of the concerted action was to initiate a project in the area of digital medical imaging where practice was evolving without structured research in radiation protection, optimisation or justification. The main issues addressed were patient and staff dosimetry, image quality, quality criteria and technical issues. The scope included computed radiography (CR), image intensifier radiography and fluoroscopy, cardiology and interventional procedures. The concerted action was based on the consolidation of work conducted in the partner's institutions together with elective new work. Protocols and approaches to dosimetry, radiological information content/image quality measurement and quality criteria were established and presented at an international workshop held in Dublin in June 1999. Details of the work conducted during the DIMOND II concerted action and a summary of the main findings and conclusions are presented in this contribution. (author)
-
Müller, Henning; Michoux, Nicolas; Bandon, David; Geissbuhler, Antoine
2004-02-01
Content-based visual information retrieval (CBVIR) or content-based image retrieval (CBIR) has been one on the most vivid research areas in the field of computer vision over the last 10 years. The availability of large and steadily growing amounts of visual and multimedia data, and the development of the Internet underline the need to create thematic access methods that offer more than simple text-based queries or requests based on matching exact database fields. Many programs and tools have been developed to formulate and execute queries based on the visual or audio content and to help browsing large multimedia repositories. Still, no general breakthrough has been achieved with respect to large varied databases with documents of differing sorts and with varying characteristics. Answers to many questions with respect to speed, semantic descriptors or objective image interpretations are still unanswered. In the medical field, images, and especially digital images, are produced in ever-increasing quantities and used for diagnostics and therapy. The Radiology Department of the University Hospital of Geneva alone produced more than 12,000 images a day in 2002. The cardiology is currently the second largest producer of digital images, especially with videos of cardiac catheterization ( approximately 1800 exams per year containing almost 2000 images each). The total amount of cardiologic image data produced in the Geneva University Hospital was around 1 TB in 2002. Endoscopic videos can equally produce enormous amounts of data. With digital imaging and communications in medicine (DICOM), a standard for image communication has been set and patient information can be stored with the actual image(s), although still a few problems prevail with respect to the standardization. In several articles, content-based access to medical images for supporting clinical decision-making has been proposed that would ease the management of clinical data and scenarios for the integration of
-
"Fitspiration" on Social Media: A Content Analysis of Gendered Images.
Carrotte, Elise Rose; Prichard, Ivanka; Lim, Megan Su Cheng
2017-03-29
"Fitspiration" (also known as "fitspo") aims to inspire individuals to exercise and be healthy, but emerging research indicates exposure can negatively impact female body image. Fitspiration is frequently accessed on social media; however, it is currently unclear the degree to which messages about body image and exercise differ by gender of the subject. The aim of our study was to conduct a content analysis to identify the characteristics of fitspiration content posted across social media and whether this differs according to subject gender. Content tagged with #fitspo across Instagram, Facebook, Twitter, and Tumblr was extracted over a composite 30-minute period. All posts were analyzed by 2 independent coders according to a codebook. Of the 415/476 (87.2%) relevant posts extracted, most posts were on Instagram (360/415, 86.8%). Most posts (308/415, 74.2%) related thematically to exercise, and 81/415 (19.6%) related thematically to food. In total, 151 (36.4%) posts depicted only female subjects and 114/415 (27.5%) depicted only male subjects. Female subjects were typically thin but toned; male subjects were often muscular or hypermuscular. Within the images, female subjects were significantly more likely to be aged under 25 years (P<.001) than the male subjects, to have their full body visible (P=.001), and to have their buttocks emphasized (P<.001). Male subjects were more likely to have their face visible in the post (P=.005) than the female subjects. Female subjects were more likely to be sexualized than the male subjects (P=.002). Female #fitspo subjects typically adhered to the thin or athletic ideal, and male subjects typically adhered to the muscular ideal. Future research and interventional efforts should consider the potential objectifying messages in fitspiration, as it relates to both female and male body image. ©Elise Rose Carrotte, Ivanka Prichard, Megan Su Cheng Lim. Originally published in the Journal of Medical Internet Research (http
-
Biased discriminant euclidean embedding for content-based image retrieval.
Bian, Wei; Tao, Dacheng
2010-02-01
With many potential multimedia applications, content-based image retrieval (CBIR) has recently gained more attention for image management and web search. A wide variety of relevance feedback (RF) algorithms have been developed in recent years to improve the performance of CBIR systems. These RF algorithms capture user's preferences and bridge the semantic gap. However, there is still a big room to further the RF performance, because the popular RF algorithms ignore the manifold structure of image low-level visual features. In this paper, we propose the biased discriminative Euclidean embedding (BDEE) which parameterises samples in the original high-dimensional ambient space to discover the intrinsic coordinate of image low-level visual features. BDEE precisely models both the intraclass geometry and interclass discrimination and never meets the undersampled problem. To consider unlabelled samples, a manifold regularization-based item is introduced and combined with BDEE to form the semi-supervised BDEE, or semi-BDEE for short. To justify the effectiveness of the proposed BDEE and semi-BDEE, we compare them against the conventional RF algorithms and show a significant improvement in terms of accuracy and stability based on a subset of the Corel image gallery.
-
Nuclear DNA content variation among central European Koeleria taxa
Czech Academy of Sciences Publication Activity Database
Pečinka, A.; Suchánková, Pavla; Lysák, Martin; Trávníček, B.; Doležel, Jaroslav
2006-01-01
Roč. 98, - (2006), s. 117-122 ISSN 0305-7364 R&D Projects: GA MŠk(CZ) LC06004 Institutional research plan: CEZ:AV0Z50380511 Keywords : Chromosome number * nuclear DNA content * flow cytometry Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.448, year: 2006
-
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.
-
Gururaj, C.; Jayadevappa, D.; Tunga, Satish
2018-06-01
Medical field has seen a phenomenal improvement over the previous years. The invention of computers with appropriate increase in the processing and internet speed has changed the face of the medical technology. However there is still scope for improvement of the technologies in use today. One of the many such technologies of medical aid is the detection of afflictions of the eye. Although a repertoire of research has been accomplished in this field, most of them fail to address how to take the detection forward to a stage where it will be beneficial to the society at large. An automated system that can predict the current medical condition of a patient after taking the fundus image of his eye is yet to see the light of the day. Such a system is explored in this paper by summarizing a number of techniques for fundus image features extraction, predominantly hard exudate mining, coupled with Content Based Image Retrieval to develop an automation tool. The knowledge of the same would bring about worthy changes in the domain of exudates extraction of the eye. This is essential in cases where the patients may not have access to the best of technologies. This paper attempts at a comprehensive summary of the techniques for Content Based Image Retrieval (CBIR) or fundus features image extraction, and few choice methods of both, and an exploration which aims to find ways to combine these two attractive features, and combine them so that it is beneficial to all.
-
Gururaj, C.; Jayadevappa, D.; Tunga, Satish
2018-02-01
Medical field has seen a phenomenal improvement over the previous years. The invention of computers with appropriate increase in the processing and internet speed has changed the face of the medical technology. However there is still scope for improvement of the technologies in use today. One of the many such technologies of medical aid is the detection of afflictions of the eye. Although a repertoire of research has been accomplished in this field, most of them fail to address how to take the detection forward to a stage where it will be beneficial to the society at large. An automated system that can predict the current medical condition of a patient after taking the fundus image of his eye is yet to see the light of the day. Such a system is explored in this paper by summarizing a number of techniques for fundus image features extraction, predominantly hard exudate mining, coupled with Content Based Image Retrieval to develop an automation tool. The knowledge of the same would bring about worthy changes in the domain of exudates extraction of the eye. This is essential in cases where the patients may not have access to the best of technologies. This paper attempts at a comprehensive summary of the techniques for Content Based Image Retrieval (CBIR) or fundus features image extraction, and few choice methods of both, and an exploration which aims to find ways to combine these two attractive features, and combine them so that it is beneficial to all.
-
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 ...
-
“Fitspiration” on Social Media: A Content Analysis of Gendered Images
Prichard, Ivanka; Lim, Megan Su Cheng
2017-01-01
Background “Fitspiration” (also known as “fitspo”) aims to inspire individuals to exercise and be healthy, but emerging research indicates exposure can negatively impact female body image. Fitspiration is frequently accessed on social media; however, it is currently unclear the degree to which messages about body image and exercise differ by gender of the subject. Objective The aim of our study was to conduct a content analysis to identify the characteristics of fitspiration content posted across social media and whether this differs according to subject gender. Methods Content tagged with #fitspo across Instagram, Facebook, Twitter, and Tumblr was extracted over a composite 30-minute period. All posts were analyzed by 2 independent coders according to a codebook. Results Of the 415/476 (87.2%) relevant posts extracted, most posts were on Instagram (360/415, 86.8%). Most posts (308/415, 74.2%) related thematically to exercise, and 81/415 (19.6%) related thematically to food. In total, 151 (36.4%) posts depicted only female subjects and 114/415 (27.5%) depicted only male subjects. Female subjects were typically thin but toned; male subjects were often muscular or hypermuscular. Within the images, female subjects were significantly more likely to be aged under 25 years (P<.001) than the male subjects, to have their full body visible (P=.001), and to have their buttocks emphasized (P<.001). Male subjects were more likely to have their face visible in the post (P=.005) than the female subjects. Female subjects were more likely to be sexualized than the male subjects (P=.002). Conclusions Female #fitspo subjects typically adhered to the thin or athletic ideal, and male subjects typically adhered to the muscular ideal. Future research and interventional efforts should consider the potential objectifying messages in fitspiration, as it relates to both female and male body image. PMID:28356239
-
General Staining and Segmentation Procedures for High Content Imaging and Analysis.
Chambers, Kevin M; Mandavilli, Bhaskar S; Dolman, Nick J; Janes, Michael S
2018-01-01
Automated quantitative fluorescence microscopy, also known as high content imaging (HCI), is a rapidly growing analytical approach in cell biology. Because automated image analysis relies heavily on robust demarcation of cells and subcellular regions, reliable methods for labeling cells is a critical component of the HCI workflow. Labeling of cells for image segmentation is typically performed with fluorescent probes that bind DNA for nuclear-based cell demarcation or with those which react with proteins for image analysis based on whole cell staining. These reagents, along with instrument and software settings, play an important role in the successful segmentation of cells in a population for automated and quantitative image analysis. In this chapter, we describe standard procedures for labeling and image segmentation in both live and fixed cell samples. The chapter will also provide troubleshooting guidelines for some of the common problems associated with these aspects of HCI.
-
Parallel imaging microfluidic cytometer.
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.
-
Effects of acute gamma-irradiation on spermatogenesis as revealed by flow cytometry
Energy Technology Data Exchange (ETDEWEB)
Hacker, U; Schumann, J; Goehde, W [Muenster Univ. (Germany, F.R.)
1980-01-01
Mice irradiated with doses ranging from 0.1 to 15 Gy using a /sup 60/Co-source and controls were killed at intervals varying from 2 to 35 days after irradiation. The DNA content of the testicular cells in single cell suspensions was measured with the pulse cytophotometer to determine the frequencies of the different stages in the spermatogenesis. The relative amount of S-phase and 4c-cells was reduced initially but increased subsequently to hypernormal values. A decrease of 2c-cells indicated a higher cell-kill of diploid spermatogonia. Gamma ray-induced spermatids with abnormal DNA-values (diploid sperm) were identified. The results suggest that the spermatogenesis can be analysed with flow cytometry and used as a biologic dosimeter even for small doses of ionizing radiation.
-
Effects of acute gamma-irradiation on spermatogenesis as revealed by flow cytometry
International Nuclear Information System (INIS)
Hacker, U.; Schumann, J.; Goehde, W.
1980-01-01
Mice irradiated with doses ranging from 0.1 to 15 Gy using a 60 Co-source and controls were killed at intervals varying from 2 to 35 days after irradiation. The DNA content of the testicular cells in single cell suspensions was measured with the pulse cytophotometer to determine the frequencies of the different stages in the spermatogenesis. The relative amount of S-phase and 4c-cells was reduced initially but increased subsequently to hypernormal values. A decrease of 2c-cells indicated a higher cell-kill of diploid spermatogonia. Gamma ray-induced spermatids with abnormal DNA-values (diploid sperm) were identified. The results suggest that the spermatogenesis can be analysed with flow cytometry and used as a biologic dosimeter even for small doses of ionizing radiation. (Auth.)
-
Determination of fat and total protein content in milk using conventional digital imaging
DEFF Research Database (Denmark)
Kucheryavskiy, Sergey; Melenteva, Anastasiia; Bogomolov, Andrey
2014-01-01
into account spatial distribution of light, diffusely transmitted through a sample. The proposed method has been tested on two sample sets prepared from industrial raw milk standards, with variable fat and protein content. Partial Least-Squares (PLS) regression on the features calculated from images......The applicability of conventional digital imaging to quantitative determination of fat and total protein in cow’s milk, based on the phenomenon of light scatter, has been proved. A new algorithm for extracting features from digital images of milk samples has been developed. The algorithm takes...... of monochromatically illuminated milk samples resulted in models with high prediction performance when analysed the sets separately (best models with cross-validated R2=0.974 for protein and R2=0.973 for fat content). However when analysed the sets jointly the obtained results were significantly worse (best models...
-
The utilization of human color categorization for content-based image retrieval
van den Broek, Egon; Rogowitz, Bernice E.; Kisters, Peter M.F.; Pappas, Thrasyvoulos N.; Vuurpijl, Louis G.
2004-01-01
We present the concept of intelligent Content-Based Image Retrieval (iCBIR), which incorporates knowledge concerning human cognition in system development. The present research focuses on the utilization of color categories (or focal colors) for CBIR purposes, in particularly considered to be useful
-
The Study on the Attenuation of X-ray and Imaging Quality by Contents in Stomach
International Nuclear Information System (INIS)
Dong, Kyung Rae; Ji, Youn Sang; Kim, Chang Bok; Choi, Seong Kwan; Moon, Sang In; Dieter, Kevin
2009-01-01
This study examined the change in the attenuation of X-rays with the ROI (Region of Interest) in DR (Digital Radiography) according to the stomach contents by manufacturing a tissue equivalent material phantom to simulate real stomach tissue based on the assumption that there is some attenuation of X-rays and a difference in imaging quality according to the stomach contents. The transit dosage by the attenuation of X-rays decreased with increasing protein thickness, which altered the average ROI values in the film and DR images. A comparison of the change in average ROI values of the film and DR image showed that the image in film caused larger density changes with varying thickness of protein than the image by DR. The results indicate that NPO (nothing by mouth) is more important in film system than in DR system.
-
The Study on the Attenuation of X-ray and Imaging Quality by Contents in Stomach
Energy Technology Data Exchange (ETDEWEB)
Dong, Kyung Rae; Ji, Youn Sang; Kim, Chang Bok; Choi, Seong Kwan; Moon, Sang In [Dept. of Radiological Technology, Gwangju Health College University, Gwangju (Korea, Republic of); Dieter, Kevin [Dept. of Physical Therapy, Gwangju Health College University, Gwangju (Korea, Republic of)
2009-03-15
This study examined the change in the attenuation of X-rays with the ROI (Region of Interest) in DR (Digital Radiography) according to the stomach contents by manufacturing a tissue equivalent material phantom to simulate real stomach tissue based on the assumption that there is some attenuation of X-rays and a difference in imaging quality according to the stomach contents. The transit dosage by the attenuation of X-rays decreased with increasing protein thickness, which altered the average ROI values in the film and DR images. A comparison of the change in average ROI values of the film and DR image showed that the image in film caused larger density changes with varying thickness of protein than the image by DR. The results indicate that NPO (nothing by mouth) is more important in film system than in DR system.
-
'Strong is the new skinny': A content analysis of #fitspiration images on Instagram.
Tiggemann, Marika; Zaccardo, Mia
2018-07-01
'Fitspiration' is an online trend designed to inspire viewers towards a healthier lifestyle by promoting exercise and healthy food. This study provides a content analysis of fitspiration imagery on the social networking site Instagram. A set of 600 images were coded for body type, activity, objectification and textual elements. Results showed that the majority of images of women contained only one body type: thin and toned. In addition, most images contained objectifying elements. Accordingly, while fitspiration images may be inspirational for viewers, they also contain a number of elements likely to have negative effects on the viewer's body image.
-
Wavelet optimization for content-based image retrieval in medical databases.
Quellec, G; Lamard, M; Cazuguel, G; Cochener, B; Roux, C
2010-04-01
We propose in this article a content-based image retrieval (CBIR) method for diagnosis aid in medical fields. In the proposed system, images are indexed in a generic fashion, without extracting domain-specific features: a signature is built for each image from its wavelet transform. These image signatures characterize the distribution of wavelet coefficients in each subband of the decomposition. A distance measure is then defined to compare two image signatures and thus retrieve the most similar images in a database when a query image is submitted by a physician. To retrieve relevant images from a medical database, the signatures and the distance measure must be related to the medical interpretation of images. As a consequence, we introduce several degrees of freedom in the system so that it can be tuned to any pathology and image modality. In particular, we propose to adapt the wavelet basis, within the lifting scheme framework, and to use a custom decomposition scheme. Weights are also introduced between subbands. All these parameters are tuned by an optimization procedure, using the medical grading of each image in the database to define a performance measure. The system is assessed on two medical image databases: one for diabetic retinopathy follow up and one for screening mammography, as well as a general purpose database. Results are promising: a mean precision of 56.50%, 70.91% and 96.10% is achieved for these three databases, when five images are returned by the system. Copyright 2009 Elsevier B.V. All rights reserved.
-
Immune response to mycobacterial infection: lessons from flow cytometry.
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.
-
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.
-
Data File Standard for Flow Cytometry, version FCS 3.1.
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.
-
Content-Based Image Retrieval Benchmarking: Utilizing color categories and color distributions
van den Broek, Egon; Kisters, Peter M.F.; Vuurpijl, Louis G.
From a human centered perspective three ingredients for Content-Based Image Retrieval (CBIR) were developed. First, with their existence confirmed by experimental data, 11 color categories were utilized for CBIR and used as input for a new color space segmentation technique. The complete HSI color
-
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
-
Content-addressable read/write memories for image analysis
Snyder, W. E.; Savage, C. D.
1982-01-01
The commonly encountered image analysis problems of region labeling and clustering are found to be cases of search-and-rename problem which can be solved in parallel by a system architecture that is inherently suitable for VLSI implementation. This architecture is a novel form of content-addressable memory (CAM) which provides parallel search and update functions, allowing speed reductions down to constant time per operation. It has been proposed in related investigations by Hall (1981) that, with VLSI, CAM-based structures with enhanced instruction sets for general purpose processing will be feasible.
-
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
-
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
-
Using Sentinel-1 and Landsat 8 satellite images to estimate surface soil moisture content.
Mexis, Philippos-Dimitrios; Alexakis, Dimitrios D.; Daliakopoulos, Ioannis N.; Tsanis, Ioannis K.
2016-04-01
Nowadays, the potential for more accurate assessment of Soil Moisture (SM) content exploiting Earth Observation (EO) technology, by exploring the use of synergistic approaches among a variety of EO instruments has emerged. This study is the first to investigate the potential of Synthetic Aperture Radar (SAR) (Sentinel-1) and optical (Landsat 8) images in combination with ground measurements to estimate volumetric SM content in support of water management and agricultural practices. SAR and optical data are downloaded and corrected in terms of atmospheric, geometric and radiometric corrections. SAR images are also corrected in terms of roughness and vegetation with the synergistic use of Oh and Topp models using a dataset consisting of backscattering coefficients and corresponding direct measurements of ground parameters (moisture, roughness). Following, various vegetation indices (NDVI, SAVI, MSAVI, EVI, etc.) are estimated to record diachronically the vegetation regime within the study area and as auxiliary data in the final modeling. Furthermore, thermal images from optical data are corrected and incorporated to the overall approach. The basic principle of Thermal InfraRed (TIR) method is that Land Surface Temperature (LST) is sensitive to surface SM content due to its impact on surface heating process (heat capacity and thermal conductivity) under bare soil or sparse vegetation cover conditions. Ground truth data are collected from a Time-domain reflectometer (TRD) gauge network established in western Crete, Greece, during 2015. Sophisticated algorithms based on Artificial Neural Networks (ANNs) and Multiple Linear Regression (MLR) approaches are used to explore the statistical relationship between backscattering measurements and SM content. Results highlight the potential of SAR and optical satellite images to contribute to effective SM content detection in support of water resources management and precision agriculture. Keywords: Sentinel-1, Landsat 8, Soil
-
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
-
An Analysis of Images of Contention and Violence in Dagara and Akan Proverbial Expressions
Directory of Open Access Journals (Sweden)
Martin Kyiileyang
2017-04-01
Full Text Available Proverbial expressions have typical linguistic and figurative features. These are normally captivating to the listener. The expressive culture of the Dagara and Akan societies is embellished by these proverbial expressions. Most African proverbs, express various images depicting both pleasant and unpleasant situations in life. Unpleasant language normally depicts several terrifying images particularly when threats, insults and other forms of abuse are traded vehemently. Dagara and Akan proverbs are no exceptions to this phenomenon. This paper seeks to examine images of contention and violence depicted in Akan and Dagara proverbial expressions. To achieve this, a variety of proverbs from Akan and Dagara were analysed for their meanings using Yankah’s and Honeck’s Theories. The result revealed that structurally, as with many proverbs, the Akan and Dagara proverbial expressions are pithy and terse. The most dominant images of contention and violence in these expressions expose negative values and perceptions about the people who speak these languages.
-
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)
-
Directory of Open Access Journals (Sweden)
Wei Long
2016-09-01
Full Text Available Fast and accurate determination of effective bentonite content in used clay bonded sand is very important for selecting the correct mixing ratio and mixing process to obtain high-performance molding sand. Currently, the effective bentonite content is determined by testing the ethylene blue absorbed in used clay bonded sand, which is usually a manual operation with some disadvantages including complicated process, long testing time and low accuracy. A rapid automatic analyzer of the effective bentonite content in used clay bonded sand was developed based on image recognition technology. The instrument consists of auto stirring, auto liquid removal, auto titration, step-rotation and image acquisition components, and processor. The principle of the image recognition method is first to decompose the color images into three-channel gray images based on the photosensitive degree difference of the light blue and dark blue in the three channels of red, green and blue, then to make the gray values subtraction calculation and gray level transformation of the gray images, and finally, to extract the outer circle light blue halo and the inner circle blue spot and calculate their area ratio. The titration process can be judged to reach the end-point while the area ratio is higher than the setting value.
