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Sample records for dual-laser flow cytometry

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

    Science.gov (United States)

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

    2014-07-01

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

  2. Detection and quantification of live, apoptotic, and necrotic human peripheral lymphocytes by single-laser flow cytometry.

    Science.gov (United States)

    Liegler, T J; Hyun, W; Yen, T S; Stites, D P

    1995-05-01

    Regulation of peripheral lymphocyte number involves a poorly understood balance between cell renewal and loss. Disrupting this balance leads to a large number of disease states. Methods which allow qualitative and quantitative measurements of cell viability are increasingly valuable to studies directed at revealing the mechanisms underlying apoptotic and necrotic cell death. Here, we have characterized a method using single-laser flow cytometry that differentiates and quantifies the relative number of live, apoptotic, and late-stage apoptotic and necrotic peripheral lymphocytes. Following in vitro gamma irradiation and staining with acridine orange in combination with ethidium bromide, three distinct populations were seen by bivariate analysis of green versus red fluorescence. The identity of each distinct fluorescent population (whether live, apoptotic, or necrotic) was determined by sorting and examination of cellular morphology by electron microscopy. This flow cytometric method is directly compared with the techniques of trypan blue exclusion and DNA fragmentation to quantify cell death following exposure to various doses of in vitro gamma irradiation and postirradiation incubation times. We extend our findings to illustrate the utility of this method beyond analyzing radiation-induced apoptotic peripheral blood mononuclear cells (PBMC); similar fluorescent patterns are shown for radiation- and corticosteroid-treated murine thymocytes, activated human PBMC, and PBMC from human immunodeficiency virus-infected individuals. Our results demonstrate that dual-parameter flow cytometric analysis of acridine orange-ethidium bromide-stained lymphocytes is overall a superior method with increased sensitivity, greater accuracy, and decreased subjectivity in comparison with the other methods tested. By using standard laser and filter settings commonly available to flow cytometric laboratories, this method allows rapid measurement of a large number of cells from a

  3. Practical flow cytometry

    National Research Council Canada - National Science Library

    Shapiro, Howard M

    2003-01-01

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

  4. Hyperchromatic laser scanning cytometry

    Science.gov (United States)

    Tárnok, Attila; Mittag, Anja

    2007-02-01

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

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

    Science.gov (United States)

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

    2014-12-01

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

  6. Femtosecond laser fabrication of fiber based optofluidic platform for flow cytometry applications

    Science.gov (United States)

    Serhatlioglu, Murat; Elbuken, Caglar; Ortac, Bulend; Solmaz, Mehmet E.

    2017-02-01

    Miniaturized optofluidic platforms play an important role in bio-analysis, detection and diagnostic applications. The advantages of such miniaturized devices are extremely low sample requirement, low cost development and rapid analysis capabilities. Fused silica is advantageous for optofluidic systems due to properties such as being chemically inert, mechanically stable, and optically transparent to a wide spectrum of light. As a three dimensional manufacturing method, femtosecond laser scanning followed by chemical etching shows great potential to fabricate glass based optofluidic chips. In this study, we demonstrate fabrication of all-fiber based, optofluidic flow cytometer in fused silica glass by femtosecond laser machining. 3D particle focusing was achieved through a straightforward planar chip design with two separately fabricated fused silica glass slides thermally bonded together. Bioparticles in a fluid stream encounter with optical interrogation region specifically designed to allocate 405nm single mode fiber laser source and two multi-mode collection fibers for forward scattering (FSC) and side scattering (SSC) signals detection. Detected signal data collected with oscilloscope and post processed with MATLAB script file. We were able to count number of events over 4000events/sec, and achieve size distribution for 5.95μm monodisperse polystyrene beads using FSC and SSC signals. Our platform shows promise for optical and fluidic miniaturization of flow cytometry systems.

  7. Cell proliferation and apoptosis in the primary enamel knot measured by flow cytometry of laser microdissected samples

    Czech Academy of Sciences Publication Activity Database

    Matalová, Eva; Dubská, L.; Fleischmannová, Jana; Chlastáková, Ivana; Janečková, Eva; Tucker, A. S.

    2010-01-01

    Roč. 55, č. 8 (2010), s. 570-575 ISSN 0003-9969 R&D Projects: GA AV ČR KJB500450802; GA AV ČR IAA600450904; GA ČR GA203/08/1680 Institutional research plan: CEZ:AV0Z50450515 Keywords : Laser capture microdissection * Flow cytometry * Primary enamel knot Subject RIV: EA - Cell Biology Impact factor: 1.463, year: 2010

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

    Energy Technology Data Exchange (ETDEWEB)

    Jett, J.H.