-
Method of detaching adherent cells for flow cytometry
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.
-
High-content analysis of single cells directly assembled on CMOS sensor based on color imaging.
Tanaka, Tsuyoshi; Saeki, Tatsuya; Sunaga, Yoshihiko; Matsunaga, Tadashi
2010-12-15
A complementary metal oxide semiconductor (CMOS) image sensor was applied to high-content analysis of single cells which were assembled closely or directly onto the CMOS sensor surface. The direct assembling of cell groups on CMOS sensor surface allows large-field (6.66 mm×5.32 mm in entire active area of CMOS sensor) imaging within a second. Trypan blue-stained and non-stained cells in the same field area on the CMOS sensor were successfully distinguished as white- and blue-colored images under white LED light irradiation. Furthermore, the chemiluminescent signals of each cell were successfully visualized as blue-colored images on CMOS sensor only when HeLa cells were placed directly on the micro-lens array of the CMOS sensor. Our proposed approach will be a promising technique for real-time and high-content analysis of single cells in a large-field area based on color imaging. Copyright © 2010 Elsevier B.V. All rights reserved.
-
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.
-
Alternatives to current flow cytometry data analysis for clinical and research studies.
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.
-
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.
-
Chmiel, P.; Ganzha, M.; Jaworska, T.; Paprzycki, M.
2017-10-01
Nowadays, as a part of systematic growth of volume, and variety, of information that can be found on the Internet, we observe also dramatic increase in sizes of available image collections. There are many ways to help users browsing / selecting images of interest. One of popular approaches are Content-Based Image Retrieval (CBIR) systems, which allow users to search for images that match their interests, expressed in the form of images (query by example). However, we believe that image search and retrieval could take advantage of semantic technologies. We have decided to test this hypothesis. Specifically, on the basis of knowledge captured in the CBIR, we have developed a domain ontology of residential real estate (detached houses, in particular). This allows us to semantically represent each image (and its constitutive architectural elements) represented within the CBIR. The proposed ontology was extended to capture not only the elements resulting from image segmentation, but also "spatial relations" between them. As a result, a new approach to querying the image database (semantic querying) has materialized, thus extending capabilities of the developed system.
-
International Nuclear Information System (INIS)
Feng Yanlin; Tan Jiaju; Yang Jie; Zhu Zheng; Yu Fengwen; He Xiaohong; Huang Kemin; Yuan Baihong; Su Shaodi
2002-01-01
Objective: To evaluate the value of 99 Tc m -methoxyisobutylisonitrile (MIBI) lung imaging in diagnosing benign/malign pulmonary disease and the relation of 99 Tc m -MIBI uptake ratio (UR) with lung cancer DNA content. Methods: Early and delay imaging were performed on 27 cases of benign lung disease and 46 cases of malign lung disease. Visual analysis of the images and T/N uptake ratio measurement were performed on every case. Cancer cell DNA content and DNA index (DI) were measured in 24 cases of malign pulmonary disease. Results: The delay phase UR was 1.13 ± 0.19 in benign disease group, and the delay phase UR was 1.45 ± 0.21 in malign disease group (t6.51, P 99 Tc m -MIBI is not an ideal imaging agent for differentiating pulmonary benign/malign disease. Lung cancer DNA content may be reflected by delay phase UR
-
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.
-
Ploidy Levels among Species in the ‘Oxalis tuberosa Alliance’ as Inferred by Flow Cytometry
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 accessions using chicken erythrocytes as internal standard. Ten Bolivian accessions of cultivated O. tuberosa were confirmed to be octoploid, with a mean nuclear DNA content of approx. 3·6 pg/2C. Two Peruvian wild Oxalis species, O. phaeotricha and O. picchensis, were inferred to be tetraploid (both with approx. 1·67 pg/2C), the latter being one of the putative progenitors of O. tuberosa identified by chloroplast‐expressed glutamine synthetase data in prior work. The remaining accessions (from 78 populations provisionally identified as 35 species) were DNA diploid, with nuclear DNA contents varying from 0·79 to 1·34 pg/2C. PMID:12102530
-
Wild immunology assessed by multidimensional mass cytometry.
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
-
Ingemi, Christopher M.; Owusu Twumasi, Jones; Yu, Tzuyang
2018-03-01
Detection and quantification of moisture content inside wood (timber) is key to ensuring safety and reliability of timber structures. Moisture inside wood attracts insects and fosters the development of fungi to attack the timber, causing significant damages and reducing the load bearing capacity during their design life. The use of non-destructive evaluation (NDE) techniques (e.g., microwave/radar, ultrasonic, stress wave, and X-ray) for condition assessment of timber structures is a good choice. NDE techniques provide information about the level of deterioration and material properties of timber structures without obstructing their functionality. In this study, microwave/radar NDE technique was selected for the characterization of wood at different moisture contents. A 12 in-by-3.5 in-by-1.5 in. white spruce specimen (picea glauca) was imaged at different moisture contents using a 10 GHz synthetic aperture radar (SAR) sensor inside an anechoic chamber. The presence of moisture was found to increase the SAR image amplitude as expected. Additionally, integrated SAR amplitude was found beneficial in modeling the moisture content inside the wood specimen.
-
Collagen Content Limits Optical Coherence Tomography Image Depth in Porcine Vocal Fold Tissue.
Garcia, Jordan A; Benboujja, Fouzi; Beaudette, Kathy; Rogers, Derek; Maurer, Rie; Boudoux, Caroline; Hartnick, Christopher J
2016-11-01
Vocal fold scarring, a condition defined by increased collagen content, is challenging to treat without a method of noninvasively assessing vocal fold structure in vivo. The goal of this study was to observe the effects of vocal fold collagen content on optical coherence tomography imaging to develop a quantifiable marker of disease. Excised specimen study. Massachusetts Eye and Ear Infirmary. Porcine vocal folds were injected with collagenase to remove collagen from the lamina propria. Optical coherence tomography imaging was performed preinjection and at 0, 45, 90, and 180 minutes postinjection. Mean pixel intensity (or image brightness) was extracted from images of collagenase- and control-treated hemilarynges. Texture analysis of the lamina propria at each injection site was performed to extract image contrast. Two-factor repeated measure analysis of variance and t tests were used to determine statistical significance. Picrosirius red staining was performed to confirm collagenase activity. Mean pixel intensity was higher at injection sites of collagenase-treated vocal folds than control vocal folds (P Fold change in image contrast was significantly increased in collagenase-treated vocal folds than control vocal folds (P = .002). Picrosirius red staining in control specimens revealed collagen fibrils most prominent in the subepithelium and above the thyroarytenoid muscle. Specimens treated with collagenase exhibited a loss of these structures. Collagen removal from vocal fold tissue increases image brightness of underlying structures. This inverse relationship may be useful in treating vocal fold scarring in patients. © American Academy of Otolaryngology—Head and Neck Surgery Foundation 2016.
-
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.
-
Combined analysis of cervical smears. Cytopathology, image cytometry and in situ hybridization
DEFF Research Database (Denmark)
Multhaupt, H; Bruder, E; Elit, L
1993-01-01
. HPV infection correlated with DNA polyploidy but was seen in 15 of 29 smears classified as cytologically normal. Morphologically abnormal Papanicolaou smears correlated with aneuploid DNA content. Smears classified as intraepithelial neoplasia correlated with aneuploid DNA content in all 12 cases...
-
[Flow cytometry in datecting lymph node micrometastasis in colorectal cancer].
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.
-
An efficient similarity measure for content based image retrieval using memetic algorithm
Directory of Open Access Journals (Sweden)
Mutasem K. Alsmadi
2017-06-01
Full Text Available Content based image retrieval (CBIR systems work by retrieving images which are related to the query image (QI from huge databases. The available CBIR systems extract limited feature sets which confine the retrieval efficacy. In this work, extensive robust and important features were extracted from the images database and then stored in the feature repository. This feature set is composed of color signature with the shape and color texture features. Where, features are extracted from the given QI in the similar fashion. Consequently, a novel similarity evaluation using a meta-heuristic algorithm called a memetic algorithm (genetic algorithm with great deluge is achieved between the features of the QI and the features of the database images. Our proposed CBIR system is assessed by inquiring number of images (from the test dataset and the efficiency of the system is evaluated by calculating precision-recall value for the results. The results were superior to other state-of-the-art CBIR systems in regard to precision.
-
Determination of chitin content in fungal cell wall: an alternative flow cytometric method.
Costa-de-Oliveira, Sofia; Silva, Ana P; Miranda, Isabel M; Salvador, Alexandre; Azevedo, Maria M; Munro, Carol A; Rodrigues, Acácio G; Pina-Vaz, Cidália
2013-03-01
The conventional methods used to evaluate chitin content in fungi, such as biochemical assessment of glucosamine release after acid hydrolysis or epifluorescence microscopy, are low throughput, laborious, time-consuming, and cannot evaluate a large number of cells. We developed a flow cytometric assay, efficient, and fast, based on Calcofluor White staining to measure chitin content in yeast cells. A staining index was defined, its value was directly related to chitin amount and taking into consideration the different levels of autofluorecence. Twenty-two Candida spp. and four Cryptococcus neoformans clinical isolates with distinct susceptibility profiles to caspofungin were evaluated. Candida albicans clinical isolate SC5314, and isogenic strains with deletions in chitin synthase 3 (chs3Δ/chs3Δ) and genes encoding predicted GlycosylPhosphatidylInositol (GPI)-anchored proteins (pga31Δ/Δ and pga62Δ/Δ), were used as controls. As expected, the wild-type strain displayed a significant higher chitin content (P relationship between chitin content and antifungal drug susceptibility phenotype was found, an association was established between the paradoxical growth effect in the presence of high caspofungin concentrations and the chitin content. This novel flow cytometry protocol revealed to be a simple and reliable assay to estimate cell wall chitin content of fungi. Copyright © 2013 International Society for Advancement of Cytometry.
-
Huang, Meiyan; Yang, Wei; Wu, Yao; Jiang, Jun; Gao, Yang; Chen, Yang; Feng, Qianjin; Chen, Wufan; Lu, Zhentai
2014-01-01
This study aims to develop content-based image retrieval (CBIR) system for the retrieval of T1-weighted contrast-enhanced MR (CE-MR) images of brain tumors. When a tumor region is fed to the CBIR system as a query, the system attempts to retrieve tumors of the same pathological category. The bag-of-visual-words (BoVW) model with partition learning is incorporated into the system to extract informative features for representing the image contents. Furthermore, a distance metric learning algorithm called the Rank Error-based Metric Learning (REML) is proposed to reduce the semantic gap between low-level visual features and high-level semantic concepts. The effectiveness of the proposed method is evaluated on a brain T1-weighted CE-MR dataset with three types of brain tumors (i.e., meningioma, glioma, and pituitary tumor). Using the BoVW model with partition learning, the mean average precision (mAP) of retrieval increases beyond 4.6% with the learned distance metrics compared with the spatial pyramid BoVW method. The distance metric learned by REML significantly outperforms three other existing distance metric learning methods in terms of mAP. The mAP of the CBIR system is as high as 91.8% using the proposed method, and the precision can reach 93.1% when the top 10 images are returned by the system. These preliminary results demonstrate that the proposed method is effective and feasible for the retrieval of brain tumors in T1-weighted CE-MR Images.
-
Directory of Open Access Journals (Sweden)
Meiyan Huang
Full Text Available This study aims to develop content-based image retrieval (CBIR system for the retrieval of T1-weighted contrast-enhanced MR (CE-MR images of brain tumors. When a tumor region is fed to the CBIR system as a query, the system attempts to retrieve tumors of the same pathological category. The bag-of-visual-words (BoVW model with partition learning is incorporated into the system to extract informative features for representing the image contents. Furthermore, a distance metric learning algorithm called the Rank Error-based Metric Learning (REML is proposed to reduce the semantic gap between low-level visual features and high-level semantic concepts. The effectiveness of the proposed method is evaluated on a brain T1-weighted CE-MR dataset with three types of brain tumors (i.e., meningioma, glioma, and pituitary tumor. Using the BoVW model with partition learning, the mean average precision (mAP of retrieval increases beyond 4.6% with the learned distance metrics compared with the spatial pyramid BoVW method. The distance metric learned by REML significantly outperforms three other existing distance metric learning methods in terms of mAP. The mAP of the CBIR system is as high as 91.8% using the proposed method, and the precision can reach 93.1% when the top 10 images are returned by the system. These preliminary results demonstrate that the proposed method is effective and feasible for the retrieval of brain tumors in T1-weighted CE-MR Images.
-
System for accessing a collection of histology images using content-based strategies
International Nuclear Information System (INIS)
Gonzalez F; Caicedo J C; Cruz Roa A; Camargo, J; Spinel, C
2010-01-01
Histology images are an important resource for research, education and medical practice. The availability of image collections with reference purposes is limited to printed formats such as books and specialized journals. When histology image sets are published in digital formats, they are composed of some tens of images that do not represent the wide diversity of biological structures that can be found in fundamental tissues; making a complete histology image collection available to the general public having a great impact on research and education in different areas such as medicine, biology and natural sciences. This work presents the acquisition process of a histology image collection with 20,000 samples in digital format, from tissue processing to digital image capturing. The main purpose of collecting these images is to make them available as reference material to the academic community. In addition, this paper presents the design and architecture of a system to query and explore the image collection, using content-based image retrieval tools and text-based search on the annotations provided by experts. The system also offers novel image visualization methods to allow easy identification of interesting images among hundreds of possible pictures. The system has been developed using a service-oriented architecture and allows web-based access in http://www.informed.unal.edu.co
-
Sorting catalytically active polymersome nanoreactors by flow cytometry
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
-
Content-based quality evaluation of color images: overview and proposals
Tremeau, Alain; Richard, Noel; Colantoni, Philippe; Fernandez-Maloigne, Christine
2003-12-01
The automatic prediction of perceived quality from image data in general, and the assessment of particular image characteristics or attributes that may need improvement in particular, becomes an increasingly important part of intelligent imaging systems. The purpose of this paper is to propose to the color imaging community in general to develop a software package available on internet to help the user to select among all these approaches which is better appropriated to a given application. The ultimate goal of this project is to propose, next to implement, an open and unified color imaging system to set up a favourable context for the evaluation and analysis of color imaging processes. Many different methods for measuring the performance of a process have been proposed by different researchers. In this paper, we will discuss the advantages and shortcomings of most of main analysis criteria and performance measures currently used. The aim is not to establish a harsh competition between algorithms or processes, but rather to test and compare the efficiency of methodologies firstly to highlight strengths and weaknesses of a given algorithm or methodology on a given image type and secondly to have these results publicly available. This paper is focused on two important unsolved problems. Why it is so difficult to select a color space which gives better results than another one? Why it is so difficult to select an image quality metric which gives better results than another one, with respect to the judgment of the Human Visual System? Several methods used either in color imaging or in image quality will be thus discussed. Proposals for content-based image measures and means of developing a standard test suite for will be then presented. The above reference advocates for an evaluation protocol based on an automated procedure. This is the ultimate goal of our proposal.
-
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 (
-
Depth-resolved incoherent and coherent wide-field high-content imaging (Conference Presentation)
So, Peter T.
2016-03-01
Recent advances in depth-resolved wide-field imaging technique has enabled many high throughput applications in biology and medicine. Depth resolved imaging of incoherent signals can be readily accomplished with structured light illumination or nonlinear temporal focusing. The integration of these high throughput systems with novel spectroscopic resolving elements further enable high-content information extraction. We will introduce a novel near common-path interferometer and demonstrate its uses in toxicology and cancer biology applications. The extension of incoherent depth-resolved wide-field imaging to coherent modality is non-trivial. Here, we will cover recent advances in wide-field 3D resolved mapping of refractive index, absorbance, and vibronic components in biological specimens.
-
Relevance Feedback in Content Based Image Retrieval: A Review
Directory of Open Access Journals (Sweden)
Manesh B. Kokare
2011-01-01
Full Text Available This paper provides an overview of the technical achievements in the research area of relevance feedback (RF in content-based image retrieval (CBIR. Relevance feedback is a powerful technique in CBIR systems, in order to improve the performance of CBIR effectively. It is an open research area to the researcher to reduce the semantic gap between low-level features and high level concepts. The paper covers the current state of art of the research in relevance feedback in CBIR, various relevance feedback techniques and issues in relevance feedback are discussed in detail.
-
Cytometric analysis of shape and DNA content in mammalian sperm
Energy Technology Data Exchange (ETDEWEB)
Gledhill, B.L.
1983-10-10
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. Sperm were analyzed by flow cytometry and slit-scan flow analysis for injury following the exposure of testes to mutagens. The utility of flow cytometry in genotoxin screening and monitoring of occupational exposure was evaluated. The technique proved valuable in separation of X- and Y-chromosome bearing sperm and the potential applicability of this technique in artificial insemination and a solution, of accurately assessing the DNA content of sperm were evaluated-with reference to determination of X- and Y-chromosome bearing sperm.
-
Cytometric analysis of shape and DNA content in 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. Sperm were analyzed by flow cytometry and slit-scan flow analysis for injury following the exposure of testes to mutagens. The utility of flow cytometry in genotoxin screening and monitoring of occupational exposure was evaluated. The technique proved valuable in separation of X- and Y-chromosome bearing sperm and the potential applicability of this technique in artificial insemination and a solution, of accurately assessing the DNA content of sperm were evaluated-with reference to determination of X- and Y-chromosome bearing sperm
-
Honey bee hemocyte profiling by flow cytometry.
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.
-
Screening of carcinoma metastasis by flow cytometry: A study of 238 cases.
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.
-
Anti-cancer agents in Saudi Arabian herbals revealed by automated high-content imaging
Hajjar, Dina A.; Kremb, Stephan Georg; Sioud, Salim; Emwas, Abdul-Hamid M.; Voolstra, Christian R.; Ravasi, Timothy
2017-01-01
in cancer therapy. Here, we used cell-based phenotypic profiling and image-based high-content screening to study the mode of action and potential cellular targets of plants historically used in Saudi Arabia's traditional medicine. We compared the cytological
-
The Means: Cytometry and Mass Spectrometry Converge in a Single Cell Deep Profiling Platform
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.
-
Methodology and application of flow cytometry for investigation of human malaria parasites.
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.
-
Heller, Rebecca; Martin-Biggers, Jennifer; Berhaupt-Glickstein, Amanda; Quick, Virginia; Byrd-Bredbenner, Carol
2015-10-01
To determine whether food label information and advertisements for foods containing no fruit cause children to have a false impression of the foods' fruit content. In the food label condition, a trained researcher showed each child sixteen different food label photographs depicting front-of-food label packages that varied with regard to fruit content (i.e. real fruit v. sham fruit) and label elements. In the food advertisement condition, children viewed sixteen, 30 s television food advertisements with similar fruit content and label elements as in the food label condition. After viewing each food label and advertisement, children responded to the question 'Did they use fruit to make this?' with responses of yes, no or don't know. Schools, day-care centres, after-school programmes and other community groups. Children aged 4-7 years. In the food label condition, χ 2 analysis of within fruit content variation differences indicated children (n 58; mean age 4·2 years) were significantly more accurate in identifying real fruit foods as the label's informational load increased and were least accurate when neither a fruit name nor an image was on the label. Children (n 49; mean age 5·4 years) in the food advertisement condition were more likely to identify real fruit foods when advertisements had fruit images compared with when no image was included, while fruit images in advertisements for sham fruit foods significantly reduced accuracy of responses. Findings suggest that labels and advertisements for sham fruit foods mislead children with regard to the food's real fruit content.
-
International Nuclear Information System (INIS)
Irwan, Roy; Edens, Mireille A.; Sijens, Paul E.
2008-01-01
A recently published Dixon-based MRI method for quantifying liver fat content using dual-echo breath-hold gradient echo imaging was validated by phantom experiments and compared with results of biopsy in two patients (Radiology 2005;237:1048-1055). We applied this method in ten healthy volunteers and compared the outcomes with the results of MR spectroscopy (MRS), the gold standard in quantifying liver fat content. Novel was the use of spectroscopic imaging yielding the variations in fat content across the liver rather than a single value obtained by single voxel MRS. Compared with the results of MRS, liver fat content according to MRI was too high in nine subjects (range 3.3-10.7% vs. 0.9-7.7%) and correct in one (21.1 vs. 21.3%). Furthermore, in one of the ten subjects the MRI fat content according to the Dixon-based MRI method was incorrect due to a (100-x) versus x percent lipid content mix-up. The second problem was fixed by a minor adjustment of the MRI algorithm. Despite systematic overestimation of liver fat contents by MRI, Spearman's correlation between the adjusted MRI liver fat contents with MRS was high (r = 0.927, P < 0.001). Even after correction of the algorithm, the problem remaining with the Dixon-based MRI method for the assessment of liver fat content,is that, at the lower end range, liver fat content is systematically overestimated by 4%. (orig.)
-
Directory of Open Access Journals (Sweden)
Sebastiaan Hammer
Full Text Available OBJECTIVE: To assess the feasibility of renal proton magnetic resonance spectroscopy for quantification of triglyceride content and to compare spectral quality and reproducibility without and with respiratory motion compensation in vivo. MATERIALS AND METHODS: The Institutional Review Board of our institution approved the study protocol, and written informed consent was obtained. After technical optimization, a total of 20 healthy volunteers underwent renal proton magnetic resonance spectroscopy of the renal cortex both without and with respiratory motion compensation and volume tracking. After the first session the subjects were repositioned and the protocol was repeated to assess reproducibility. Spectral quality (linewidth of the water signal and triglyceride content were quantified. Bland-Altman analyses and a test by Pitman were performed. RESULTS: Linewidth changed from 11.5±0.4 Hz to 10.7±0.4 Hz (all data pooled, p<0.05, without and with respiratory motion compensation respectively. Mean % triglyceride content in the first and second session without respiratory motion compensation were respectively 0.58±0.12% and 0.51±0.14% (P = NS. Mean % triglyceride content in the first and second session with respiratory motion compensation were respectively 0.44±0.10% and 0.43±0.10% (P = NS between sessions and P = NS compared to measurements with respiratory motion compensation. Bland-Altman analyses showed narrower limits of agreement and a significant difference in the correlated variances (correlation of -0.59, P<0.05. CONCLUSION: Metabolic imaging of the human kidney using renal proton magnetic resonance spectroscopy is a feasible tool to assess cortical triglyceride content in humans in vivo and the use of respiratory motion compensation significantly improves spectral quality and reproducibility. Therefore, respiratory motion compensation seems a necessity for metabolic imaging of renal triglyceride content in vivo.
-
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.
-
International Society for the Advancement of Cytometry cell sorter biosafety standards.
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.
-
Analysis of repetitive DNA in chromosomes by flow cytometry
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
-
DIRECT FLOW-CYTOMETRY OF ANAEROBIC-BACTERIA IN HUMAN FECES
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
-
Improved and Reproducible Flow Cytometry Methodology for Nuclei Isolation from Single Root Meristem
Directory of Open Access Journals (Sweden)
Thaís Cristina Ribeiro Silva
2010-01-01
Full Text Available Root meristems have increasingly been target of cell cycle studies by flow cytometric DNA content quantification. Moreover, roots can be an alternative source of nuclear suspension when leaves become unfeasible and for chromosome analysis and sorting. In the present paper, a protocol for intact nuclei isolation from a single root meristem was developed. This proceeding was based on excision of the meristematic region using a prototypical slide, followed by short enzymatic digestion and mechanical isolation of nuclei during homogenization with a hand mixer. Such parameters were optimized for reaching better results. Satisfactory nuclei amounts were extracted and analyzed by flow cytometry, producing histograms with reduced background noise and CVs between 3.2 and 4.1%. This improved and reproducible technique was shown to be rapid, inexpensive, and simple for nuclear extraction from a single root tip, and can be adapted for other plants and purposes.
-
Gating-ML: XML-based gating descriptions in flow cytometry.
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.
-
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.
-
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...
-
FT-IR imaging for quantitative determination of liver fat content in non-alcoholic fatty liver.
Kochan, K; Maslak, E; Chlopicki, S; Baranska, M
2015-08-07
In this work we apply FT-IR imaging of large areas of liver tissue cross-section samples (∼5 cm × 5 cm) for quantitative assessment of steatosis in murine model of Non-Alcoholic Fatty Liver (NAFLD). We quantified the area of liver tissue occupied by lipid droplets (LDs) by FT-IR imaging and Oil Red O (ORO) staining for comparison. Two alternative FT-IR based approaches are presented. The first, straightforward method, was based on average spectra from tissues and provided values of the fat content by using a PLS regression model and the reference method. The second one – the chemometric-based method – enabled us to determine the values of the fat content, independently of the reference method by means of k-means cluster (KMC) analysis. In summary, FT-IR images of large size liver sections may prove to be useful for quantifying liver steatosis without the need of tissue staining.
-
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.
-
The content of social media's shared images about Ebola: a retrospective study.
Seltzer, E K; Jean, N S; Kramer-Golinkoff, E; Asch, D A; Merchant, R M
2015-09-01
Social media have strongly influenced awareness and perceptions of public health emergencies, but a considerable amount of social media content is now carried through images, rather than just text. This study's objective is to explore how image-sharing platforms are used for information dissemination in public health emergencies. Retrospective review of images posted on two popular image-sharing platforms to characterize public discourse about Ebola. Using the keyword '#ebola' we identified a 1% sample of images posted on Instagram and Flickr across two sequential weeks in November 2014. Images from both platforms were independently coded by two reviewers and characterized by themes. We reviewed 1217 images posted on Instagram and Flickr and identified themes. Nine distinct themes were identified. These included: images of health care workers and professionals [308 (25%)], West Africa [75 (6%)], the Ebola virus [59 (5%)], and artistic renderings of Ebola [64 (5%)]. Also identified were images with accompanying embedded text related to Ebola and associated: facts [68 (6%)], fears [40 (3%)], politics [46 (4%)], and jokes [284 (23%)]. Several [273 (22%)] images were unrelated to Ebola or its sequelae. Instagram images were primarily coded as jokes [255 (42%)] or unrelated [219 (36%)], while Flickr images primarily depicted health care workers and other professionals [281 (46%)] providing care or other services for prevention or treatment. Image sharing platforms are being used for information exchange about public health crises, like Ebola. Use differs by platform and discerning these differences can help inform future uses for health care professionals and researchers seeking to assess public fears and misinformation or provide targeted education/awareness interventions. Copyright © 2015 The Royal Institute of Public Health. All rights reserved.
-
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.
-
Disability in physical education textbooks: an analysis of image content.
Táboas-Pais, María Inés; Rey-Cao, Ana
2012-10-01
The aim of this paper is to show how images of disability are portrayed in physical education textbooks for secondary schools in Spain. The sample was composed of 3,316 images published in 36 textbooks by 10 publishing houses. A content analysis was carried out using a coding scheme based on categories employed in other similar studies and adapted to the requirements of this study with additional categories. The variables were camera angle, gender, type of physical activity, field of practice, space, and level. Univariate and bivariate descriptive analyses were also carried out. The Pearson chi-square statistic was used to identify associations between the variables. Results showed a noticeable imbalance between people with disabilities and people without disabilities, and women with disabilities were less frequently represented than men with disabilities. People with disabilities were depicted as participating in a very limited variety of segregated, competitive, and elite sports activities.
-
Candidiasis and the impact of flow cytometry on antifungal drug discovery.
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.
-
Nedosekin, D. A.; Sarimollaoglu, M.; Foster, S.; Galanzha, E. I.; Zharov, V. P.
2013-03-01
Fluorescence flow cytometry is a well-established analytical tool that provides quantification of multiple biological parameters of cells at molecular levels, including their functional states, morphology, composition, proliferation, and protein expression. However, only the fluorescence and scattering parameters of the cells or labels are available for detection. Cell pigmentation, presence of non-fluorescent dyes or nanoparticles cannot be reliably quantified. Herewith, we present a novel photoacoustic (PA) flow cytometry design for simple integration of absorbance measurements into schematics of conventional in vitro flow cytometers. The integrated system allow simultaneous measurements of light absorbance, scattering and of multicolor fluorescence from single cells in the flow at rates up to 2 m/s. We compared various combinations of excitation laser sources for multicolor detection, including simultaneous excitation of PA and fluorescence using a single 500 kHz pulsed nanosecond laser. Multichannel detection scheme allows simultaneous detection of up to 8 labels, including 4 fluorescent tags and 4 PA colors. In vitro PA-fluorescence flow cytometer was used for studies of nanoparticles uptake and for the analysis of cell line pigmentation, including genetically encoded melanin expression in breast cancer cell line. We demonstrate that this system can be used for direct nanotoxicity studies with simultaneous quantification of nanoparticles content and assessment of cell viability using a conventional fluorescent apoptosis assays.
-
Nucleus fingerprinting for the unique identification of Feulgen-stained nuclei
Friedrich, David; Brozio, Matthias; Bell, André; Biesterfeld, Stefan; Böcking, Alfred; Aach, Til
2012-03-01
DNA Image Cytometry is a method for non-invasive cancer diagnosis which measures the DNA content of Feulgen-stained nuclei. DNA content is measured using a microscope system equipped with a digital camera as a densitometer and estimating the DNA content from the absorption of light when passing through the nuclei. However, a DNA Image Cytometry measurement is only valid if each nucleus is only measured once. To assist the user in preventing multiple measurements of the same nucleus, we have developed a unique digital identifier for the characterization of Feulgen-stained nuclei, the so called Nucleus Fingerprint. Only nuclei with a new fingerprint can be added to the measurement. This fingerprint is based on basic nucleus features, the contour of the nucleus and the spatial relationship to nuclei in the vicinity. Based on this characterization, a classifier for testing two nuclei for identity is presented. In a pairwise comparison of ~40000 pairs of mutually different nuclei, 99.5% were classified as different. In another 450 tests, the fingerprints of the same nucleus recorded a second time were in all cases judged identical. We therefore conclude that our Nucleus Fingerprint approach robustly prevents the repeated measurement of nuclei in DNA Image Cytometry.
-
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
-
Optimizing transformations for automated, high throughput analysis of flow cytometry data.
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
-
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
-
Evolutionary and Modern Image Content Differentially Influence the Processing of Emotional Pictures
Directory of Open Access Journals (Sweden)
Matthias Dhum
2017-08-01
Full Text Available From an evolutionary perspective, environmental threats relevant for survival constantly challenged human beings. Current research suggests the evolution of a fear processing module in the brain to cope with these threats. Recently, humans increasingly encountered modern threats (e.g., guns or car accidents in addition to evolutionary threats (e.g., snakes or predators which presumably required an adaptation of perception and behavior. However, the neural processes underlying the perception of these different threats remain to be elucidated. We investigated the effect of image content (i.e., evolutionary vs. modern threats on the activation of neural networks of emotion processing. During functional magnetic resonance imaging (fMRI 41 participants watched affective pictures displaying evolutionary-threatening, modern-threatening, evolutionary-neutral and modern-neutral content. Evolutionary-threatening stimuli evoked stronger activations than modern-threatening stimuli in left inferior frontal gyrus and thalamus, right middle frontal gyrus and parietal regions as well as bilaterally in parietal regions, fusiform gyrus and bilateral amygdala. We observed the opposite effect, i.e., higher activity for modern-threatening than for evolutionary-threatening stimuli, bilaterally in the posterior cingulate and the parahippocampal gyrus. We found no differences in subjective arousal ratings between the two threatening conditions. On the valence scale though, subjects rated modern-threatening pictures significantly more negative than evolutionary-threatening pictures, indicating a higher level of perceived threat. The majority of previous studies show a positive relationship between arousal rating and amygdala activity. However, comparing fMRI results with behavioral findings we provide evidence that neural activity in fear processing areas is not only driven by arousal or valence, but presumably also by the evolutionary content of the stimulus. This has
-
Near-Infrared Imaging for Spatial Mapping of Organic Content in Petroleum Source Rocks
Mehmani, Y.; Burnham, A. K.; Vanden Berg, M. D.; Tchelepi, H.
2017-12-01
Natural gas from unconventional petroleum source rocks (shales) plays a key role in our transition towards sustainable low-carbon energy production. The potential for carbon storage (in adsorbed state) in these formations further aligns with efforts to mitigate climate change. Optimizing production and development from these resources requires knowledge of the hydro-thermo-mechanical properties of the rock, which are often strong functions of organic content. This work demonstrates the potential of near-infrared (NIR) spectral imaging in mapping the spatial distribution of organic content with O(100µm) resolution on cores that can span several hundred feet in depth (Mehmani et al., 2017). We validate our approach for the immature oil shale of the Green River Formation (GRF), USA, and show its applicability potential in other formations. The method is a generalization of a previously developed optical approach specialized to the GRF (Mehmani et al., 2016a). The implications of this work for spatial mapping of hydro-thermo-mechanical properties of excavated cores, in particular thermal conductivity, are discussed (Mehmani et al., 2016b). References:Mehmani, Y., A.K. Burnham, M.D. Vanden Berg, H. Tchelepi, "Quantification of organic content in shales via near-infrared imaging: Green River Formation." Fuel, (2017). Mehmani, Y., A.K. Burnham, M.D. Vanden Berg, F. Gelin, and H. Tchelepi. "Quantification of kerogen content in organic-rich shales from optical photographs." Fuel, (2016a). Mehmani, Y., A.K. Burnham, H. Tchelepi, "From optics to upscaled thermal conductivity: Green River oil shale." Fuel, (2016b).
-
Absolute counting of neutrophils in whole blood using flow cytometry.
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.
-
Determination of Ploidy Level and Nuclear DNA Content in the Droseraceae by Flow Cytometry
Czech Academy of Sciences Publication Activity Database
Hoshi, Y.; Azumatani, M.; Suyama, T.; Adamec, Lubomír
2017-01-01
Roč. 82, č. 3 (2017), s. 321-327 ISSN 0011-4545 Institutional support: RVO:67985939 Keywords : nuclear DNA content * genome size * Droseraceae Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Plant sciences, botany Impact factor: 0.913, year: 2016
-
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.
-
Measurement and Characterization of Apoptosis by Flow Cytometry.
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.
-
Multiplex polymerase chain reaction on FTA cards vs. flow cytometry for B-lymphocyte clonality.
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.
-
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.
-
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.
-
High content live cell imaging for the discovery of new antimalarial marine natural products
Directory of Open Access Journals (Sweden)
Cervantes Serena
2012-01-01
Full Text Available Abstract Background The human malaria parasite remains a burden in developing nations. It is responsible for up to one million deaths a year, a number that could rise due to increasing multi-drug resistance to all antimalarial drugs currently available. Therefore, there is an urgent need for the discovery of new drug therapies. Recently, our laboratory developed a simple one-step fluorescence-based live cell-imaging assay to integrate the complex biology of the human malaria parasite into drug discovery. Here we used our newly developed live cell-imaging platform to discover novel marine natural products and their cellular phenotypic effects against the most lethal malaria parasite, Plasmodium falciparum. Methods A high content live cell imaging platform was used to screen marine extracts effects on malaria. Parasites were grown in vitro in the presence of extracts, stained with RNA sensitive dye, and imaged at timed intervals with the BD Pathway HT automated confocal microscope. Results Image analysis validated our new methodology at a larger scale level and revealed potential antimalarial activity of selected extracts with a minimal cytotoxic effect on host red blood cells. To further validate our assay, we investigated parasite's phenotypes when incubated with the purified bioactive natural product bromophycolide A. We show that bromophycolide A has a strong and specific morphological effect on parasites, similar to the ones observed from the initial extracts. Conclusion Collectively, our results show that high-content live cell-imaging (HCLCI can be used to screen chemical libraries and identify parasite specific inhibitors with limited host cytotoxic effects. All together we provide new leads for the discovery of novel antimalarials.
-
High content live cell imaging for the discovery of new antimalarial marine natural products.
Cervantes, Serena; Stout, Paige E; Prudhomme, Jacques; Engel, Sebastian; Bruton, Matthew; Cervantes, Michael; Carter, David; Tae-Chang, Young; Hay, Mark E; Aalbersberg, William; Kubanek, Julia; Le Roch, Karine G
2012-01-03
The human malaria parasite remains a burden in developing nations. It is responsible for up to one million deaths a year, a number that could rise due to increasing multi-drug resistance to all antimalarial drugs currently available. Therefore, there is an urgent need for the discovery of new drug therapies. Recently, our laboratory developed a simple one-step fluorescence-based live cell-imaging assay to integrate the complex biology of the human malaria parasite into drug discovery. Here we used our newly developed live cell-imaging platform to discover novel marine natural products and their cellular phenotypic effects against the most lethal malaria parasite, Plasmodium falciparum. A high content live cell imaging platform was used to screen marine extracts effects on malaria. Parasites were grown in vitro in the presence of extracts, stained with RNA sensitive dye, and imaged at timed intervals with the BD Pathway HT automated confocal microscope. Image analysis validated our new methodology at a larger scale level and revealed potential antimalarial activity of selected extracts with a minimal cytotoxic effect on host red blood cells. To further validate our assay, we investigated parasite's phenotypes when incubated with the purified bioactive natural product bromophycolide A. We show that bromophycolide A has a strong and specific morphological effect on parasites, similar to the ones observed from the initial extracts. Collectively, our results show that high-content live cell-imaging (HCLCI) can be used to screen chemical libraries and identify parasite specific inhibitors with limited host cytotoxic effects. All together we provide new leads for the discovery of novel antimalarials. © 2011 Cervantes et al; licensee BioMed Central Ltd.
-
Directory of Open Access Journals (Sweden)
Tiago Severo Garcia
Full Text Available Imaging studies are expected to produce reliable information regarding the size and fat content of the pancreas. However, the available studies have produced inconclusive results. The aim of this study was to perform a systematic review and meta-analysis of imaging studies assessing pancreas size and fat content in patients with type 1 diabetes (T1DM and type 2 diabetes (T2DM.Medline and Embase databases were performed. Studies evaluating pancreatic size (diameter, area or volume and/or fat content by ultrasound, computed tomography, or magnetic resonance imaging in patients with T1DM and/or T2DM as compared to healthy controls were selected. Seventeen studies including 3,403 subjects (284 T1DM patients, 1,139 T2DM patients, and 1,980 control subjects were selected for meta-analyses. Pancreas diameter, area, volume, density, and fat percentage were evaluated.Pancreatic volume was reduced in T1DM and T2DM vs. controls (T1DM vs. controls: -38.72 cm3, 95%CI: -52.25 to -25.19, I2 = 70.2%, p for heterogeneity = 0.018; and T2DM vs. controls: -12.18 cm3, 95%CI: -19.1 to -5.25, I2 = 79.3%, p for heterogeneity = 0.001. Fat content was higher in T2DM vs. controls (+2.73%, 95%CI 0.55 to 4.91, I2 = 82.0%, p for heterogeneity<0.001.Individuals with T1DM and T2DM have reduced pancreas size in comparison with control subjects. Patients with T2DM have increased pancreatic fat content.
-
International Nuclear Information System (INIS)
Carita, Edilson Carlos; Seraphim, Enzo; Honda, Marcelo Ossamu; Azevedo-Marques, Paulo Mazzoncini de
2008-01-01
Objective: the present paper describes the implementation and evaluation of a medical images management system with content-based retrieval support (PACS-CBIR) integrating modules focused on images acquisition, storage and distribution, and text retrieval by keyword and images retrieval by similarity. Materials and methods: internet-compatible technologies were utilized for the system implementation with free ware, and C ++ , PHP and Java languages on a Linux platform. There is a DICOM-compatible image management module and two query modules, one of them based on text and the other on similarity of image texture attributes. Results: results demonstrate an appropriate images management and storage, and that the images retrieval time, always < 15 sec, was found to be good by users. The evaluation of retrieval by similarity has demonstrated that the selected images extractor allowed the sorting of images according to anatomical areas. Conclusion: based on these results, one can conclude that the PACS-CBIR implementation is feasible. The system has demonstrated to be DICOM-compatible, and that it can be integrated with the local information system. The similar images retrieval functionality can be enhanced by the introduction of further descriptors. (author)
-
DNA content analysis allows discrimination between Trypanosoma cruzi and Trypanosoma rangeli.
Naves, Lucila Langoni; da Silva, Marcos Vinícius; Fajardo, Emanuella Francisco; da Silva, Raíssa Bernardes; De Vito, Fernanda Bernadelli; Rodrigues, Virmondes; Lages-Silva, Eliane; Ramírez, Luis Eduardo; Pedrosa, André Luiz
2017-01-01
Trypanosoma cruzi, a human protozoan parasite, is the causative agent of Chagas disease. Currently the species is divided into six taxonomic groups. The genome of the CL Brener clone has been estimated to be 106.4-110.7 Mb, and DNA content analyses revealed that it is a diploid hybrid clone. Trypanosoma rangeli is a hemoflagellate that has the same reservoirs and vectors as T. cruzi; however, it is non-pathogenic to vertebrate hosts. The haploid genome of T. rangeli was previously estimated to be 24 Mb. The parasitic strains of T. rangeli are divided into KP1(+) and KP1(-). Thus, the objective of this study was to investigate the DNA content in different strains of T. cruzi and T. rangeli by flow cytometry. All T. cruzi and T. rangeli strains yielded cell cycle profiles with clearly identifiable G1-0 (2n) and G2-M (4n) peaks. T. cruzi and T. rangeli genome sizes were estimated using the clone CL Brener and the Leishmania major CC1 as reference cell lines because their genome sequences have been previously determined. The DNA content of T. cruzi strains ranged from 87,41 to 108,16 Mb, and the DNA content of T. rangeli strains ranged from 63,25 Mb to 68,66 Mb. No differences in DNA content were observed between KP1(+) and KP1(-) T. rangeli strains. Cultures containing mixtures of the epimastigote forms of T. cruzi and T. rangeli strains resulted in cell cycle profiles with distinct G1 peaks for strains of each species. These results demonstrate that DNA content analysis by flow cytometry is a reliable technique for discrimination between T. cruzi and T. rangeli isolated from different hosts.
-
Liu, Changhong; Liu, Wei; Chen, Wei; Yang, Jianbo; Zheng, Lei
2015-04-15
Tomato is an important health-stimulating fruit because of the antioxidant properties of its main bioactive compounds, dominantly lycopene and phenolic compounds. Nowadays, product differentiation in the fruit market requires an accurate evaluation of these value-added compounds. An experiment was conducted to simultaneously and non-destructively measure lycopene and phenolic compounds content in intact tomatoes using multispectral imaging combined with chemometric methods. Partial least squares (PLS), least squares-support vector machines (LS-SVM) and back propagation neural network (BPNN) were applied to develop quantitative models. Compared with PLS and LS-SVM, BPNN model considerably improved the performance with coefficient of determination in prediction (RP(2))=0.938 and 0.965, residual predictive deviation (RPD)=4.590 and 9.335 for lycopene and total phenolics content prediction, respectively. It is concluded that multispectral imaging is an attractive alternative to the standard methods for determination of bioactive compounds content in intact tomatoes, providing a useful platform for infield fruit sorting/grading. Copyright © 2014 Elsevier Ltd. All rights reserved.
-
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.
-
Multisite Thrombus Imaging and Fibrin Content Estimation With a Single Whole-Body PET Scan in Rats.
Blasi, Francesco; Oliveira, Bruno L; Rietz, Tyson A; Rotile, Nicholas J; Naha, Pratap C; Cormode, David P; Izquierdo-Garcia, David; Catana, Ciprian; Caravan, Peter
2015-10-01
Thrombosis is a leading cause of morbidity and mortality worldwide. Current diagnostic strategies rely on imaging modalities that are specific for distinct vascular territories, but a thrombus-specific whole-body imaging approach is still missing. Moreover, imaging techniques to assess thrombus composition are underdeveloped, although therapeutic strategies may benefit from such technology. Therefore, our goal was to test whether positron emission tomography (PET) with the fibrin-binding probe (64)Cu-FBP8 allows multisite thrombus detection and fibrin content estimation. Thrombosis was induced in Sprague-Dawley rats (n=32) by ferric chloride application on both carotid artery and femoral vein. (64)Cu-FBP8-PET/CT imaging was performed 1, 3, or 7 days after thrombosis to detect thrombus location and to evaluate age-dependent changes in target uptake. Ex vivo biodistribution, autoradiography, and histopathology were performed to validate imaging results. Arterial and venous thrombi were localized on fused PET/CT images with high accuracy (97.6%; 95% confidence interval, 92-100). A single whole-body PET/MR imaging session was sufficient to reveal the location of both arterial and venous thrombi after (64)Cu-FBP8 administration. PET imaging showed that probe uptake was greater in younger clots than in older ones for both arterial and venous thrombosis (P<0.0001). Quantitative histopathology revealed an age-dependent reduction of thrombus fibrin content (P<0.001), consistent with PET results. Biodistribution and autoradiography further confirmed the imaging findings. We demonstrated that (64)Cu-FBP8-PET is a feasible approach for whole-body thrombus detection and that molecular imaging of fibrin can provide, noninvasively, insight into clot composition. © 2015 American Heart Association, Inc.