    1995-04-27

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

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

    Science.gov (United States)

    Sun, Q; Ding, Y; Zhang, J

    2001-01-25

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

  10. Microfluidic devices and methods for integrated flow cytometry

    Science.gov (United States)

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

    2011-08-16

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

  11. 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.

  12. Flow Cytometry Section

    Data.gov (United States)

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

  13. 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....

  14. Flow cytometry protocols

    National Research Council Canada - National Science Library

    Jaroszeski, Mark J; Heller, Richard

    1998-01-01

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

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

    Science.gov (United States)

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

    2017-06-28

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

  16. Flow cytometry in diagnostic cytology.

    Science.gov (United States)

    O'Leary, T J

    1998-01-01

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

  17. Flow Cytometry Technician | Center for Cancer Research

    Science.gov (United States)

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

  18. Sensitivity of hemozoin detection by automated flow cytometry in non- and semi-immune malaria patients

    NARCIS (Netherlands)

    Grobusch, Martin P.; Hänscheid, Thomas; Krämer, Benedikt; Neukammer, Jörg; May, Jürgen; Seybold, Joachim; Kun, Jürgen F. J.; Suttorp, Norbert

    2003-01-01

    BACKGROUND: Cell-Dyn automated blood cell analyzers use laser flow cytometry technology, allowing detection of malaria pigment (hemozoin) in monocytes. We evaluated the value of such an instrument to diagnose malaria in febrile travelers returning to Berlin, Germany, the relation between the

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

    Science.gov (United States)

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

    2016-06-01

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

  20. 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

  1. 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.

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

    Science.gov (United States)

    Spidlen, Josef; Breuer, Karin; Brinkman, Ryan

    2012-07-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

  4. Immuno flow cytometry in marine phytoplankton research

    NARCIS (Netherlands)

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

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

  5. Preparation and flow cytometry of uniform silica-fluorescent dye microspheres.

    Science.gov (United States)

    Bele, Marjan; Siiman, Olavi; Matijević, Egon

    2002-10-15

    Uniform fluorescent silica-dye microspheres have been prepared by coating preformed monodispersed silica particles with silica layers containing rhodamine 6G or acridine orange. The resulting dispersions exhibit intense fluorescent emission between 500 and 600 nm, over a broad excitation wavelength range of 460 to 550 nm, even with exceedingly small amounts of dyes incorporated into the silica particles (10-30 ppm, expressed as weight of dye relative to weight of dry particles). The fluorescent particles can be prepared in micrometer diameters suitable for analyses using flow cytometry with 488-nm laser excitation.

  6. Spaceflight Flow Cytometry: Design Challenges and Applications

    Science.gov (United States)

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

    2004-01-01

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

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

    Science.gov (United States)

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

    2017-12-19

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

  8. 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

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

    Science.gov (United States)

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

    2009-06-16

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

  10. Analysis of UV-excited fluorochromes by flow cytometry using near-ultraviolet laser diodes.

    Science.gov (United States)

    Telford, William G

    2004-09-01

    Violet laser diodes have become common and reliable laser sources for benchtop flow cytometers. While these lasers are very useful for a variety of violet and some ultraviolet-excited fluorochromes (e.g., DAPI), they do not efficiently excite most UV-stimulated probes. In this study, the next generation of InGaN near-UV laser diodes (NUVLDs) emitting in the 370-375-nm range have been evaluated as laser sources for cuvette-based flow cytometers. Several NUVLDs, ranging in wavelength from 370 to 374 nm and in power level from 1.5 to 10 mW, were mounted on a BD Biosciences LSR II and evaluated for their ability to excite cells labeled with the UV DNA binding dye DAPI, several UV phenotyping fluorochromes (including Alexa Fluor 350, Marina Blue, and quantum dots), and the fluorescent calcium chelator indo-1. NUVLDs at the 8-10-mW power range gave detection sensitivity levels comparable to more powerful solid-state and ion laser sources, using low-fluorescence microsphere beads as measurement standards. NUVLDs at all tested power levels allowed extremely high-resolution DAPI cell cycle analysis, and sources in the 8-10-mW power range excited Alexa Fluor 350, Marina Blue, and a variety of quantum dots at virtually the same signal-to-noise ratios as more powerful UV sources. These evaluations indicate that near-UV laser diodes installed on a cuvette-based flow cytometer performed nearly as well as more powerful solid-state UV lasers on the same instrumentation, and comparably to more powerful ion lasers on a jet-in-air system, and. Despite their limited power, integration of these small and inexpensive lasers into benchtop flow cytometers should allow the use of flow cytometric applications requiring UV excitation on a wide variety of instruments. Copyright 2004 Wiley-Liss, Inc.