-
Coconut genome size determined by flow cytometry: Tall versus Dwarf types.
Freitas Neto, M; Pereira, T N S; Geronimo, I G C; Azevedo, A O N; Ramos, S R R; Pereira, M G
2016-02-11
Coconuts (Cocos nucifera L.) are tropical palm trees that are classified into Tall and Dwarf types based on height, and both types are diploid (2n = 2x = 32 chromosomes). The reproduction mode is autogamous for Dwarf types and allogamous for Tall types. One hypothesis for the origin of the Dwarf coconut suggests that it is a Tall variant that resulted from either mutation or inbreeding, and differences in genome size between the two types would support this hypothesis. In this study, we estimated the genome sizes of 14 coconut accessions (eight Tall and six Dwarf types) using flow cytometry. Nuclei were extracted from leaf discs and stained with propidium iodide, and Pisum sativum (2C = 9.07 pg DNA) was used as an internal standard. Histograms with good resolution and low coefficients of variation (2.5 to 3.2%) were obtained. The 2C DNA content ranged from 5.72 to 5.48 pg for Tall accessions and from 5.58 to 5.52 pg for Dwarf accessions. The mean genome sizes for Tall and Dwarf specimens were 5.59 and 5.55 pg, respectively. Among all accessions, Rennel Island Tall had the highest mean DNA content (5.72 pg), whereas West African Tall had the lowest (5.48 pg). The mean coconut genome size (2C = 5.57 pg, corresponding to 2723.73 Mbp/haploid set) was classified as small. Only small differences in genome size existed among the coconut accessions, suggesting that the Dwarf type did not evolve from the Tall type.
-
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.
-
Biology and flow cytometry of proangiogenic hematopoietic progenitors cells.
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.
-
Content-based image retrieval applied to bone age assessment
Fischer, Benedikt; Brosig, André; Welter, Petra; Grouls, Christoph; Günther, Rolf W.; Deserno, Thomas M.
2010-03-01
Radiological bone age assessment is based on local image regions of interest (ROI), such as the epiphysis or the area of carpal bones. These are compared to a standardized reference and scores determining the skeletal maturity are calculated. For computer-aided diagnosis, automatic ROI extraction and analysis is done so far mainly by heuristic approaches. Due to high variations in the imaged biological material and differences in age, gender and ethnic origin, automatic analysis is difficult and frequently requires manual interactions. On the contrary, epiphyseal regions (eROIs) can be compared to previous cases with known age by content-based image retrieval (CBIR). This requires a sufficient number of cases with reliable positioning of the eROI centers. In this first approach to bone age assessment by CBIR, we conduct leaving-oneout experiments on 1,102 left hand radiographs and 15,428 metacarpal and phalangeal eROIs from the USC hand atlas. The similarity of the eROIs is assessed by cross-correlation of 16x16 scaled eROIs. The effects of the number of eROIs, two age computation methods as well as the number of considered CBIR references are analyzed. The best results yield an error rate of 1.16 years and a standard deviation of 0.85 years. As the appearance of the hand varies naturally by up to two years, these results clearly demonstrate the applicability of the CBIR approach for bone age estimation.
-
Lipid nanoparticles assessment by flow cytometry.
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.
-
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
-
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.
-
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...
-
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
-
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.
-
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.
-
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.
-
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
-
Liang, Ru-Ze
2017-04-24
In this paper we study the problem of content-based image retrieval. In this problem, the most popular performance measure is the top precision measure, and the most important component of a retrieval system is the similarity function used to compare a query image against a database image. However, up to now, there is no existing similarity learning method proposed to optimize the top precision measure. To fill this gap, in this paper, we propose a novel similarity learning method to maximize the top precision measure. We model this problem as a minimization problem with an objective function as the combination of the losses of the relevant images ranked behind the top-ranked irrelevant image, and the squared Frobenius norm of the similarity function parameter. This minimization problem is solved as a quadratic programming problem. The experiments over two benchmark data sets show the advantages of the proposed method over other similarity learning methods when the top precision is used as the performance measure.
-
Liang, Ru-Ze; Shi, Lihui; Wang, Haoxiang; Meng, Jiandong; Wang, Jim Jing-Yan; Sun, Qingquan; Gu, Yi
2017-01-01
In this paper we study the problem of content-based image retrieval. In this problem, the most popular performance measure is the top precision measure, and the most important component of a retrieval system is the similarity function used to compare a query image against a database image. However, up to now, there is no existing similarity learning method proposed to optimize the top precision measure. To fill this gap, in this paper, we propose a novel similarity learning method to maximize the top precision measure. We model this problem as a minimization problem with an objective function as the combination of the losses of the relevant images ranked behind the top-ranked irrelevant image, and the squared Frobenius norm of the similarity function parameter. This minimization problem is solved as a quadratic programming problem. The experiments over two benchmark data sets show the advantages of the proposed method over other similarity learning methods when the top precision is used as the performance measure.
-
Quantifying the margin sharpness of lesions on radiological images for content-based image retrieval
International Nuclear Information System (INIS)
Xu Jiajing; Napel, Sandy; Greenspan, Hayit; Beaulieu, Christopher F.; Agrawal, Neeraj; Rubin, Daniel
2012-01-01
. Equivalence across deformations was assessed using Schuirmann's paired two one-sided tests. Results: In simulated images, the concordance correlation between measured gradient and actual gradient was 0.994. The mean (s.d.) and standard deviation NDCG score for the retrieval of K images, K = 5, 10, and 15, were 84% (8%), 85% (7%), and 85% (7%) for CT images containing liver lesions, and 82% (7%), 84% (6%), and 85% (4%) for CT images containing lung nodules, respectively. The authors’ proposed method outperformed the two existing margin characterization methods in average NDCG scores over all K, by 1.5% and 3% in datasets containing liver lesion, and 4.5% and 5% in datasets containing lung nodules. Equivalence testing showed that the authors’ feature is more robust across all margin deformations (p < 0.05) than the two existing methods for margin sharpness characterization in both simulated and clinical datasets. Conclusions: The authors have described a new image feature to quantify the margin sharpness of lesions. It has strong correlation with known margin sharpness in simulated images and in clinical CT images containing liver lesions and lung nodules. This image feature has excellent performance for retrieving images with similar margin characteristics, suggesting potential utility, in conjunction with other lesion features, for content-based image retrieval applications.
-
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.
-
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.
-
McDonough, Patrick M; Prigozhina, Natalie L; Basa, Ranor C B; Price, Jeffrey H
2017-07-01
Postchemotherapy cognitive impairment (PCCI) is commonly exhibited by cancer patients treated with a variety of chemotherapeutic agents, including the endocrine disruptor tamoxifen (TAM). The etiology of PCCI is poorly understood. Our goal was to develop high-throughput assay methods to test the effects of chemicals on neuronal function applicable to PCCI. Rat hippocampal neurons (RHNs) were plated in 96- or 384-well dishes and exposed to test compounds (forskolin [FSK], 17β-estradiol [ES]), TAM or fulvestrant [FUL], aka ICI 182,780) for 6-14 days. Kinetic Image Cytometry™ (KIC™) methods were developed to quantify spontaneously occurring intracellular calcium transients representing the activity of the neurons, and high-content analysis (HCA) methods were developed to quantify the expression, colocalization, and puncta formed by synaptic proteins (postsynaptic density protein-95 [PSD-95] and presynaptic protein Synapsin-1 [Syn-1]). As quantified by KIC, FSK increased the occurrence and synchronization of the calcium transients indicating stimulatory effects on RHN activity, whereas TAM had inhibitory effects. As quantified by HCA, FSK also increased PSD-95 puncta and PSD-95:Syn-1 colocalization, whereas ES increased the puncta of both PSD-95 and Syn-1 with little effect on colocalization. The estrogen receptor antagonist FUL also increased PSD-95 puncta. In contrast, TAM reduced Syn-1 and PSD-95:Syn-1 colocalization, consistent with its inhibitory effects on the calcium transients. Thus TAM reduced activity and synapse formation by the RHNs, which may relate to the ability of this agent to cause PCCI. The results illustrate that KIC and HCA can be used to quantify neurotoxic and neuroprotective effects of chemicals in RHNs to investigate mechanisms and potential therapeutics for PCCI.
-
Bravo-Zanoguera, Miguel E; Laris, Casey A; Nguyen, Lam K; Oliva, Mike; Price, Jeffrey H
2007-01-01
Efficient image cytometry of a conventional microscope slide means rapid acquisition and analysis of 20 gigapixels of image data (at 0.3-microm sampling). The voluminous data motivate increased acquisition speed to enable many biomedical applications. Continuous-motion time-delay-and-integrate (TDI) scanning has the potential to speed image acquisition while retaining sensitivity, but the challenge of implementing high-resolution autofocus operating simultaneously with acquisition has limited its adoption. We develop a dynamic autofocus system for this need using: 1. a "volume camera," consisting of nine fiber optic imaging conduits to charge-coupled device (CCD) sensors, that acquires images in parallel from different focal planes, 2. an array of mixed analog-digital processing circuits that measure the high spatial frequencies of the multiple image streams to create focus indices, and 3. a software system that reads and analyzes the focus data streams and calculates best focus for closed feedback loop control. Our system updates autofocus at 56 Hz (or once every 21 microm of stage travel) to collect sharply focused images sampled at 0.3x0.3 microm(2)/pixel at a stage speed of 2.3 mms. The system, tested by focusing in phase contrast and imaging long fluorescence strips, achieves high-performance closed-loop image-content-based autofocus in continuous scanning for the first time.
-
Directory of Open Access Journals (Sweden)
M Sedghi
2012-03-01
Full Text Available The relationship between sorghum grain color and tannin content was reported in several references. In this study, 33 phenotypes of sorghum grain differing in seed characteristics were collected and analyzed by Folin-Ciocalteu method. A computer image analysis method was used to determine the color characteristics of all 33 sorghum phenotypes. Two methods of multiple linear regression and artificial neural network (ANN models were developed to describe tannin content in sorghum grain from three input parameters of color characteristics. The goodness of fit of the models was tested using R², MS error, and bias. The computer image analysis technique was a suitable method to estimate tannin through sorghum grain color strength. Therefore, the color quality of the samples was described according three color parameters: L* (lightness, a* (redness - from green to red and b* (blueness - from blue to yellow. The developed regression and ANN models showed a strong relationship between color and tannin content of samples. The goodness of fit (in terms of R², which corresponds to training the ANN model, showed higher accuracy of prediction of ANN compared with the equation established by the regression method (0.96 vs. 0.88. The ANN models in term of MS error showed lower residuals distribution than that of regression model (0.002 vs. 0.006. The platform of computer image analysis technique and ANN-based model may be used to estimate the tannin content of sorghum.
-
Chlorophyll content of Plešné Lake phytoplankton cells studied with image analysis
Czech Academy of Sciences Publication Activity Database
Nedoma, Jiří; Nedbalová, Linda
2006-01-01
Roč. 61, Suppl. 20 (2006), S491-S498 ISSN 0006-3088 Grant - others:MSMT(CZ) 0021620828 Institutional research plan: CEZ:AV0Z60170517; CEZ:AV0Z60050516 Keywords : cellular chlorophyll content * image analysis * vertical profile Subject RIV: EH - Ecology, Behaviour Impact factor: 0.213, year: 2006
-
Impact of image segmentation on high-content screening data quality for SK-BR-3 cells
Directory of Open Access Journals (Sweden)
Li Yizheng
2007-09-01
Full Text Available Abstract Background High content screening (HCS is a powerful method for the exploration of cellular signalling and morphology that is rapidly being adopted in cancer research. HCS uses automated microscopy to collect images of cultured cells. The images are subjected to segmentation algorithms to identify cellular structures and quantitate their morphology, for hundreds to millions of individual cells. However, image analysis may be imperfect, especially for "HCS-unfriendly" cell lines whose morphology is not well handled by current image segmentation algorithms. We asked if segmentation errors were common for a clinically relevant cell line, if such errors had measurable effects on the data, and if HCS data could be improved by automated identification of well-segmented cells. Results Cases of poor cell body segmentation occurred frequently for the SK-BR-3 cell line. We trained classifiers to identify SK-BR-3 cells that were well segmented. On an independent test set created by human review of cell images, our optimal support-vector machine classifier identified well-segmented cells with 81% accuracy. The dose responses of morphological features were measurably different in well- and poorly-segmented populations. Elimination of the poorly-segmented cell population increased the purity of DNA content distributions, while appropriately retaining biological heterogeneity, and simultaneously increasing our ability to resolve specific morphological changes in perturbed cells. Conclusion Image segmentation has a measurable impact on HCS data. The application of a multivariate shape-based filter to identify well-segmented cells improved HCS data quality for an HCS-unfriendly cell line, and could be a valuable post-processing step for some HCS datasets.
-
Flow Cytometry in Diagnosis of Myelomatous Pleural Effusion: A Case Report.
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.
-
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.
-
Routine detection of Epstein-Barr virus specific T-cells in the peripheral blood by flow cytometry
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
-
A content-based digital image watermarking scheme resistant to local geometric distortions
International Nuclear Information System (INIS)
Yang, Hong-ying; Chen, Li-li; Wang, Xiang-yang
2011-01-01
Geometric distortion is known as one of the most difficult attacks to resist, as it can desynchronize the location of the watermark and hence cause incorrect watermark detection. Geometric distortion can be decomposed into two classes: global affine transforms and local geometric distortions. Most countermeasures proposed in the literature only address the problem of global affine transforms. It is a challenging problem to design a robust image watermarking scheme against local geometric distortions. In this paper, we propose a new content-based digital image watermarking scheme with good visual quality and reasonable resistance against local geometric distortions. Firstly, the robust feature points, which can survive various common image processing and global affine transforms, are extracted by using a multi-scale SIFT (scale invariant feature transform) detector. Then, the affine covariant local feature regions (LFRs) are constructed adaptively according to the feature scale and local invariant centroid. Finally, the digital watermark is embedded into the affine covariant LFRs by modulating the magnitudes of discrete Fourier transform (DFT) coefficients. By binding the watermark with the affine covariant LFRs, the watermark detection can be done without synchronization error. Experimental results show that the proposed image watermarking is not only invisible and robust against common image processing operations such as sharpening, noise addition, and JPEG compression, etc, but also robust against global affine transforms and local geometric distortions
-
Piccinini, Filippo; Balassa, Tamas; Szkalisity, Abel; Molnar, Csaba; Paavolainen, Lassi; Kujala, Kaisa; Buzas, Krisztina; Sarazova, Marie; Pietiainen, Vilja; Kutay, Ulrike; Smith, Kevin; Horvath, Peter
2017-06-28
High-content, imaging-based screens now routinely generate data on a scale that precludes manual verification and interrogation. Software applying machine learning has become an essential tool to automate analysis, but these methods require annotated examples to learn from. Efficiently exploring large datasets to find relevant examples remains a challenging bottleneck. Here, we present Advanced Cell Classifier (ACC), a graphical software package for phenotypic analysis that addresses these difficulties. ACC applies machine-learning and image-analysis methods to high-content data generated by large-scale, cell-based experiments. It features methods to mine microscopic image data, discover new phenotypes, and improve recognition performance. We demonstrate that these features substantially expedite the training process, successfully uncover rare phenotypes, and improve the accuracy of the analysis. ACC is extensively documented, designed to be user-friendly for researchers without machine-learning expertise, and distributed as a free open-source tool at www.cellclassifier.org. Copyright © 2017 Elsevier Inc. All rights reserved.
-
Dhaliwal, Anandika; Brenner, Matthew; Wolujewicz, Paul; Zhang, Zheng; Mao, Yong; Batish, Mona; Kohn, Joachim; Moghe, Prabhas V
2016-11-01
A predictive framework for the evolution of stem cell biology in 3-D is currently lacking. In this study we propose deep image informatics of the nuclear biology of stem cells to elucidate how 3-D biomaterials steer stem cell lineage phenotypes. The approach is based on high content imaging informatics to capture minute variations in the 3-D spatial organization of splicing factor SC-35 in the nucleoplasm as a marker to classify emergent cell phenotypes of human mesenchymal stem cells (hMSCs). The cells were cultured in varied 3-D culture systems including hydrogels, electrospun mats and salt leached scaffolds. The approach encompasses high resolution 3-D imaging of SC-35 domains and high content image analysis (HCIA) to compute quantitative 3-D nuclear metrics for SC-35 organization in single cells in concert with machine learning approaches to construct a predictive cell-state classification model. Our findings indicate that hMSCs cultured in collagen hydrogels and induced to differentiate into osteogenic or adipogenic lineages could be classified into the three lineages (stem, adipogenic, osteogenic) with ⩾80% precision and sensitivity, within 72h. Using this framework, the augmentation of osteogenesis by scaffold design exerted by porogen leached scaffolds was also profiled within 72h with ∼80% high sensitivity. Furthermore, by employing 3-D SC-35 organizational metrics, differential osteogenesis induced by novel electrospun fibrous polymer mats incorporating decellularized matrix could also be elucidated and predictably modeled at just 3days with high precision. We demonstrate that 3-D SC-35 organizational metrics can be applied to model the stem cell state in 3-D scaffolds. We propose that this methodology can robustly discern minute changes in stem cell states within complex 3-D architectures and map single cell biological readouts that are critical to assessing population level cell heterogeneity. The sustained development and validation of bioactive
-
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
-
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.
-
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.
-
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.
-
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)
-
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.
-
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.
-
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)
-
Stochastic Measurement Models for Quantifying Lymphocyte Responses Using Flow Cytometry
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
-
Biomass measurement by flow cytometry during solid-state fermentation of basidiomycetes.
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.
-
Bezborodkina, Natalia N; Chestnova, Anna Yu; Vorobev, Mikhail L; Kudryavtsev, Boris N
2016-04-01
Hepatocytes differ from one another by the degree of the ploidy, size, position in the liver lobule, and level of the DNA-synthetic processes. It is believed, that the cell size exerts substantial influence on the metabolism of the hepatocytes and the glycogen content in them. The aim of the present study was to test this hypothesis. Dry weight of hepatocytes, their ploidy and glycogen content were determined in the normal and the cirrhotic rat liver. Liver cirrhosis in rats was produced by chronic inhalation of CCl4 vapours in the course of 6 months. A combined cytophotometric method was used. Dry weight of the cell, its glycogen and DNA content were successively measured on a mapped preparation. Hepatocytes of each ploidy class in the normal and the cirrhotic rat liver accumulated glycogen at the same rate. In the normal liver, there was a distinct correlation between the size of hepatocytes and glycogen content in them. This correlation was observed in each ploidy class, and was especially pronounced in the class of mononucleate tetraploid hepatocytes. In the cirrhotic liver, there was no correlation between the size of the cells and their glycogen content. The impairment of liver lobular structure probably explains the observed lack of correlation between hepatocyte size and their glycogen content in the cirrhotic liver. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.
-
A general method for bead-enhanced quantitation by flow cytometry
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
-
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
-
A multi-scale convolutional neural network for phenotyping high-content cellular images.
Godinez, William J; Hossain, Imtiaz; Lazic, Stanley E; Davies, John W; Zhang, Xian
2017-07-01
Identifying phenotypes based on high-content cellular images is challenging. Conventional image analysis pipelines for phenotype identification comprise multiple independent steps, with each step requiring method customization and adjustment of multiple parameters. Here, we present an approach based on a multi-scale convolutional neural network (M-CNN) that classifies, in a single cohesive step, cellular images into phenotypes by using directly and solely the images' pixel intensity values. The only parameters in the approach are the weights of the neural network, which are automatically optimized based on training images. The approach requires no a priori knowledge or manual customization, and is applicable to single- or multi-channel images displaying single or multiple cells. We evaluated the classification performance of the approach on eight diverse benchmark datasets. The approach yielded overall a higher classification accuracy compared with state-of-the-art results, including those of other deep CNN architectures. In addition to using the network to simply obtain a yes-or-no prediction for a given phenotype, we use the probability outputs calculated by the network to quantitatively describe the phenotypes. This study shows that these probability values correlate with chemical treatment concentrations. This finding validates further our approach and enables chemical treatment potency estimation via CNNs. The network specifications and solver definitions are provided in Supplementary Software 1. william_jose.godinez_navarro@novartis.com or xian-1.zhang@novartis.com. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com
-
Research Techniques Made Simple: Experimental Methodology for Single-Cell Mass Cytometry
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
-
Immune Response to Mycobacterial Infection: Lessons from Flow Cytometry
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...
-
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.
-
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.
-
Assessment of Abdominal Adipose Tissue and Organ Fat Content by Magnetic Resonance Imaging
Hu, Houchun H.; Nayak, Krishna S.; Goran, Michael I.
2010-01-01
As the prevalence of obesity continues to rise, rapid and accurate tools for assessing abdominal body and organ fat quantity and distribution are critically needed to assist researchers investigating therapeutic and preventive measures against obesity and its comorbidities. Magnetic resonance imaging (MRI) is the most promising modality to address such need. It is non-invasive, utilizes no ionizing radiation, provides unmatched 3D visualization, is repeatable, and is applicable to subject cohorts of all ages. This article is aimed to provide the reader with an overview of current and state-of-the-art techniques in MRI and associated image analysis methods for fat quantification. The principles underlying traditional approaches such as T1-weighted imaging and magnetic resonance spectroscopy as well as more modern chemical-shift imaging techniques are discussed and compared. The benefits of contiguous 3D acquisitions over 2D multi-slice approaches are highlighted. Typical post-processing procedures for extracting adipose tissue depot volumes and percent organ fat content from abdominal MRI data sets are explained. Furthermore, the advantages and disadvantages of each MRI approach with respect to imaging parameters, spatial resolution, subject motion, scan time, and appropriate fat quantitative endpoints are also provided. Practical considerations in implementing these methods are also presented. PMID:21348916
-
Artymovich, Katherine; Appledorn, Daniel M
2015-01-01
In vitro cell proliferation and apoptosis assays are widely used to study cancer cell biology. Commonly used methodologies are however performed at a single, user-defined endpoint. We describe a kinetic multiplex assay incorporating the CellPlayer(TM) NucLight Red reagent to measure proliferation and the CellPlayer(TM) Caspase-3/7 reagent to measure apoptosis using the two-color, live-content imaging platform, IncuCyte(TM) ZOOM. High-definition phase-contrast images provide an additional qualitative validation of cell death based on morphological characteristics. The kinetic data generated using this strategy can be used to derive informed pharmacology measurements to screen potential cancer therapeutics.
-
Transfer, imaging, and analysis plate for facile handling of 384 hanging drop 3D tissue spheroids.
Cavnar, Stephen P; Salomonsson, Emma; Luker, Kathryn E; Luker, Gary D; Takayama, Shuichi
2014-04-01
Three-dimensional culture systems bridge the experimental gap between in vivo and in vitro physiology. However, nonstandardized formation and limited downstream adaptability of 3D cultures have hindered mainstream adoption of these systems for biological applications, especially for low- and moderate-throughput assays commonly used in biomedical research. Here we build on our recent development of a 384-well hanging drop plate for spheroid culture to design a complementary spheroid transfer and imaging (TRIM) plate. The low-aspect ratio wells of the TRIM plate facilitated high-fidelity, user-independent, contact-based collection of hanging drop spheroids. Using the TRIM plate, we demonstrated several downstream analyses, including bulk tissue collection for flow cytometry, high-resolution low working-distance immersion imaging, and timely reagent delivery for enzymatic studies. Low working-distance multiphoton imaging revealed a cell type-dependent, macroscopic spheroid structure. Unlike ovarian cancer spheroids, which formed loose, disk-shaped spheroids, human mammary fibroblasts formed tight, spherical, and nutrient-limited spheroids. Beyond the applications we describe here, we expect the hanging drop spheroid plate and complementary TRIM plate to facilitate analyses of spheroids across the spectrum of throughput, particularly for bulk collection of spheroids and high-content imaging.
-
DEFF Research Database (Denmark)
Grønholdt, Marie-Louise M.; Nordestgaard, Børge; Weibe, Britt M.
1997-01-01
Relatioin between low gray scale values in computerized images of carotid plaques and 1) plasma levels of triglyceride-rich lipoproteins and 2) plaque lipid content......Relatioin between low gray scale values in computerized images of carotid plaques and 1) plasma levels of triglyceride-rich lipoproteins and 2) plaque lipid content...
-
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
-
Hoffman, Ewelina; Patel, Aateka; Ball, Doug; Klapwijk, Jan; Millar, Val; Kumar, Abhinav; Martin, Abigail; Mahendran, Rhamiya; Dailey, Lea Ann; Forbes, Ben; Hutter, Victoria
2017-12-01
Progress to the clinic may be delayed or prevented when vacuolated or "foamy" alveolar macrophages are observed during non-clinical inhalation toxicology assessment. The first step in developing methods to study this response in vitro is to characterize macrophage cell lines and their response to drug exposures. Human (U937) and rat (NR8383) cell lines and primary rat alveolar macrophages obtained by bronchoalveolar lavage were characterized using high content fluorescence imaging analysis quantification of cell viability, morphometry, and phospholipid and neutral lipid accumulation. Cell health, morphology and lipid content were comparable (p content. Responses to amiodarone, a known inducer of phospholipidosis, required analysis of shifts in cell population profiles (the proportion of cells with elevated vacuolation or lipid content) rather than average population data which was insensitive to the changes observed. A high content image analysis assay was developed and used to provide detailed morphological characterization of rat and human alveolar-like macrophages and their response to a phospholipidosis-inducing agent. This provides a basis for development of assays to predict or understand macrophage vacuolation following inhaled drug exposure.
-
Czech Academy of Sciences Publication Activity Database
Loureiro, J.; Rodriguez, E.; Doležel, Jaroslav; Santos, C.
2006-01-01
Roč. 98, - (2006), s. 515-527 ISSN 0305-7364 R&D Projects: GA MŠk(CZ) LC06004 Institutional research plan: CEZ:AV0Z50380511 Keywords : genome size * flow cytometry * nuclear DNA content Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.448, year: 2006
-
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.
-
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,
-
External quality assessment in flow cytometry: educational aspects and trends toward improvement
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
-
Analyse de données de cytometrie de flux pour un grand nombre d'échantillons
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...
-
Detection of circulating breast cancer cells using photoacoustic flow cytometry
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.
-
Riboflavin content in autofluorescent earthworm coelomocytes is species-specific.
Directory of Open Access Journals (Sweden)
Joanna Homa
2007-01-01
Full Text Available We have recently shown that a large proproportion of earthworm coelomocytes exhibit strong autofluorescence in some species (Dendrobaena veneta, Allolobophora chlorotica, Dendrodrilus rubidus, Eisenia fetida, and Octolasion spp., while autofluorescent coelomocytes are very scarce in representatives of Lumbricus spp. and Aporrectodea spp. Riboflavin (vitamin B2 was identified as a major fluorophore in Eisenia jetida coelomocytes. The main aim of the present experiments was to quantify riboflavin content in autofluorescent coelomocytes (eleocytes from several earthworm species through a combination of flow cytometric and spectrofluorometric measurements. Spectrofluorometry of coelomocyte lysates showed that riboflavin was non-detectable in the coelomocytes of Aporrectodea spp. and Lumbricus spp., but was a prominent constituent of lysates from species with autofluorescent eleocytes. In the latter case, riboflavin content was the highest in E. fetida, followed by Octolasion spp. > A. chlorotica > D. rubidus. The riboflavin content of coelomocytes correlates positively with eleocyte autofluorescence intensity measured by flow cytometry and visible with fluorescence microscopy.
-
Ubiquitous picture-rich content representation
Wang, Wiley; Dean, Jennifer; Muzzolini, Russ
2010-02-01
The amount of digital images taken by the average consumer is consistently increasing. People enjoy the convenience of storing and sharing their pictures through online (digital) and offline (traditional) media. A set of pictures can be uploaded to: online photo services, web blogs and social network websites. Alternatively, these images can be used to generate: prints, cards, photo books or other photo products. Through uploading and sharing, images are easily transferred from one format to another. And often, a different set of associated content (text, tags) is created across formats. For example, on his web blog, a user may journal his experiences of his recent travel; on his social network website, his friends tag and comment on the pictures; in his online photo album, some pictures are titled and keyword-tagged. When the user wants to tell a complete story, perhaps in a photo book, he must collect, across all formats: the pictures, writings and comments, etc. and organize them in a book format. The user has to arrange the content of his trip in each format. The arrangement, the associations between the images, tags, keywords and text, cannot be shared with other formats. In this paper, we propose a system that allows the content to be easily created and shared across various digital media formats. We define a uniformed data association structure to connect: images, documents, comments, tags, keywords and other data. This content structure allows the user to switch representation formats without reediting. The framework under each format can emphasize (display or hide) content elements based on preference. For example, a slide show view will emphasize the display of pictures with limited text; a blog view will display highlighted images and journal text; and the photo book will try to fit in all images and text content. In this paper, we will discuss the strategy to associate pictures with text content, so that it can naturally tell a story. We will also list
-
In vitro flow cytometry-based screening platform for cellulase engineering
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
-
Determining ice water content from 2D crystal images in convective cloud systems
Leroy, Delphine; Coutris, Pierre; Fontaine, Emmanuel; Schwarzenboeck, Alfons; Strapp, J. Walter
2016-04-01
Cloud microphysical in-situ instrumentation measures bulk parameters like total water content (TWC) and/or derives particle size distributions (PSD) (utilizing optical spectrometers and optical array probes (OAP)). The goal of this work is to introduce a comprehensive methodology to compute TWC from OAP measurements, based on the dataset collected during recent HAIC (High Altitude Ice Crystals)/HIWC (High Ice Water Content) field campaigns. Indeed, the HAIC/HIWC field campaigns in Darwin (2014) and Cayenne (2015) provide a unique opportunity to explore the complex relationship between cloud particle mass and size in ice crystal environments. Numerous mesoscale convective systems (MCSs) were sampled with the French Falcon 20 research aircraft at different temperature levels from -10°C up to 50°C. The aircraft instrumentation included an IKP-2 (isokinetic probe) to get reliable measurements of TWC and the optical array probes 2D-S and PIP recording images over the entire ice crystal size range. Based on the known principle relating crystal mass and size with a power law (m=α•Dβ), Fontaine et al. (2014) performed extended 3D crystal simulations and thereby demonstrated that it is possible to estimate the value of the exponent β from OAP data, by analyzing the surface-size relationship for the 2D images as a function of time. Leroy et al. (2015) proposed an extended version of this method that produces estimates of β from the analysis of both the surface-size and perimeter-size relationships. Knowing the value of β, α then is deduced from the simultaneous IKP-2 TWC measurements for the entire HAIC/HIWC dataset. The statistical analysis of α and β values for the HAIC/HIWC dataset firstly shows that α is closely linked to β and that this link changes with temperature. From these trends, a generalized parameterization for α is proposed. Finally, the comparison with the initial IKP-2 measurements demonstrates that the method is able to predict TWC values
-
Content metamorphosis in synthetic holography
International Nuclear Information System (INIS)
Desbiens, Jacques
2013-01-01
A synthetic hologram is an optical system made of hundreds of images amalgamated in a structure of holographic cells. Each of these images represents a point of view on a three-dimensional space which makes us consider synthetic holography as a multiple points of view perspective system. In the composition of a computer graphics scene for a synthetic hologram, the field of view of the holographic image can be divided into several viewing zones. We can attribute these divisions to any object or image feature independently and operate different transformations on image content. In computer generated holography, we tend to consider content variations as a continuous animation much like a short movie. However, by composing sequential variations of image features in relation with spatial divisions, we can build new narrative forms distinct from linear cinematographic narration. When observers move freely and change their viewing positions, they travel from one field of view division to another. In synthetic holography, metamorphoses of image content are within the observer's path. In all imaging Medias, the transformation of image features in synchronisation with the observer's position is a rare occurrence. However, this is a predominant characteristic of synthetic holography. This paper describes some of my experimental works in the development of metamorphic holographic images.
-
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
-
A roughly mapped terra incognita: Image of the child in adult-oriented media contents
Directory of Open Access Journals (Sweden)
Korać Nada M.
2003-01-01
Full Text Available The study analyzes the image of the child in the media contents intended for adult audiences in Serbia, considering the importance of the role media play in shaping public opinion on children, as well as the influence of such public opinion on adults' attitudes, decisions and actions concerning children. The study focuses on visibility and portrayal of children in the media, in order to determine to what extent children are present in them, and in what way. Relevant data were collected for three media - press, radio and television - mostly covering the entire territory of Serbia, over two consecutive months (April - May 2001. Content analysis revealed that children are not only underrepresented, but also misrepresented, in Serbian media.
-
Thorson, Esther; And Others
1991-01-01
This study analyzed four characteristics of political commercials to determine their impact on television viewers' reactions: (1) issue versus image strategies; (2) attack versus support appeals; (3) presence and absence of music; and (4) visual content, either with families or in professional campaign settings. Memory measures and attitudes are…
-
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.
-
Keyframes Global Map Establishing Method for Robot Localization through Content-Based Image Matching
Directory of Open Access Journals (Sweden)
Tianyang Cao
2017-01-01
Full Text Available Self-localization and mapping are important for indoor mobile robot. We report a robust algorithm for map building and subsequent localization especially suited for indoor floor-cleaning robots. Common methods, for example, SLAM, can easily be kidnapped by colliding or disturbed by similar objects. Therefore, keyframes global map establishing method for robot localization in multiple rooms and corridors is needed. Content-based image matching is the core of this method. It is designed for the situation, by establishing keyframes containing both floor and distorted wall images. Image distortion, caused by robot view angle and movement, is analyzed and deduced. And an image matching solution is presented, consisting of extraction of overlap regions of keyframes extraction and overlap region rebuild through subblocks matching. For improving accuracy, ceiling points detecting and mismatching subblocks checking methods are incorporated. This matching method can process environment video effectively. In experiments, less than 5% frames are extracted as keyframes to build global map, which have large space distance and overlap each other. Through this method, robot can localize itself by matching its real-time vision frames with our keyframes map. Even with many similar objects/background in the environment or kidnapping robot, robot localization is achieved with position RMSE <0.5 m.
-
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...
-
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.
-
Vreugdenburg, Thomas D; Laurence, Caroline O; Willis, Cameron D; Mundy, Linda; Hiller, Janet E
2014-09-01
To describe the nature and frequency of information presented on direct-to-consumer websites for emerging breast cancer imaging devices. Content analysis of Australian website advertisements from 2 March 2011 to 30 March 2012, for three emerging breast cancer imaging devices: digital infrared thermal imaging, electrical impedance scanning and electronic palpation imaging. Type of imaging offered, device safety, device performance, application of device, target population, supporting evidence and comparator tests. Thirty-nine unique Australian websites promoting a direct-to-consumer breast imaging device were identified. Despite a lack of supporting evidence, 22 websites advertised devices for diagnosis, 20 advertised devices for screening, 13 advertised devices for prevention and 13 advertised devices for identifying breast cancer risk factors. Similarly, advertised ranges of diagnostic sensitivity (78%-99%) and specificity (44%-91%) were relatively high compared with published literature. Direct comparisons with conventional screening tools that favoured the new device were highly prominent (31 websites), and one-third of websites (12) explicitly promoted their device as a suitable alternative. Australian websites for emerging breast imaging devices, which are also available internationally, promote the use of such devices as safe and effective solutions for breast cancer screening and diagnosis in a range of target populations. Many of these claims are not supported by peer-reviewed evidence, raising questions about the manner in which these devices and their advertising material are regulated, particularly when they are promoted as direct alternatives to established screening interventions.
-
Novel high-throughput cell-based hybridoma screening methodology using the Celigo Image Cytometer.
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.
-
Conturo, Thomas Edward
Tissue blood flow, blood content, and water state have been characterized in-situ with new nuclear magnetic resonance imaging techniques. The sensitivities of standard techniques to the physiologic tissue parameters spin density (N_{rm r}) and relaxation times (T_1 and T_2 ) are mathematically defined. A new driven inversion method is developed so that tissue T_1 and T_2 changes produce cooperative intensity changes, yielding high contrast, high signal to noise, and sensitivity to a wider range of tissue parameters. The actual tissue parameters were imaged by automated collection of multiple-echo data having multiple T _1 dependence. Data are simultaneously fit by three-parameters to a closed-form expression, producing lower inter-parameter correlation and parameter noise than in separate T_1 or T_2 methods or pre-averaged methods. Accurate parameters are obtained at different field strengths. Parametric images of pathology demonstrate high sensitivity to tissue heterogeneity, and water content is determined in many tissues. Erythrocytes were paramagnetically labeled to study blood content and relaxation mechanisms. Liver and spleen relaxation were enhanced following 10% exchange of animal blood volumes. Rapid water exchange between intracellular and extracellular compartments was validated. Erythrocytes occupied 12.5% of renal cortex volume, and blood content was uniform in the liver, spleen and kidney. The magnitude and direction of flow velocity was then imaged. To eliminate directional artifacts, a bipolar gradient technique sensitized to flow in different directions was developed. Phase angle was reconstructed instead of intensity since the former has a 2pi -fold higher dynamic range. Images of flow through curves demonstrated secondary flow with a centrifugally-biased laminar profile and stationary velocity peaks along the curvature. Portal vein flow velocities were diminished or reversed in cirrhosis. Image artifacts have been characterized and removed. The
-
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
-
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.
-
TBIdoc: 3D content-based CT image retrieval system for traumatic brain injury
Li, Shimiao; Gong, Tianxia; Wang, Jie; Liu, Ruizhe; Tan, Chew Lim; Leong, Tze Yun; Pang, Boon Chuan; Lim, C. C. Tchoyoson; Lee, Cheng Kiang; Tian, Qi; Zhang, Zhuo
2010-03-01
Traumatic brain injury (TBI) is a major cause of death and disability. Computed Tomography (CT) scan is widely used in the diagnosis of TBI. Nowadays, large amount of TBI CT data is stacked in the hospital radiology department. Such data and the associated patient information contain valuable information for clinical diagnosis and outcome prediction. However, current hospital database system does not provide an efficient and intuitive tool for doctors to search out cases relevant to the current study case. In this paper, we present the TBIdoc system: a content-based image retrieval (CBIR) system which works on the TBI CT images. In this web-based system, user can query by uploading CT image slices from one study, retrieval result is a list of TBI cases ranked according to their 3D visual similarity to the query case. Specifically, cases of TBI CT images often present diffuse or focal lesions. In TBIdoc system, these pathological image features are represented as bin-based binary feature vectors. We use the Jaccard-Needham measure as the similarity measurement. Based on these, we propose a 3D similarity measure for computing the similarity score between two series of CT slices. nDCG is used to evaluate the system performance, which shows the system produces satisfactory retrieval results. The system is expected to improve the current hospital data management in TBI and to give better support for the clinical decision-making process. It may also contribute to the computer-aided education in TBI.
-
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.
-
IMAGE DESCRIPTIONS FOR SKETCH BASED IMAGE RETRIEVAL
SAAVEDRA RONDO, JOSE MANUEL; SAAVEDRA RONDO, JOSE MANUEL
2008-01-01
Due to the massive use of Internet together with the proliferation of media devices, content based image retrieval has become an active discipline in computer science. A common content based image retrieval approach requires that the user gives a regular image (e.g, a photo) as a query. However, having a regular image as query may be a serious problem. Indeed, people commonly use an image retrieval system because they do not count on the desired image. An easy alternative way t...
-
Computer-aided diagnostics of screening mammography using content-based image retrieval
Deserno, Thomas M.; Soiron, Michael; de Oliveira, Júlia E. E.; de A. Araújo, Arnaldo
2012-03-01
Breast cancer is one of the main causes of death among women in occidental countries. In the last years, screening mammography has been established worldwide for early detection of breast cancer, and computer-aided diagnostics (CAD) is being developed to assist physicians reading mammograms. A promising method for CAD is content-based image retrieval (CBIR). Recently, we have developed a classification scheme of suspicious tissue pattern based on the support vector machine (SVM). In this paper, we continue moving towards automatic CAD of screening mammography. The experiments are based on in total 10,509 radiographs that have been collected from different sources. From this, 3,375 images are provided with one and 430 radiographs with more than one chain code annotation of cancerous regions. In different experiments, this data is divided into 12 and 20 classes, distinguishing between four categories of tissue density, three categories of pathology and in the 20 class problem two categories of different types of lesions. Balancing the number of images in each class yields 233 and 45 images remaining in each of the 12 and 20 classes, respectively. Using a two-dimensional principal component analysis, features are extracted from small patches of 128 x 128 pixels and classified by means of a SVM. Overall, the accuracy of the raw classification was 61.6 % and 52.1 % for the 12 and the 20 class problem, respectively. The confusion matrices are assessed for detailed analysis. Furthermore, an implementation of a SVM-based CBIR system for CADx in screening mammography is presented. In conclusion, with a smarter patch extraction, the CBIR approach might reach precision rates that are helpful for the physicians. This, however, needs more comprehensive evaluation on clinical data.
-
Clinical cytometry and progress in HLA antibody detection.
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.
-
Using Lanthanide Nanoparticles as Isotopic Tags for Biomarker Detection by Mass Cytometry
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
-
The impact of digital imaging in the field of cytopathology.
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.
-
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
-
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
-
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.
-
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.
-
Lebaron, P; Servais, P; Agogué, H; Courties, C; Joux, F
2001-04-01
The nucleic acid contents of individual bacterial cells as determined with three different nucleic acid-specific fluorescent dyes (SYBR I, SYBR II, and SYTO 13) and flow cytometry were compared for different seawater samples. Similar fluorescence patterns were observed, and bacteria with high apparent nucleic acid contents (HNA) could be discriminated from bacteria with low nucleic acid contents (LNA). The best discrimination between HNA and LNA cells was found when cells were stained with SYBR II. Bacteria in different water samples collected from seven freshwater, brackish water, and seawater ecosystems were prelabeled with tritiated leucine and then stained with SYBR II. After labeling and staining, HNA, LNA, and total cells were sorted by flow cytometry, and the specific activity of each cellular category was determined from leucine incorporation rates. The HNA cells were responsible for most of the total bacterial production, and the specific activities of cells in the HNA population varied between samples by a factor of seven. We suggest that nucleic acid content alone can be a better indicator of the fraction of growing cells than total counts and that this approach should be combined with other fluorescent physiological probes to improve detection of the most active cells in aquatic systems.
-
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...
-
Charoenkwan, Phasit; Hwang, Eric; Cutler, Robert W; Lee, Hua-Chin; Ko, Li-Wei; Huang, Hui-Ling; Ho, Shinn-Ying
2013-01-01
High-content screening (HCS) has become a powerful tool for drug discovery. However, the discovery of drugs targeting neurons is still hampered by the inability to accurately identify and quantify the phenotypic changes of multiple neurons in a single image (named multi-neuron image) of a high-content screen. Therefore, it is desirable to develop an automated image analysis method for analyzing multi-neuron images. We propose an automated analysis method with novel descriptors of neuromorphology features for analyzing HCS-based multi-neuron images, called HCS-neurons. To observe multiple phenotypic changes of neurons, we propose two kinds of descriptors which are neuron feature descriptor (NFD) of 13 neuromorphology features, e.g., neurite length, and generic feature descriptors (GFDs), e.g., Haralick texture. HCS-neurons can 1) automatically extract all quantitative phenotype features in both NFD and GFDs, 2) identify statistically significant phenotypic changes upon drug treatments using ANOVA and regression analysis, and 3) generate an accurate classifier to group neurons treated by different drug concentrations using support vector machine and an intelligent feature selection method. To evaluate HCS-neurons, we treated P19 neurons with nocodazole (a microtubule depolymerizing drug which has been shown to impair neurite development) at six concentrations ranging from 0 to 1000 ng/mL. The experimental results show that all the 13 features of NFD have statistically significant difference with respect to changes in various levels of nocodazole drug concentrations (NDC) and the phenotypic changes of neurites were consistent to the known effect of nocodazole in promoting neurite retraction. Three identified features, total neurite length, average neurite length, and average neurite area were able to achieve an independent test accuracy of 90.28% for the six-dosage classification problem. This NFD module and neuron image datasets are provided as a freely downloadable
-
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.