  11. 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

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

    Directory of Open Access Journals (Sweden)

    Michael S Bono

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

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

    Science.gov (United States)

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

    2015-01-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

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

    Science.gov (United States)

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

    2013-01-31

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

  16. 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

  17. Photoacoustic-fluorescence in vitro flow cytometry for quantification of absorption, scattering and fluorescence properties of the cells

    Science.gov (United States)

    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.

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1992-06-01

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

  20. Flow cytometry, fluorescent probes, and flashing bacteria

    NARCIS (Netherlands)

    Bunthof, C.J.

    2002-01-01


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

  1. 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)

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

    Science.gov (United States)

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

    2012-07-01

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

  3. Sorting catalytically active polymersome nanoreactors by flow cytometry

    NARCIS (Netherlands)

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

    2009-01-01

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

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

    Science.gov (United States)

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

    1999-01-01

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

  5. Method of detaching adherent cells for flow cytometry

    KAUST Repository

    Kaur, Mandeep

    2015-12-24

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

  6. 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 ...

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

    Science.gov (United States)

    Rosebrock, Adam P

    2017-01-03

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

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

    Science.gov (United States)

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

    2017-11-01

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

  9. Analysis of repetitive DNA in chromosomes by flow cytometry

    NARCIS (Netherlands)

    Brind'Amour, Julie; Lansdorp, Peter M.

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

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

    Science.gov (United States)

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

    2013-01-01

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

  11. 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.

  12. Development of a Dual-PIV system for high-speed flow applications

    Science.gov (United States)

    Schreyer, Anne-Marie; Lasserre, Jean J.; Dupont, Pierre

    2015-10-01

    A new Dual-particle image velocimetry (Dual-PIV) system for application in supersonic flows was developed. The system was designed for shock wave/turbulent boundary layer interactions with separation. This type of flow places demanding requirements on the system, from the large range of characteristic frequencies O(100 Hz-100 kHz) to spatial and temporal resolutions necessary for the measurement of turbulent quantities (Dolling in AIAA J 39(8):1517-1531, 2001; Dupont et al. in J Fluid Mech 559:255-277, 2006; Smits and Dussauge in Turbulent shear layers in supersonic flow, 2nd edn. Springer, New York, 2006). While classic PIV systems using high-resolution CCD sensors allow high spatial resolution, these systems cannot provide the required temporal resolution. Existing high-speed PIV systems provide temporal and CMOS sensor resolutions, and even laser pulse energies, that are not adapted to our needs. The only obvious solution allowing sufficiently high spatial resolution, access to high frequencies, and a high laser pulse energy is a multi-frame system: a Dual-PIV system, consisting of two synchronized PIV systems observing the same field of view, will give access to temporal characteristics of the flow. The key technology of our system is frequency-based image separation: two lasers of different wavelengths illuminate the field of view. The cross-pollution with laser light from the respective other branches was quantified during system validation. The overall system noise was quantified, and the prevailing error of only 2 % reflects the good spatial and temporal alignment. The quality of the measurement system is demonstrated with some results on a subsonic jet flow including the spatio-temporal inter-correlation functions between the systems. First measurements in a turbulent flat-plate boundary layer at Mach 2 show the same satisfactory data quality and are also presented and discussed.

  13. 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.

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

    Science.gov (United States)

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

    2014-01-01

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

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

    Science.gov (United States)

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

    2010-01-01

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

  16. Flow Cytometry of the Side Population: Tips & Tricks

    Directory of Open Access Journals (Sweden)

    Irene Sales-Pardo

    2006-01-01

    Full Text Available Background: The Side Population (SP has become an important hallmark for the definition of the stem cell compartment, especially in the detection of these cells and in their physical isolation by fluorescence-activated cell sorting (FACS. SP cells are CD34neg and were discovered using ultraviolet excitation based on the efflux of Hoechst 33342 (Ho342. Although the method works as originally described, we believe that this method is difficult for most investigators. First, because the ability to discriminate SP cells is based on the differential retention of Ho342 during a functional assay; second, because of the difficulties in setting the right experimental and acquisition conditions; and third, because the analysis of the acquired data requires an extensive expertise on flow cytometry to accurately detect the SP events. Methods: First of all and mainly for the SP application, the laser beam paths were exhaustively checked to ensure the lowest coefficients of variation. Blood suspensions were prepared by erythrocyte lysis with ammonium chloride and hematopoietic cells were labeled with Ho342. Results: The Ho342 concentration and the staining procedure are critical for the optimal resolution of the SP cells. Although UV laser alignment is very important to resolve the dim tail that outlines the SP, the problem with Ho342 excitation is not the Hoechst Blue emission, but rather the Hoechst Red's (because of the weak emission. Conclusions: Each laboratory must establish its own expected ranges based on its instrument and results may vary slightly due to instrument differences such as the narrowness of the band pass filters, laser power, laser emission wavelength, nozzle type, differential of pressure, light collection system (cuvette versus jet-in-air and beam shaping optics.