-
A CLIPS expert system for clinical flow cytometry data analysis
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.
-
Hatta, Tomoko; Fujinaga, Yasunari; Kadoya, Masumi; Ueda, Hitoshi; Murayama, Hiroaki; Kurozumi, Masahiro; Ueda, Kazuhiko; Komatsu, Michiharu; Nagaya, Tadanobu; Joshita, Satoru; Kodama, Ryo; Tanaka, Eiji; Uehara, Tsuyoshi; Sano, Kenji; Tanaka, Naoki
2010-12-01
To assess the degree of hepatic fat content, simple and noninvasive methods with high objectivity and reproducibility are required. Magnetic resonance imaging (MRI) is one such candidate, although its accuracy remains unclear. We aimed to validate an MRI method for quantifying hepatic fat content by calibrating MRI reading with a phantom and comparing MRI measurements in human subjects with estimates of liver fat content in liver biopsy specimens. The MRI method was performed by a combination of MRI calibration using a phantom and double-echo chemical shift gradient-echo sequence (double-echo fast low-angle shot sequence) that has been widely used on a 1.5-T scanner. Liver fat content in patients with nonalcoholic fatty liver disease (NAFLD, n = 26) was derived from a calibration curve generated by scanning the phantom. Liver fat was also estimated by optical image analysis. The correlation between the MRI measurements and liver histology findings was examined prospectively. Magnetic resonance imaging measurements showed a strong correlation with liver fat content estimated from the results of light microscopic examination (correlation coefficient 0.91, P hepatic steatosis. Moreover, the severity of lobular inflammation or fibrosis did not influence the MRI measurements. This MRI method is simple and noninvasive, has excellent ability to quantify hepatic fat content even in NAFLD patients with mild steatosis or advanced fibrosis, and can be performed easily without special devices.
-
International Nuclear Information System (INIS)
Flaberg, Emilie; Sabelström, Per; Strandh, Christer; Szekely, Laszlo
2008-01-01
Confocal laser scanning microscopy has revolutionized cell biology. However, the technique has major limitations in speed and sensitivity due to the fact that a single laser beam scans the sample, allowing only a few microseconds signal collection for each pixel. This limitation has been overcome by the introduction of parallel beam illumination techniques in combination with cold CCD camera based image capture. Using the combination of microlens enhanced Nipkow spinning disc confocal illumination together with fully automated image capture and large scale in silico image processing we have developed a system allowing the acquisition, presentation and analysis of maximum resolution confocal panorama images of several Gigapixel size. We call the method Extended Field Laser Confocal Microscopy (EFLCM). We show using the EFLCM technique that it is possible to create a continuous confocal multi-colour mosaic from thousands of individually captured images. EFLCM can digitize and analyze histological slides, sections of entire rodent organ and full size embryos. It can also record hundreds of thousands cultured cells at multiple wavelength in single event or time-lapse fashion on fixed slides, in live cell imaging chambers or microtiter plates. The observer independent image capture of EFLCM allows quantitative measurements of fluorescence intensities and morphological parameters on a large number of cells. EFLCM therefore bridges the gap between the mainly illustrative fluorescence microscopy and purely quantitative flow cytometry. EFLCM can also be used as high content analysis (HCA) instrument for automated screening processes
-
Analysis of nuclear localization of interleukin-1 family cytokines by flow cytometry.
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.
-
Webb, Jennifer B; Vinoski, Erin R; Bonar, Adrienne S; Davies, Alexandria E; Etzel, Lena
2017-09-01
In step with the proliferation of Thinspiration and Fitspiration content disseminated in popular web-based media, the fat acceptance movement has garnered heightened visibility within mainstream culture via the burgeoning Fatosphere weblog community. The present study extended previous Fatosphere research by comparing the shared and distinct strategies used to represent and motivate a fat-accepting lifestyle among 400 images sourced from Fatspiration- and Health at Every Size ® -themed hashtags on Instagram. Images were systematically analyzed for the socio-demographic and body size attributes of the individuals portrayed alongside content reflecting dimensions of general fat acceptance, physical appearance pride, physical activity and health, fat shaming, and eating and weight loss-related themes. #fatspiration/#fatspo-tagged images more frequently promoted fat acceptance through fashion and beauty-related activism; #healthateverysize/#haes posts more often featured physically-active portrayals, holistic well-being, and weight stigma. Findings provide insight into the common and unique motivational factors and contradictory messages encountered in these fat-accepting social media communities. Copyright © 2017 Elsevier Ltd. All rights reserved.
-
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.
-
Reticulocyte analysis using flow cytometry.
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.
-
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.
-
ESIM: Edge Similarity for Screen Content Image Quality Assessment.
Ni, Zhangkai; Ma, Lin; Zeng, Huanqiang; Chen, Jing; Cai, Canhui; Ma, Kai-Kuang
2017-10-01
In this paper, an accurate full-reference image quality assessment (IQA) model developed for assessing screen content images (SCIs), called the edge similarity (ESIM), is proposed. It is inspired by the fact that the human visual system (HVS) is highly sensitive to edges that are often encountered in SCIs; therefore, essential edge features are extracted and exploited for conducting IQA for the SCIs. The key novelty of the proposed ESIM lies in the extraction and use of three salient edge features-i.e., edge contrast, edge width, and edge direction. The first two attributes are simultaneously generated from the input SCI based on a parametric edge model, while the last one is derived directly from the input SCI. The extraction of these three features will be performed for the reference SCI and the distorted SCI, individually. The degree of similarity measured for each above-mentioned edge attribute is then computed independently, followed by combining them together using our proposed edge-width pooling strategy to generate the final ESIM score. To conduct the performance evaluation of our proposed ESIM model, a new and the largest SCI database (denoted as SCID) is established in our work and made to the public for download. Our database contains 1800 distorted SCIs that are generated from 40 reference SCIs. For each SCI, nine distortion types are investigated, and five degradation levels are produced for each distortion type. Extensive simulation results have clearly shown that the proposed ESIM model is more consistent with the perception of the HVS on the evaluation of distorted SCIs than the multiple state-of-the-art IQA methods.
-
Bleaching response of Symbiodinium (zooxanthellae): determination by flow cytometry.
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.
-
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.
-
Kawasaki, M; Sasaki, K; Satoh, T; Kurose, A; Kamada, T; Furuya, T; Murakami, T; Todoroki, T
1997-01-01
We have demonstrated a method for the in situ determination of the cell cycle phases of TIG-7 fibroblasts using a laser scanning cytometer (LSC) which has not only a function equivalent to flow cytometry (FCM) but also has a capability unique in itself. LSC allows a more detailed analysis of the cell cycle in cells stained with propidium iodide (PI) than FCM. With LSC it is possible to discriminate between mitotic cells and G2 cells, between post-mitotic cells and G1 cells, and between quiescent cells and cycling cells in a PI fluorescence peak (chromatin condensation) vs. fluorescence value (DNA content) cytogram for cells stained with PI. These were amply confirmed by experiments using colcemid and adriamycin. We were able to identify at least six cell subpopulations for PI stained cells using LSC; namely G1, S, G2, M, postmitotic and quiescent cell populations. LSC analysis facilitates the monitoring of effects of drugs on the cell cycle.
-
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.
-
Directory of Open Access Journals (Sweden)
Yansheng Li
2016-08-01
Full Text Available With the urgent demand for automatic management of large numbers of high-resolution remote sensing images, content-based high-resolution remote sensing image retrieval (CB-HRRS-IR has attracted much research interest. Accordingly, this paper proposes a novel high-resolution remote sensing image retrieval approach via multiple feature representation and collaborative affinity metric fusion (IRMFRCAMF. In IRMFRCAMF, we design four unsupervised convolutional neural networks with different layers to generate four types of unsupervised features from the fine level to the coarse level. In addition to these four types of unsupervised features, we also implement four traditional feature descriptors, including local binary pattern (LBP, gray level co-occurrence (GLCM, maximal response 8 (MR8, and scale-invariant feature transform (SIFT. In order to fully incorporate the complementary information among multiple features of one image and the mutual information across auxiliary images in the image dataset, this paper advocates collaborative affinity metric fusion to measure the similarity between images. The performance evaluation of high-resolution remote sensing image retrieval is implemented on two public datasets, the UC Merced (UCM dataset and the Wuhan University (WH dataset. Large numbers of experiments show that our proposed IRMFRCAMF can significantly outperform the state-of-the-art approaches.
-
Directory of Open Access Journals (Sweden)
Liu Shanmei
2016-01-01
Full Text Available Non-destructive determination of TVB-N content in beef using hyperspectral imaging (HSI technique was evaluated. In order to create a robust model to predict the TVB-N content in beef, partition of sample set, spectral pretreatment, and the optimum wavelength selection were discussed. After the beef sample set was parted by concentration gradient (CG algortithm, and the spectra of beef samples were preprocessed by standard normalized variate (SNV combined with auto scale(AS, the partial least square regression (PLSR model was established using the full spectral range, which had the best prediction abilities with Rcv2 of 0.9124, Rp2 of 0.8816, RMSECV of 1.5889, and RMSEP of 1.7719, respectively. After the optimum wavelengths which is closely related to the TVB-N content of beef samples was obtained using the competitive adaptive re-weighted (CARS algorithm, a new PLSR model was established using the optimum wavelengths, which had outstanding prediction abilities with Rcv2 of 0.9235, Rp2 of 0.9241, RMSECV of 1.4881, and RMSEP of 1.4882, respectively.The study showed that HSI is a powerful technique to predict the TVB-N content in beef by a nondestructive way.
-
Enhancing Image Retrieval System Using Content Based Search ...
African Journals Online (AJOL)
The output shows more efficiency in retrieval because instead of performing the search on the entire image database, the image category option directs the retrieval engine to the specified category. Also, there is provision to update or modify the different image categories in the image database as need arise. Keywords: ...
-
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...
-
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.
-
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.
-
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.
-
Bradford, Jolene A; Clarke, Scott T
2011-01-01
Adult human mesenchymal stem cells (hMSC) are rare fibroblast-like cells capable of differentiation into a variety of cell tissues which include bone, cartilage, muscle, ligament, tendon, and adipose. Normal adult bone marrow and adipose tissue are the most common sources of these cells. The International Society for Cellular Therapy (ISCT) has proposed a set of standards to define hMSC for laboratory investigations and preclinical studies: adherence to plastic in standard culture conditions; in vitro differentiation into osteoblasts, adipocytes, and chondroblasts; and specific surface antigen expression. Direct measurement of proliferation combined with simultaneous detection of the ISCT-consensus immunophenotypic profile provides data that is used to determine the differentiation status and health of the cells. Flow cytometry provides a powerful technology that is routinely used to simultaneously and rapidly measure multiple parameters in a single sample. This chapter describes a flow cytometric panel for the simultaneous detection of immunophenotypic profile, proliferative capacity, and DNA content measurement in hMSC. Because a relatively small number of cells are needed with this approach, measurements can be made with minimal impact on expansion potential. The ability to assess antigen expression and proliferative status enables the investigator to make informed decisions on expansion and harvesting.
-
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...
-
Brosnahan, Michael L.; Farzan, Shahla; Keafer, Bruce A.; Sosik, Heidi M.; Olson, Robert J.; Anderson, Donald M.
2014-05-01
Measurements of the DNA content of different protist populations can shed light on a variety of processes, including cell division, sex, prey ingestion, and parasite invasion. Here, we modified an Imaging FlowCytobot (IFCB), a custom-built flow cytometer that records images of microplankton, to measure the DNA content of large dinoflagellates and other high-DNA content species. The IFCB was also configured to measure fluorescence from Cy3-labeled rRNA probes, aiding the identification of Alexandrium fundyense (syn. A. tamarense Group I), a photosynthetic dinoflagellate that causes paralytic shellfish poisoning (PSP). The modified IFCB was used to analyze samples from the development, peak and termination phases of an inshore A. fundyense bloom (Salt Pond, Eastham, MA, USA), and from a rare A. fundyense ‘red tide’ that occurred in the western Gulf of Maine, offshore of Portsmouth, NH (USA). Diploid or G2 phase (‘2C’) A. fundyense cells were frequently enriched at the near-surface, suggesting an important role for aggregation at the air-sea interface during sexual events. Also, our analysis showed that large proportions of A. fundyense cells in both the Salt Pond and red tide blooms were planozygotes during bloom decline, highlighting the importance of sexual fusion to bloom termination. At Salt Pond, bloom decline also coincided with a dramatic rise in infections by the parasite genus Amoebophrya. The samples that were most heavily infected contained many large cells with higher DNA-associated fluorescence than 2C vegetative cells, but these cells' nuclei were also frequently consumed by Amoebophrya trophonts. Neither large cell size nor increased DNA-associated fluorescence could be replicated by infecting an A. fundyense culture of vegetative cells. Therefore, we attribute these characteristics of the large Salt Pond cells to planozygote maturation rather than Amoebophrya infection, though an interaction between infection and planozygote maturation may
-
Preparation and flow cytometry of uniform silica-fluorescent dye microspheres.
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.
-
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.
-
Directory of Open Access Journals (Sweden)
Lassi Rieppo
Full Text Available Fourier Transform Infrared (FT-IR spectroscopic imaging has been earlier applied for the spatial estimation of the collagen and the proteoglycan (PG contents of articular cartilage (AC. However, earlier studies have been limited to the use of univariate analysis techniques. Current analysis methods lack the needed specificity for collagen and PGs. The aim of the present study was to evaluate the suitability of partial least squares regression (PLSR and principal component regression (PCR methods for the analysis of the PG content of AC. Multivariate regression models were compared with earlier used univariate methods and tested with a sample material consisting of healthy and enzymatically degraded steer AC. Chondroitinase ABC enzyme was used to increase the variation in PG content levels as compared to intact AC. Digital densitometric measurements of Safranin O-stained sections provided the reference for PG content. The results showed that multivariate regression models predict PG content of AC significantly better than earlier used absorbance spectrum (i.e. the area of carbohydrate region with or without amide I normalization or second derivative spectrum univariate parameters. Increased molecular specificity favours the use of multivariate regression models, but they require more knowledge of chemometric analysis and extended laboratory resources for gathering reference data for establishing the models. When true molecular specificity is required, the multivariate models should be used.
-
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.
-
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. ...
-
Facilitating personal content management in smart phones
Aaltonen, Antti
2007-01-01
Smart phones, which combine, e.g., communication and mobile multimedia features, store increasing amount of media content and so they face content management challenges similar to what desktop computers are experiencing. Content management refers to actions performed on content (e.g., capture image,
-
Directory of Open Access Journals (Sweden)
Fernández Prieto, José Antonio
2011-12-01
Full Text Available Flow cytometry analysis has been widely applied in the determination of nuclear DNA content and ploidy level in many organisms. Despite being the most appropriate method for DNA content measurement, flow cytometry also presents some limitations. A fairly common, but little-studied problem is the effect on measurements of the presence of secondary metabolites. A good example is the genus Viola, which is composed of 525-600 species distributed worldwide. These species have proved to be problematic for flow cytometric analyses due to the release of extremely mucilaginous compounds into the nuclear suspension. In this work, the genome size of 13 species of Viola using flow cytometry are presented for the first time. Despite obtaining histograms with high coefficients of variation, we here present an optimized protocol to remove cytoplasmic compounds, particularly mucilaginous ones, from plant nuclei that pave the way for its application to estimate the genome size of other species exhibiting similar problems. Statistical analyses revealed significant differences between sections Viola and Melanium, and within each section (P El análisis mediante citometría de flujo ha sido aplicado de modo general para determinar el contenido de ADN nuclear y el nivel de ploidía en muchos organismos. A pesar de ser el método más adecuado para medir la cantidad de ADN, esta técnica también presenta algunas limitaciones. Un problema bastante común, aunque poco estudiado, es el efecto de los metabolitos secundarios en los resultados obtenidos. Un ejemplo respecto a la presencia de estos compuestos se encuentra en el género Viola, compuesto por 525-600 especies distribuidas por todo el mundo. Las especies de este género ya han sido previamente descritas como problemáticas en los análisis de citometría de flujo debido a la presencia de compuestos extremadamente mucilaginosos en las suspensiones de núcleos. En el presente trabajo se analiza por primera vez
-
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.
-
De Silva, T.; Uneri, A.; Ketcha, M. D.; Reaungamornrat, S.; Kleinszig, G.; Vogt, S.; Aygun, N.; Lo, S.-F.; Wolinsky, J.-P.; Siewerdsen, J. H.
2016-04-01
In image-guided spine surgery, robust three-dimensional to two-dimensional (3D-2D) registration of preoperative computed tomography (CT) and intraoperative radiographs can be challenged by the image content mismatch associated with the presence of surgical instrumentation and implants as well as soft-tissue resection or deformation. This work investigates image similarity metrics in 3D-2D registration offering improved robustness against mismatch, thereby improving performance and reducing or eliminating the need for manual masking. The performance of four gradient-based image similarity metrics (gradient information (GI), gradient correlation (GC), gradient information with linear scaling (GS), and gradient orientation (GO)) with a multi-start optimization strategy was evaluated in an institutional review board-approved retrospective clinical study using 51 preoperative CT images and 115 intraoperative mobile radiographs. Registrations were tested with and without polygonal masks as a function of the number of multistarts employed during optimization. Registration accuracy was evaluated in terms of the projection distance error (PDE) and assessment of failure modes (PDE > 30 mm) that could impede reliable vertebral level localization. With manual polygonal masking and 200 multistarts, the GC and GO metrics exhibited robust performance with 0% gross failures and median PDE interquartile range (IQR)) and a median runtime of 84 s (plus upwards of 1-2 min for manual masking). Excluding manual polygonal masks and decreasing the number of multistarts to 50 caused the GC-based registration to fail at a rate of >14% however, GO maintained robustness with a 0% gross failure rate. Overall, the GI, GC, and GS metrics were susceptible to registration errors associated with content mismatch, but GO provided robust registration (median PDE = 5.5 mm, 2.6 mm IQR) without manual masking and with an improved runtime (29.3 s). The GO metric
-
Siadat, Mohammad-Reza; Soltanian-Zadeh, Hamid; Fotouhi, Farshad A.; Elisevich, Kost
2003-01-01
This paper presents the development of a human brain multimedia database for surgical candidacy determination in temporal lobe epilepsy. The focus of the paper is on content-based image management, navigation and retrieval. Several medical image-processing methods including our newly developed segmentation method are utilized for information extraction/correlation and indexing. The input data includes T1-, T2-Weighted MRI and FLAIR MRI and ictal and interictal SPECT modalities with associated clinical data and EEG data analysis. The database can answer queries regarding issues such as the correlation between the attribute X of the entity Y and the outcome of a temporal lobe epilepsy surgery. The entity Y can be a brain anatomical structure such as the hippocampus. The attribute X can be either a functionality feature of the anatomical structure Y, calculated with SPECT modalities, such as signal average, or a volumetric/morphological feature of the entity Y such as volume or average curvature. The outcome of the surgery can be any surgery assessment such as memory quotient. A determination is made regarding surgical candidacy by analysis of both textual and image data. The current database system suggests a surgical determination for the cases with relatively small hippocampus and high signal intensity average on FLAIR images within the hippocampus. This indication pretty much fits with the surgeons" expectations/observations. Moreover, as the database gets more populated with patient profiles and individual surgical outcomes, using data mining methods one may discover partially invisible correlations between the contents of different modalities of data and the outcome of the surgery.
-
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.
-
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.
-
Menon, K Venugopal; Kumar, Dinesh; Thomas, Tessamma
2014-02-01
Study Design Preliminary evaluation of new tool. Objective To ascertain whether the newly developed content-based image retrieval (CBIR) software can be used successfully to retrieve images of similar cases of adolescent idiopathic scoliosis (AIS) from a database to help plan treatment without adhering to a classification scheme. Methods Sixty-two operated cases of AIS were entered into the newly developed CBIR database. Five new cases of different curve patterns were used as query images. The images were fed into the CBIR database that retrieved similar images from the existing cases. These were analyzed by a senior surgeon for conformity to the query image. Results Within the limits of variability set for the query system, all the resultant images conformed to the query image. One case had no similar match in the series. The other four retrieved several images that were matching with the query. No matching case was left out in the series. The postoperative images were then analyzed to check for surgical strategies. Broad guidelines for treatment could be derived from the results. More precise query settings, inclusion of bending films, and a larger database will enhance accurate retrieval and better decision making. Conclusion The CBIR system is an effective tool for accurate documentation and retrieval of scoliosis images. Broad guidelines for surgical strategies can be made from the postoperative images of the existing cases without adhering to any classification scheme.
-
Pila, Eva; Mond, Jonathan M; Griffiths, Scott; Mitchison, Deborah; Murray, Stuart B
2017-06-01
Despite the pervasive social endorsement of "cheat meals" within pro-muscularity online communities, there is an absence of empirical work examining this dietary phenomenon. The present study aimed to characterize cheat meals, and explore the meaning ascribed to engagement in this practice. Thematic content analysis was employed to code the photographic and textual elements of a sample (n = 600) that was extracted from over 1.6 million images marked with the #cheatmeal tag on the social networking site, Instagram. Analysis of the volume and type of food revealed the presence of very large quantities (54.5%) of calorie-dense foods (71.3%) that was rated to qualify as an objective binge episode. Photographic content of people commonly portrayed highly-muscular bodies (60.7%) in the act of intentional body exposure (40.0%). Meanwhile, textual content exemplified the idealization of overconsumption, a strict commitment to fitness, and a reward-based framework around diet and fitness. Collectively, these findings position cheat meals as goal-oriented dietary practices in the pursuit of physique-ideals, thus underscoring the potential clinical repercussions of this socially-endorsed dietary phenomenon. © 2017 Wiley Periodicals, Inc.
-
Peláez, Jesús; Long, Julie A
2007-01-01
The aim of the present work was to use a battery of lectins to 1) delineate the carbohydrate content of sperm glycocalyx in the turkey and chicken using flow cytometry analysis, and 2) evaluate the distribution of existing sugars over the sperm plasma membrane surface with epifluorescent microscopy. Carbohydrate groups (corresponding lectins) that were investigated included galactose (GS-I, Jacalin, RCA-I, PNA), glucose and/or mannose (Con A, PSA, GNA), N-acetyl-glucosamine (GS-II, s-WGA, STA), N-acetyl-galactosamine (SBA, WFA), fucose (Lotus, UEA-I), sialic acid (LFA, LPA), and N-acetyl-lactosamine (ECA). Spermatozoa were assessed before and after treatment with neuraminidase to remove sialic acid. Mean fluorescence intensity (MnFI) was used as indicator of lectin binding for flow cytometry analysis. Nontreated spermatozoa from both species showed high MnFI when incubated with RCA-I, Con A, LFA, and LPA, as did chicken spermatozoa incubated with s-WGA. Neuraminidase treatment increased the MnFI for most lectins except LFA and LPA, as expected. Differences in MnFI between species included higher values for s-WGA and ECA in chicken spermatozoa and for WFA in turkey spermatozoa. Microscopy revealed segregation of some sugar residues into membrane-specific domains; however, the 2 staining techniques (cell suspension vs fixed preparation) differed in identifying lectin binding patterns, with fixed preparations yielding a high degree of nonspecific binding. We conclude that 1) the glycocalyx of turkey and chicken spermatozoa contains a diversity of carbohydrate groups, 2) these residues are extensively masked by sialic acid, 3) the glycocalyx composition is species-specific, and 4) some glycoconjugates appear to be segregated into membrane-specific domains. Characterization of the poultry sperm glycocalyx is the first step in identifying the physiological impact of semen storage on sperm function.
-
Leskovjan, Andreana C; Kretlow, Ariane; Miller, Lisa M
2010-04-01
Polyunsaturated fatty acids are essential to brain functions such as membrane fluidity, signal transduction, and cell survival. It is also thought that low levels of unsaturated lipid in the brain may contribute to Alzheimer's disease (AD) risk or severity. However, it is not known how accumulation of unsaturated lipids is affected in different regions of the hippocampus, which is a central target of AD plaque pathology, during aging. In this study, we used Fourier transform infrared imaging (FTIRI) to visualize the unsaturated lipid content in specific regions of the hippocampus in the PSAPP mouse model of AD as a function of plaque formation. Specifically, the unsaturated lipid content was imaged using the olefinic =CH stretching mode at 3012 cm(-1). The axonal, dendritic, and somatic layers of the hippocampus were examined in the mice at 13, 24, 40, and 56 weeks old. Results showed that lipid unsaturation in the axonal layer was significantly increased with normal aging in control (CNT) mice (p avoiding progression of the disease.
-
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....
-
Lockwood, S F; Bickham, J W
1991-01-01
Intraspecific variation in cellular DNA content was measured in five Coregonus autumnalis spawning populations from the Mackenzie River drainage, Canada, using flow cytometry. The rivers assayed were the Peel, Arctic Red, Mountain, Carcajou, and Liard rivers. DNA content was determined from whole blood preparations of fish from all rivers except the Carcajou, for which kidney tissue was used. DNA content measurements of kidney and blood preparations of the same fish from the Mountain River revealed statistically indistinguishable results. Mosaicism was found in blood preparations from the Peel, Arctic Red, Mountain, and Liard rivers, but was not observed in kidney tissue preparations from the Mountain or Carcajou rivers. The Liard River sample had significantly elevated mean DNA content relative to the other four samples; all other samples were statistically indistinguishable. Significant differences in mean DNA content among spawning stocks of a single species reinforces the need for adequate sample sizes of both individuals and populations when reporting "C" values for a particular species.
-
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
-
Yang, Wei; Feng, Qianjin; Yu, Mei; Lu, Zhentai; Gao, Yang; Xu, Yikai; Chen, Wufan
2012-11-01
A content-based image retrieval (CBIR) method for T1-weighted contrast-enhanced MRI (CE-MRI) images of brain tumors is presented for diagnosis aid. The method is thoroughly evaluated on a large image dataset. Using the tumor region as a query, the authors' CBIR system attempts to retrieve tumors of the same pathological category. Aside from commonly used features such as intensity, texture, and shape features, the authors use a margin information descriptor (MID), which is capable of describing the characteristics of tissue surrounding a tumor, for representing image contents. In addition, the authors designed a distance metric learning algorithm called Maximum mean average Precision Projection (MPP) to maximize the smooth approximated mean average precision (mAP) to optimize retrieval performance. The effectiveness of MID and MPP algorithms was evaluated using a brain CE-MRI dataset consisting of 3108 2D scans acquired from 235 patients with three categories of brain tumors (meningioma, glioma, and pituitary tumor). By combining MID and other features, the mAP of retrieval increased by more than 6% with the learned distance metrics. The distance metric learned by MPP significantly outperformed the other two existing distance metric learning methods in terms of mAP. The CBIR system using the proposed strategies achieved a mAP of 87.3% and a precision of 89.3% when top 10 images were returned by the system. Compared with scale-invariant feature transform, the MID, which uses the intensity profile as descriptor, achieves better retrieval performance. Incorporating tumor margin information represented by MID with the distance metric learned by the MPP algorithm can substantially improve the retrieval performance for brain tumors in CE-MRI.
-
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
-
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.
-
[Quantitative image of bone mineral content--dual energy subtraction in a single exposure].
Katoh, T
1990-09-25
A dual energy subtraction system was constructed on an experimental basis for the quantitative image of bone mineral content. The system consists of a radiography system and an image processor. Two radiograms were taken with dual x-ray energy in a single exposure using an x-ray beam dichromized by a tin filter. In this system, a film cassette was used where a low speed film-screen system, a copper filter and a high speed film-screen system were layered on top of each other. The images were read by a microdensitometer and processed by a personal computer. The image processing included the corrections of the film characteristics and heterogeneity in the x-ray field, and the dual energy subtraction in which the effect of the high energy component of the dichromized beam on the tube side image was corrected. In order to determine the accuracy of the system, experiments using wedge phantoms made of mixtures of epoxy resin and bone mineral-equivalent materials in various fractions were performed for various tube potentials and film processing conditions. The results indicated that the relative precision of the system was within +/- 4% and that the propagation of the film noise was within +/- 11 mg/cm2 for the 0.2 mm pixels. The results also indicated that the system response was independent of the tube potential and the film processing condition. The bone mineral weight in each phalanx of the freshly dissected hand of a rhesus monkey was measured by this system and compared with the ash weight. The results showed an error of +/- 10%, slightly larger than that of phantom experiments, which is probably due to the effect of fat and the variation of focus-object distance. The air kerma in free air at the object was approximately 0.5 mGy for one exposure. The results indicate that this system is applicable to clinical use and provides useful information for evaluating a time-course of localized bone disease.
-
International Nuclear Information System (INIS)
Kurokawa, Hiroaki; Nakano, Yoshihisa; Ikeda, Koshi; Tanaka, Yoshimasa; Fujisawa, Ichiro
1998-01-01
We investigated the correlation between the signal intensity on T 1 -weighted MR images and vasopressin (VP) content in the posterior pituitary lobe. Fourteen rabbits were studied. There were 12 water-deprived rabbits (48, 72, 96, 120, 144 and 168 hours: 2 each) and 2 controls. Sagittal T 1 -weighted SE (spin-echo) MR images were obtained before and after dehydration. The signal intensity ratio of the posterior pituitary lobe to the pons was correlated with the VP content in the posterior lobe as measured by radioimmunoassay. Before water deprivation, high signal intensity in the posterior lobe was demonstrated clearly in all 14 rabbits. After water deprivation, the hyperintense signal gradually decreased and became indistinguishable from anterior lobe in four animals. The mean signal intensity ratio before water deprivation was 1.55±0.12 (mean±SD) and after water deprivation, gradually decreased over time and reached to 1.19 after 168 hours of water deprivation. Pituitary VP content and concentration decreased in parallel with the signal intensity ratio of the posterior pituitary. Significantly correlation was observed between the signal intensity ratio and VP concentration of posterior pituitary (r=0.809, p 1 -weighted image may reflect a indicator of pituitary VP content and thus may enable evaluation of disorders of water metabolism. (author)
-
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.
-
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)
-
Mustapha, Aouache; Hussain, Aini; Samad, Salina Abdul; Zulkifley, Mohd Asyraf; Diyana Wan Zaki, Wan Mimi; Hamid, Hamzaini Abdul
2015-01-16
Content-based medical image retrieval (CBMIR) system enables medical practitioners to perform fast diagnosis through quantitative assessment of the visual information of various modalities. In this paper, a more robust CBMIR system that deals with both cervical and lumbar vertebrae irregularity is afforded. It comprises three main phases, namely modelling, indexing and retrieval of the vertebrae image. The main tasks in the modelling phase are to improve and enhance the visibility of the x-ray image for better segmentation results using active shape model (ASM). The segmented vertebral fractures are then characterized in the indexing phase using region-based fracture characterization (RB-FC) and contour-based fracture characterization (CB-FC). Upon a query, the characterized features are compared to the query image. Effectiveness of the retrieval phase is determined by its retrieval, thus, we propose an integration of the predictor model based cross validation neural network (PMCVNN) and similarity matching (SM) in this stage. The PMCVNN task is to identify the correct vertebral irregularity class through classification allowing the SM process to be more efficient. Retrieval performance between the proposed and the standard retrieval architectures are then compared using retrieval precision (Pr@M) and average group score (AGS) measures. Experimental results show that the new integrated retrieval architecture performs better than those of the standard CBMIR architecture with retrieval results of cervical (AGS > 87%) and lumbar (AGS > 82%) datasets. The proposed CBMIR architecture shows encouraging results with high Pr@M accuracy. As a result, images from the same visualization class are returned for further used by the medical personnel.
-
Directory of Open Access Journals (Sweden)
Allison DaSilva
2014-10-01
Full Text Available A comparative analysis of content enrichment services, Syndetic Solutions and Content Café 2, was undertaken to explore which service would provide public library users with a superior online search and discovery experience through the enriched data elements offered, specifically looking at the cover image data element and exploring what factors impact the display of said element. A data-set of 250 items in five different formats, including books, CDs, DVDs, e-books, and video games, was searched in four North American public libraries’ discovery layers to compare the integration, extent, and quality of the cover image data element supplied by Syndetic Solutions and Content Café 2. Based on an analysis of the URLs, ISBNs, and UPCs for each of the 250 items, it was determined that the integration, and therefore the display, of the cover image data element was impacted by: (1 whether or not an ISBN or UPC was listed in the MARC bibliographic record; (2 which ISBN or UPC was listed, as items could potentially have more than one; (3 the inclusion of both ISBNs and UPCs in the record and the settings of the discovery tool; (4 the order in which the ISBNs or UPCs were listed in the record; and (5 whether or not Syndetic Solutions or Content Café 2 had the image data in its database at the time of the search. The quality of the cover image displayed was found to be impacted by the size requested by the library and the size of the image provided by the publisher. These findings may also have implications for the integration of other enriched data elements.
-
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.
-
Deal, Samantha; Wambaugh, John; Judson, Richard; Mosher, Shad; Radio, Nick; Houck, Keith; Padilla, Stephanie
2016-09-01
One of the rate-limiting procedures in a developmental zebrafish screen is the morphological assessment of each larva. Most researchers opt for a time-consuming, structured visual assessment by trained human observer(s). The present studies were designed to develop a more objective, accurate and rapid method for screening zebrafish for dysmorphology. Instead of the very detailed human assessment, we have developed the computational malformation index, which combines the use of high-content imaging with a very brief human visual assessment. Each larva was quickly assessed by a human observer (basic visual assessment), killed, fixed and assessed for dysmorphology with the Zebratox V4 BioApplication using the Cellomics® ArrayScan® V(TI) high-content image analysis platform. The basic visual assessment adds in-life parameters, and the high-content analysis assesses each individual larva for various features (total area, width, spine length, head-tail length, length-width ratio, perimeter-area ratio). In developing the computational malformation index, a training set of hundreds of embryos treated with hundreds of chemicals were visually assessed using the basic or detailed method. In the second phase, we assessed both the stability of these high-content measurements and its performance using a test set of zebrafish treated with a dose range of two reference chemicals (trans-retinoic acid or cadmium). We found the measures were stable for at least 1 week and comparison of these automated measures to detailed visual inspection of the larvae showed excellent congruence. Our computational malformation index provides an objective manner for rapid phenotypic brightfield assessment of individual larva in a developmental zebrafish assay. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.
-
International Nuclear Information System (INIS)
De Silva, T; Ketcha, M D; Siewerdsen, J H; Uneri, A; Reaungamornrat, S; Kleinszig, G; Vogt, S; Aygun, N; Lo, S-F; Wolinsky, J-P
2016-01-01
In image-guided spine surgery, robust three-dimensional to two-dimensional (3D–2D) registration of preoperative computed tomography (CT) and intraoperative radiographs can be challenged by the image content mismatch associated with the presence of surgical instrumentation and implants as well as soft-tissue resection or deformation. This work investigates image similarity metrics in 3D–2D registration offering improved robustness against mismatch, thereby improving performance and reducing or eliminating the need for manual masking. The performance of four gradient-based image similarity metrics (gradient information (GI), gradient correlation (GC), gradient information with linear scaling (GS), and gradient orientation (GO)) with a multi-start optimization strategy was evaluated in an institutional review board-approved retrospective clinical study using 51 preoperative CT images and 115 intraoperative mobile radiographs. Registrations were tested with and without polygonal masks as a function of the number of multistarts employed during optimization. Registration accuracy was evaluated in terms of the projection distance error (PDE) and assessment of failure modes (PDE > 30 mm) that could impede reliable vertebral level localization. With manual polygonal masking and 200 multistarts, the GC and GO metrics exhibited robust performance with 0% gross failures and median PDE < 6.4 mm (±4.4 mm interquartile range (IQR)) and a median runtime of 84 s (plus upwards of 1–2 min for manual masking). Excluding manual polygonal masks and decreasing the number of multistarts to 50 caused the GC-based registration to fail at a rate of >14%; however, GO maintained robustness with a 0% gross failure rate. Overall, the GI, GC, and GS metrics were susceptible to registration errors associated with content mismatch, but GO provided robust registration (median PDE = 5.5 mm, 2.6 mm IQR) without manual masking and with an improved
-
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...
-
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...
-
Smarda, P; Stancík, D
2006-01-01
Ploidy levels and chromosome numbers for 24 species of Festuca L. from 29 sites in Bolivia, Colombia, Ecuador and Venezuela are given. The ploidy level of 22 species is reported for the first time. A higher proportion of tetraploids in northern South America and the high frequency of polyploids in the whole continent are documented. In combination with chromosome counts, ploidy level was determined using flow cytometry in 4- to 5 1/2 -year-old herbarium specimens and mature caryopses. Flow cytometric determination from seeds was more reliable than determination from herbarium specimens. In herbarium specimens, the youngest, fresh green leaves, still hidden in sheaths, seem to be most suitable for cytometric determination. In old, brownish leaves, or poorly preserved herbarium specimens, the degradation of DNA signal in flow histograms was documented. DNA content measured in seeds was always higher than that measured in herbarium specimens, which may be caused by the presence of different cytosolic compounds. Differences of about 15% in relative DNA content of F. sodiroana and F. vaginalis was found in simultaneous measurements in seeds.
-
Rydberg, Hanna A.; Matson, Maria; Å mand, Helene L.; Esbjö rner, Elin K.; Nordé n, Bengt
2012-01-01
Cell-penetrating peptides (CPPs) are able to traverse cellular membranes and deliver macromolecular cargo. Uptake occurs through both endocytotic and nonendocytotic pathways, but the molecular requirements for efficient internalization are not fully understood. Here we investigate how the presence of tryptophans and their position within an oligoarginine influence uptake mechanism and efficiency. Flow cytometry and confocal fluorescence imaging are used to estimate uptake efficiency, intracellular distribution and toxicity in Chinese hamster ovarian cells. Further, membrane leakage and lipid membrane affinity are investigated. The peptides contain eight arginine residues and one to four tryptophans, the tryptophans positioned either at the N-terminus, in the middle, or evenly distributed along the amino acid sequence. Our data show that the intracellular distribution varies among peptides with different tryptophan content and backbone spacing. Uptake efficiency is higher for the peptides with four tryptophans in the middle, or evenly distributed along the peptide sequence, than for the peptide with four tryptophans at the N-terminus. All peptides display low cytotoxicity except for the one with four tryptophans at the N-terminus, which was moderately toxic. This finding is consistent with their inability to induce efficient leakage of dye from lipid vesicles. All peptides have comparable affinities for lipid vesicles, showing that lipid binding is not a decisive parameter for uptake. Our results indicate that tryptophan content and backbone spacing can affect both the CPP uptake efficiency and the CPP uptake mechanism. The low cytotoxicity of these peptides and the possibilities of tuning their uptake mechanism are interesting from a therapeutic point of view. © 2012 American Chemical Society.
-
Rydberg, Hanna A.
2012-07-10
Cell-penetrating peptides (CPPs) are able to traverse cellular membranes and deliver macromolecular cargo. Uptake occurs through both endocytotic and nonendocytotic pathways, but the molecular requirements for efficient internalization are not fully understood. Here we investigate how the presence of tryptophans and their position within an oligoarginine influence uptake mechanism and efficiency. Flow cytometry and confocal fluorescence imaging are used to estimate uptake efficiency, intracellular distribution and toxicity in Chinese hamster ovarian cells. Further, membrane leakage and lipid membrane affinity are investigated. The peptides contain eight arginine residues and one to four tryptophans, the tryptophans positioned either at the N-terminus, in the middle, or evenly distributed along the amino acid sequence. Our data show that the intracellular distribution varies among peptides with different tryptophan content and backbone spacing. Uptake efficiency is higher for the peptides with four tryptophans in the middle, or evenly distributed along the peptide sequence, than for the peptide with four tryptophans at the N-terminus. All peptides display low cytotoxicity except for the one with four tryptophans at the N-terminus, which was moderately toxic. This finding is consistent with their inability to induce efficient leakage of dye from lipid vesicles. All peptides have comparable affinities for lipid vesicles, showing that lipid binding is not a decisive parameter for uptake. Our results indicate that tryptophan content and backbone spacing can affect both the CPP uptake efficiency and the CPP uptake mechanism. The low cytotoxicity of these peptides and the possibilities of tuning their uptake mechanism are interesting from a therapeutic point of view. © 2012 American Chemical Society.
-
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.
-
Optimization of reference library used in content-based medical image retrieval scheme
International Nuclear Information System (INIS)
Park, Sang Cheol; Sukthankar, Rahul; Mummert, Lily; Satyanarayanan, Mahadev; Zheng Bin
2007-01-01
Building an optimal image reference library is a critical step in developing the interactive computer-aided detection and diagnosis (I-CAD) systems of medical images using content-based image retrieval (CBIR) schemes. In this study, the authors conducted two experiments to investigate (1) the relationship between I-CAD performance and size of reference library and (2) a new reference selection strategy to optimize the library and improve I-CAD performance. The authors assembled a reference library that includes 3153 regions of interest (ROI) depicting either malignant masses (1592) or CAD-cued false-positive regions (1561) and an independent testing data set including 200 masses and 200 false-positive regions. A CBIR scheme using a distance-weighted K-nearest neighbor algorithm is applied to retrieve references that are considered similar to the testing sample from the library. The area under receiver operating characteristic curve (A z ) is used as an index to evaluate the I-CAD performance. In the first experiment, the authors systematically increased reference library size and tested I-CAD performance. The result indicates that scheme performance improves initially from A z =0.715 to 0.874 and then plateaus when the library size reaches approximately half of its maximum capacity. In the second experiment, based on the hypothesis that a ROI should be removed if it performs poorly compared to a group of similar ROIs in a large and diverse reference library, the authors applied a new strategy to identify 'poorly effective' references. By removing 174 identified ROIs from the reference library, I-CAD performance significantly increases to A z =0.914 (p<0.01). The study demonstrates that increasing reference library size and removing poorly effective references can significantly improve I-CAD performance
-
Directory of Open Access Journals (Sweden)
Cervantes Serena
2009-06-01
Full Text Available Abstract Background Malaria, a major public health issue in developing nations, is responsible for more than one million deaths a year. The most lethal species, Plasmodium falciparum, causes up to 90% of fatalities. Drug resistant strains to common therapies have emerged worldwide and recent artemisinin-based combination therapy failures hasten the need for new antimalarial drugs. Discovering novel compounds to be used as antimalarials is expedited by the use of a high-throughput screen (HTS to detect parasite growth and proliferation. Fluorescent dyes that bind to DNA have replaced expensive traditional radioisotope incorporation for HTS growth assays, but do not give additional information regarding the parasite stage affected by the drug and a better indication of the drug's mode of action. Live cell imaging with RNA dyes, which correlates with cell growth and proliferation, has been limited by the availability of successful commercial dyes. Results After screening a library of newly synthesized stryrl dyes, we discovered three RNA binding dyes that provide morphological details of live parasites. Utilizing an inverted confocal imaging platform, live cell imaging of parasites increases parasite detection, improves the spatial and temporal resolution of the parasite under drug treatments, and can resolve morphological changes in individual cells. Conclusion This simple one-step technique is suitable for automation in a microplate format for novel antimalarial compound HTS. We have developed a new P. falciparum RNA high-content imaging growth inhibition assay that is robust with time and energy efficiency.
-
Garbett, Kirsty M; Diedrichs, Phillippa C
2016-12-01
Mothers are a key influence on adolescent girls' body image. This study aimed to improve understanding of mothers' and daughters' preferences for content in body image interventions designed to assist mothers to promote positive body image among their daughters. British mother-daughter dyads (N=190) viewed descriptions of five evidence-based influences on body image (family, friends, and relationships; appearance-based teasing; media and celebrities; appearance conversations; body acceptance and care). Mothers and daughters each selected the two most important influences to learn about in these interventions. Overall, both mothers and daughters most frequently opted for family, friends, and relationships and body acceptance and care, whereas media and celebrities was their least preferred topic. While the overall sample of mothers and daughters agreed on preferences, Fisher's exact tests showed that within-dyad agreement was low. Recommendations for improving parent and child engagement with, and effectiveness of, child body image interventions delivered to parents are discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.
-
Noninvasive imaging of multiple myeloma using near infrared fluorescent molecular probe
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.
-
DEFF Research Database (Denmark)
Fernandes, Armando M.; Franco, Camilo; Mendes-Ferreira, Ana
2015-01-01
This work presents the results of measuring pH, sugars, and anthocyanin content of whole grape berries. The spectrum of each sample, composed of six whole grape berries, was collected using hyperspectral imaging in reflectance mode from 380 to 1028 nm. The spectra were converted to enological...... parameters by multilayer perceptrons created using 240 samples that were split for 7-fold cross-validation and test. The test set with 30 samples revealed R2 values of 0.73, 0.92 and 0.95 and RMSE of 0.18, 0.95 °Brix and 14 mg/l for pH, sugars and anthocyanin content, respectively. This is the only work, up...
-
Transformation invariant image indexing and retrieval for image databases
Gevers, Th.; Smeulders, A.W.M.
1994-01-01
This paper presents a novel design of an image database system which supports storage, indexing and retrieval of images by content. The image retrieval methodology is based on the observation that images can be discriminated by the presence of image objects and their spatial relations. Images in the
-
Profiling pleural effusion cells by a diffraction imaging method
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.
-
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.
-
Changes in distribution of cell cycle phases and DNA content in HeLa S3 cell after irradiation
International Nuclear Information System (INIS)
Wang Shunbao
1992-01-01
The effects of irradiation and hyperthermia on the distribution in various phases and DNA content of HeLa S 3 cells were analyzed by flow cytometry and an image analysis instrument. A marked increase in DNA content from 6.718 to 9.614(AU) in HeLa S 3 cells after 6 Gy irradiation was seen to correspond with the changes in the distribution of various phases in G 2 + M, from 22% to 52%. Meanwhile, the surviving fraction of HeLa S 3 cells after 6 Gy irradiation was less than 1%. However, after heating at 44 deg C for 10 min, the amount of cells in G 2 + M increased from 22.5% to 52.5% and the surviving fraction after hyperthermia was less than 2.65%. The changes in distribution of various phases after Ir-192 irradiation were similar to those seen after X-ray irradiation. The delay of G 2 + M phase after treatment with 8 Gy plus heating at 44 deg C for 7 min in HeLa S 3 cells was similar to that seen in the case of treatment with 8 Gy alone. As the surviving fraction accompanying the G 2 + M delay after irradiation plus heat treatment was very low, we suggest that the changes of distribution in various phases of HeLa S 3 cells after treatment might be used as a rapid indicator of serious injury
-
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
-
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
-
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.
-
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.
-
Directory of Open Access Journals (Sweden)
Ольга Геннадьевна Лопухова
2013-09-01
Full Text Available Purpose of the study is investigation of correlation between a content of gender appearance of image “successful person” and parameters of psychic and social adjustment of women belonging to different generations.Methodology. Subjective image “successful person” means a stable and possibly gender differentiated element of “Ideal Me” images system. It includes cognitive component (conscious and verbalization representations of typical description of successful person, and affective component (positive or negative positions in regards to image “successful person”. We have compared results of survey and valuation of 265 women belonging to different generations and staying in different social situations: involuntary unemployment situation, employment and getting professional education. Subjective and projective methods were used in survey of cognitive and affective components of “successful person” image. Valuation of psychic adjustment based on parameters of internal conflict in comparison with manifestations of anxiety, frustration, aggression, rigidity, neurotic. Data analysis used parametric and nonparametric methods, including ANOVA/MANOVA.Results. It has been discovered that level of psychic adjustment of women depends much more on proper integration of subjective “successful person” image content with individual features of self-conception than on objective “social status” (being in involuntary unemployment situation.Practical implications are psychology consulting and correction of social or psychic unadjustment.DOI: http://dx.doi.org/10.12731/2218-7405-2013-6-41
-
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.
-
Content-Based Information Retrieval from Forensic Databases
Geradts, Z.J.M.H.
2002-01-01
In forensic science, the number of image databases is growing rapidly. For this reason, it is necessary to have a proper procedure for searching in these images databases based on content. The use of image databases results in more solved crimes; furthermore, statistical information can be obtained
-
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.
-
International Nuclear Information System (INIS)
Plytycz, Barbara; Lis-Molenda, Urszula; Cygal, Malgorzata; Kielbasa, Edyta; Grebosz, Anna; Duchnowski, Michal; Andre, Jane; Morgan, A. John
2009-01-01
The effect of Pb + Zn on coelomocyte riboflavin content in the epigeic earthworm Dendrodrilus rubidus inhabiting three metalliferous soils and one reference soil was measured by flow cytometry and spectrofluorimetry. A reciprocal polluted↔unpolluted worm transfer experiment (4-week exposure) was also performed. High proportions of autofluorescent eleocytes were counted in worms from all localities, but intense riboflavin-derived autofluorescence was detectable only in reference worm eleocytes. Other findings were: (i) fluorophore(s) other than riboflavin is/are responsible for eleocyte autofluorescence in residents of metalliferous soils; (ii) riboflavin content was reduced in the eleocytes of worms transferred from unpolluted to metal-polluted soil; (iii) the riboflavin content of D. rubidus eleocytes is a promising biomarker of exposure; (iv) COII mitochondrial genotyping revealed that the reference population is genetically distinct from the three mine populations; (v) metal exposure rather than genotype is probably the main determinant of inter-population differences in eleocyte riboflavin status. - Soil metal pollution reduces riboflavin content of earthworm eleocytes.
-
Energy Technology Data Exchange (ETDEWEB)
Plytycz, Barbara, E-mail: barbara.plytycz@uj.edu.p [Institute of Zoology, Jagiellonian University, Ingardena 6, PL 30-060 Krakow (Poland); Lis-Molenda, Urszula; Cygal, Malgorzata; Kielbasa, Edyta; Grebosz, Anna [Institute of Zoology, Jagiellonian University, Ingardena 6, PL 30-060 Krakow (Poland); Duchnowski, Michal [Institute of Environmental Sciences, Jagiellonian University, Krakow (Poland); Andre, Jane; Morgan, A. John [Cardiff School of Biosciences, Main Building, Cardiff University, Cardiff CF10 3TL, Wales (United Kingdom)
2009-11-15
The effect of Pb + Zn on coelomocyte riboflavin content in the epigeic earthworm Dendrodrilus rubidus inhabiting three metalliferous soils and one reference soil was measured by flow cytometry and spectrofluorimetry. A reciprocal pollutedreversibleunpolluted worm transfer experiment (4-week exposure) was also performed. High proportions of autofluorescent eleocytes were counted in worms from all localities, but intense riboflavin-derived autofluorescence was detectable only in reference worm eleocytes. Other findings were: (i) fluorophore(s) other than riboflavin is/are responsible for eleocyte autofluorescence in residents of metalliferous soils; (ii) riboflavin content was reduced in the eleocytes of worms transferred from unpolluted to metal-polluted soil; (iii) the riboflavin content of D. rubidus eleocytes is a promising biomarker of exposure; (iv) COII mitochondrial genotyping revealed that the reference population is genetically distinct from the three mine populations; (v) metal exposure rather than genotype is probably the main determinant of inter-population differences in eleocyte riboflavin status. - Soil metal pollution reduces riboflavin content of earthworm eleocytes.
-
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.
-
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
-
DEFF Research Database (Denmark)
Munar, Ana Maria
2011-01-01
study of social media sites and destination brands, relying on qualitative research methods, content analysis and field research. Findings – Tourists are largely contributing to destination image formation, while avoiding the use of the formal elements of the brands. The most popular strategies used...
-
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
-
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
-
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
-
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.
-
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.
-
Content layer progressive coding of digital maps
DEFF Research Database (Denmark)
Forchhammer, Søren; Jensen, Ole Riis
2000-01-01
A new lossless context based method is presented for content progressive coding of limited bits/pixel images, such as maps, company logos, etc., common on the WWW. Progressive encoding is achieved by separating the image into content layers based on other predefined information. Information from...... already coded layers are used when coding subsequent layers. This approach is combined with efficient template based context bi-level coding, context collapsing methods for multi-level images and arithmetic coding. Relative pixel patterns are used to collapse contexts. The number of contexts are analyzed....... The new methods outperform existing coding schemes coding digital maps and in addition provide progressive coding. Compared to the state-of-the-art PWC coder, the compressed size is reduced to 60-70% on our layered test images....
-
Content Layer progressive Coding of Digital Maps
DEFF Research Database (Denmark)
Forchhammer, Søren; Jensen, Ole Riis
2002-01-01
A new lossless context based method is presented for content progressive coding of limited bits/pixel images, such as maps, company logos, etc., common on the World Wide Web. Progressive encoding is achieved by encoding the image in content layers based on color level or other predefined...... information. Information from already coded layers are used when coding subsequent layers. This approach is combined with efficient template based context bilevel coding, context collapsing methods for multilevel images and arithmetic coding. Relative pixel patterns are used to collapse contexts. Expressions...... for calculating the resulting number of contexts are given. The new methods outperform existing schemes coding digital maps and in addition provide progressive coding. Compared to the state-of-the-art PWC coder, the compressed size is reduced to 50-70% on our layered map test images....
-
Energy Technology Data Exchange (ETDEWEB)
Kurokawa, Hiroaki; Nakano, Yoshihisa; Ikeda, Koshi; Tanaka, Yoshimasa [Kansai Medical Univ., Moriguchi, Osaka (Japan); Fujisawa, Ichiro
1998-06-01
We investigated the correlation between the signal intensity on T{sub 1}-weighted MR images and vasopressin (VP) content in the posterior pituitary lobe. Fourteen rabbits were studied. There were 12 water-deprived rabbits (48, 72, 96, 120, 144 and 168 hours: 2 each) and 2 controls. Sagittal T{sub 1}-weighted SE (spin-echo) MR images were obtained before and after dehydration. The signal intensity ratio of the posterior pituitary lobe to the pons was correlated with the VP content in the posterior lobe as measured by radioimmunoassay. Before water deprivation, high signal intensity in the posterior lobe was demonstrated clearly in all 14 rabbits. After water deprivation, the hyperintense signal gradually decreased and became indistinguishable from anterior lobe in four animals. The mean signal intensity ratio before water deprivation was 1.55{+-}0.12 (mean{+-}SD) and after water deprivation, gradually decreased over time and reached to 1.19 after 168 hours of water deprivation. Pituitary VP content and concentration decreased in parallel with the signal intensity ratio of the posterior pituitary. Significantly correlation was observed between the signal intensity ratio and VP concentration of posterior pituitary (r=0.809, p<0.001) . In conclusion, the results indicate that the signal intensity ratio on T{sub 1}-weighted image may reflect a indicator of pituitary VP content and thus may enable evaluation of disorders of water metabolism. (author)
-
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.
-
Lago Cooke, Santiago Guillermo
2016-01-01
The current work entails the development of a novel high content platform for the measurement of kinetic ligand responses across cell signalling networks at the single-cell level in distinct PBMC subtypes ex vivo. Using automated sample preparation, fluorescent cellular barcoding and flow cytometry the platform is capable of detecting 21, 840 parallel cell signalling responses in each PBMC sample. We apply this platform to characterize the effects of neuropsychiatric treatments and CNS ligand...
-
Toward privacy-preserving JPEG image retrieval
Cheng, Hang; Wang, Jingyue; Wang, Meiqing; Zhong, Shangping
2017-07-01
This paper proposes a privacy-preserving retrieval scheme for JPEG images based on local variance. Three parties are involved in the scheme: the content owner, the server, and the authorized user. The content owner encrypts JPEG images for privacy protection by jointly using permutation cipher and stream cipher, and then, the encrypted versions are uploaded to the server. With an encrypted query image provided by an authorized user, the server may extract blockwise local variances in different directions without knowing the plaintext content. After that, it can calculate the similarity between the encrypted query image and each encrypted database image by a local variance-based feature comparison mechanism. The authorized user with the encryption key can decrypt the returned encrypted images with plaintext content similar to the query image. The experimental results show that the proposed scheme not only provides effective privacy-preserving retrieval service but also ensures both format compliance and file size preservation for encrypted JPEG images.
-
Magnetic Resonance Imaging (MRI) -- Head
Full Text Available ... Imaging (MRI) procedure View full size with caption Pediatric Content Some imaging tests and treatments have special pediatric considerations. The teddy bear denotes child-specific content. ...
-
International Nuclear Information System (INIS)
De Silva, T; Ketcha, M; Siewerdsen, J H; Uneri, A; Reaungamornrat, S; Vogt, S; Kleinszig, G; Lo, S F; Wolinsky, J P; Gokaslan, Z L; Aygun, N
2015-01-01
Purpose: In image-guided spine surgery, mapping 3D preoperative images to 2D intraoperative images via 3D-2D registration can provide valuable assistance in target localization. However, the presence of surgical instrumentation, hardware implants, and soft-tissue resection/displacement causes mismatches in image content, confounding existing registration methods. Manual/semi-automatic methods to mask such extraneous content is time consuming, user-dependent, error prone, and disruptive to clinical workflow. We developed and evaluated 2 novel similarity metrics within a robust registration framework to overcome such challenges in target localization. Methods: An IRB-approved retrospective study in 19 spine surgery patients included 19 preoperative 3D CT images and 50 intraoperative mobile radiographs in cervical, thoracic, and lumbar spine regions. A neuroradiologist provided truth definition of vertebral positions in CT and radiography. 3D-2D registration was performed using the CMA-ES optimizer with 4 gradient-based image similarity metrics: (1) gradient information (GI); (2) gradient correlation (GC); (3) a novel variant referred to as gradient orientation (GO); and (4) a second variant referred to as truncated gradient correlation (TGC). Registration accuracy was evaluated in terms of the projection distance error (PDE) of the vertebral levels. Results: Conventional similarity metrics were susceptible to gross registration error and failure modes associated with the presence of surgical instrumentation: for GI, the median PDE and interquartile range was 33.0±43.6 mm; similarly for GC, PDE = 23.0±92.6 mm respectively. The robust metrics GO and TGC, on the other hand, demonstrated major improvement in PDE (7.6 ±9.4 mm and 8.1± 18.1 mm, respectively) and elimination of gross failure modes. Conclusion: The proposed GO and TGC similarity measures improve registration accuracy and robustness to gross failure in the presence of strong image content mismatch. Such
-
Energy Technology Data Exchange (ETDEWEB)
De Silva, T; Ketcha, M; Siewerdsen, J H [Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD (United States); Uneri, A; Reaungamornrat, S [Department of Computer Science, Johns Hopkins University, Baltimore, MD (United States); Vogt, S; Kleinszig, G [Siemens Healthcare XP Division, Erlangen, DE (Germany); Lo, S F; Wolinsky, J P; Gokaslan, Z L [Department of Neurosurgery, The Johns Hopkins Hospital, Baltimore, MD (United States); Aygun, N [Department of Raiology and Radiological Sciences, The Johns Hopkins Hospital, Baltimore, MD (United States)
2015-06-15
Purpose: In image-guided spine surgery, mapping 3D preoperative images to 2D intraoperative images via 3D-2D registration can provide valuable assistance in target localization. However, the presence of surgical instrumentation, hardware implants, and soft-tissue resection/displacement causes mismatches in image content, confounding existing registration methods. Manual/semi-automatic methods to mask such extraneous content is time consuming, user-dependent, error prone, and disruptive to clinical workflow. We developed and evaluated 2 novel similarity metrics within a robust registration framework to overcome such challenges in target localization. Methods: An IRB-approved retrospective study in 19 spine surgery patients included 19 preoperative 3D CT images and 50 intraoperative mobile radiographs in cervical, thoracic, and lumbar spine regions. A neuroradiologist provided truth definition of vertebral positions in CT and radiography. 3D-2D registration was performed using the CMA-ES optimizer with 4 gradient-based image similarity metrics: (1) gradient information (GI); (2) gradient correlation (GC); (3) a novel variant referred to as gradient orientation (GO); and (4) a second variant referred to as truncated gradient correlation (TGC). Registration accuracy was evaluated in terms of the projection distance error (PDE) of the vertebral levels. Results: Conventional similarity metrics were susceptible to gross registration error and failure modes associated with the presence of surgical instrumentation: for GI, the median PDE and interquartile range was 33.0±43.6 mm; similarly for GC, PDE = 23.0±92.6 mm respectively. The robust metrics GO and TGC, on the other hand, demonstrated major improvement in PDE (7.6 ±9.4 mm and 8.1± 18.1 mm, respectively) and elimination of gross failure modes. Conclusion: The proposed GO and TGC similarity measures improve registration accuracy and robustness to gross failure in the presence of strong image content mismatch. Such
-
Short-term memory for scenes with affective content
Maljkovic, Vera; Martini, Paolo
2005-01-01
The emotional content of visual images can be parameterized along two dimensions: valence (pleasantness) and arousal (intensity of emotion). In this study we ask how these distinct emotional dimensions affect the short-term memory of human observers viewing a rapid stream of images and trying to remember their content. We show that valence and arousal modulate short-term memory as independent factors. Arousal influences dramatically the average speed of data accumulation in memory: Higher aro...
-
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
-
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.
-
Interactive classification and content-based retrieval of tissue images
Aksoy, Selim; Marchisio, Giovanni B.; Tusk, Carsten; Koperski, Krzysztof
2002-11-01
We describe a system for interactive classification and retrieval of microscopic tissue images. Our system models tissues in pixel, region and image levels. Pixel level features are generated using unsupervised clustering of color and texture values. Region level features include shape information and statistics of pixel level feature values. Image level features include statistics and spatial relationships of regions. To reduce the gap between low-level features and high-level expert knowledge, we define the concept of prototype regions. The system learns the prototype regions in an image collection using model-based clustering and density estimation. Different tissue types are modeled using spatial relationships of these regions. Spatial relationships are represented by fuzzy membership functions. The system automatically selects significant relationships from training data and builds models which can also be updated using user relevance feedback. A Bayesian framework is used to classify tissues based on these models. Preliminary experiments show that the spatial relationship models we developed provide a flexible and powerful framework for classification and retrieval of tissue images.
-
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.
-
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
-
Alibhai, Dominic; Kumar, Sunil; Kelly, Douglas; Warren, Sean; Alexandrov, Yuriy; Munro, Ian; McGinty, James; Talbot, Clifford; Murray, Edward J.; Stuhmeier, Frank; Neil, Mark A. A.; Dunsby, Chris; French, Paul M. W.
2011-03-01
We describe an optically-sectioned FLIM multiwell plate reader that combines Nipkow microscopy with wide-field time-gated FLIM, and its application to high content analysis of FRET. The system acquires sectioned FLIM images in fluorescent protein. It has been applied to study the formation of immature HIV virus like particles (VLPs) in live cells by monitoring Gag-Gag protein interactions using FLIM FRET of HIV-1 Gag transfected with CFP or YFP. VLP formation results in FRET between closely packed Gag proteins, as confirmed by our FLIM analysis that includes automatic image segmentation.
-
García Carrillo, Mercedes; Ferrario, Mariana; Guerrero, Sandra
2018-08-01
The aim of this study was to analyze the effectiveness of UV-C light (0-10.6 kJ/m 2 ) assisted by mild heat treatment (50 °C) on the inactivation of Saccharomyces cerevisiae KE 162 in peptone water and fresh carrot-orange juice blend (pH: 3.8; 9.8°Brix; 707 NTU; absorption coefficient: 0.17 cm -1 ). Yeast induced damage by single UV-C and mild heat (H) and the combined treatment UV-C/H, was investigated by flow cytometry (FC) and transmission electron microscopy (TEM). When studying induced damage by FC, cells were labeled with fluorescein diacetate (FDA) and propidium iodide (PI) to monitor membrane integrity and esterase activity. UV-C/H provoked up to 4.7 log-reductions of S. cerevisiae; whereas, only 2.6-3.3 log-reductions were achieved by single UV-C and H treatments. FC revealed a shift with treatment time from cells with esterase activity and intact membrane to cells with permeabilized membrane. This shift was more noticeable in peptone water and UV-C/H treated juice. In the UV-C treated juice, double stained cells were detected, suggesting the possibility of being sub-lethally damaged, with compromised membrane but still metabolically active. TEM images of treated cells revealed severe damage, encompassing coagulated inner content, disorganized lumen and cell debris. FC and TEM provided additional information regarding degree and type of damage, complementing information revealed by the traditional plate count technique. Copyright © 2018 Elsevier Ltd. All rights reserved.
-
High throughput imaging cytometer with acoustic focussing.
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.
-
Directory of Open Access Journals (Sweden)
Tiziana de-Magistris
2016-12-01
Full Text Available The aim of this study is to assess the influence of body image on consumers’ willingness to pay (WTP for potato chips carrying nutritional claims among obese and non-obese people. About 309 non-clinical individuals participated in a Real Choice Experiment. They were recruited by a company and grouped in: (i non-obese with good body image; (ii non-obese with body image dissatisfaction; (iii obese with good body image; (iv obese with body image dissatisfaction. Results indicate differences in consumers’ willingness to pay among consumer groups. Body image dissatisfaction of normal people did not influence the WTP for healthier chips. Obese people with body image dissatisfaction were willing to pay more for healthier chips (i.e., low-salt content potato chips than normal ones with body image dissatisfaction. Examining the role of knowledge in the light of how this could impact on body image is relevant to improve the health status of individuals and their diet. Knowledge about nutrition could improve the body image of obese people.
-
de-Magistris, Tiziana; López-Galán, Belinda; Caputo, Vincenzina
2016-12-21
The aim of this study is to assess the influence of body image on consumers' willingness to pay (WTP) for potato chips carrying nutritional claims among obese and non-obese people. About 309 non-clinical individuals participated in a Real Choice Experiment. They were recruited by a company and grouped in: (i) non-obese with good body image; (ii) non-obese with body image dissatisfaction; (iii) obese with good body image; (iv) obese with body image dissatisfaction. Results indicate differences in consumers' willingness to pay among consumer groups. Body image dissatisfaction of normal people did not influence the WTP for healthier chips. Obese people with body image dissatisfaction were willing to pay more for healthier chips (i.e., low-salt content potato chips) than normal ones with body image dissatisfaction. Examining the role of knowledge in the light of how this could impact on body image is relevant to improve the health status of individuals and their diet. Knowledge about nutrition could improve the body image of obese people.