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

    Science.gov (United States)

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

    2017-08-01

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

  18. Laser doppler velocimetry and confined flows

    Directory of Open Access Journals (Sweden)

    Ilić Jelena T.

    2017-01-01

    Full Text Available Finding the mode, in which two component laser Doppler velocimetry can be applied to flows confined in cylindrical tubes or vessels, was the aim of this study. We have identified principle issues that influence the propagation of laser beams in laser Doppler velocimetry system, applied to flow confined in cylindrical tube. Among them, the most important are influences of fluid and wall refractive indices, wall thickness and internal radius ratio and beam intersection angle. In analysis of the degrees of these influences, we have applied mathematical model, based on geometrical optics. The separation of measurement volumes, that measure different velocity components, has been recognized as the main drawback. To overcome this, we propose a lens with dual focal length – primary focal length for the measurement of one velocity component and secondary focal length for the measurement of the other velocity component. We present here the procedure for calculating the optimal value of secondary focal length, depending on experimental set-up parameters. The mathematical simulation of the application of the dual focal length lens, for chosen cases presented here, confirmed the accuracy of the proposed procedure.

  19. 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.

  20. 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

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

    Czech Academy of Sciences Publication Activity Database

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

    2004-01-01

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

  2. Flow Cytometry Pulse Width Data Enables Rapid and Sensitive Estimation of Biomass Dry Weight in the Microalgae Chlamydomonas reinhardtii and Chlorella vulgaris

    Science.gov (United States)

    Chioccioli, Maurizio; Hankamer, Ben; Ross, Ian L.

    2014-01-01

    Dry weight biomass is an important parameter in algaculture. Direct measurement requires weighing milligram quantities of dried biomass, which is problematic for small volume systems containing few cells, such as laboratory studies and high throughput assays in microwell plates. In these cases indirect methods must be used, inducing measurement artefacts which vary in severity with the cell type and conditions employed. Here, we utilise flow cytometry pulse width data for the estimation of cell density and biomass, using Chlorella vulgaris and Chlamydomonas reinhardtii as model algae and compare it to optical density methods. Measurement of cell concentration by flow cytometry was shown to be more sensitive than optical density at 750 nm (OD750) for monitoring culture growth. However, neither cell concentration nor optical density correlates well to biomass when growth conditions vary. Compared to the growth of C. vulgaris in TAP (tris-acetate-phosphate) medium, cells grown in TAP + glucose displayed a slowed cell division rate and a 2-fold increased dry biomass accumulation compared to growth without glucose. This was accompanied by increased cellular volume. Laser scattering characteristics during flow cytometry were used to estimate cell diameters and it was shown that an empirical but nonlinear relationship could be shown between flow cytometric pulse width and dry weight biomass per cell. This relationship could be linearised by the use of hypertonic conditions (1 M NaCl) to dehydrate the cells, as shown by density gradient centrifugation. Flow cytometry for biomass estimation is easy to perform, sensitive and offers more comprehensive information than optical density measurements. In addition, periodic flow cytometry measurements can be used to calibrate OD750 measurements for both convenience and accuracy. This approach is particularly useful for small samples and where cellular characteristics, especially cell size, are expected to vary during growth. PMID

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-12-04

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

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

    International Nuclear Information System (INIS)

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

    2013-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Pipy Bernard

    2010-02-01

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

  6. 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 (

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

    Science.gov (United States)

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

    2018-02-01

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

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

    Science.gov (United States)

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

    2010-11-04

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

  9. 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

  10. Honey bee hemocyte profiling by flow cytometry.

    Science.gov (United States)

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

    2014-01-01

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

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

    Science.gov (United States)

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

    2016-05-01

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

  12. Lipid nanoparticles assessment by flow cytometry.

    Science.gov (United States)

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

    2017-03-30

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

  13. Dual frequency comb metrology with one fiber laser

    Science.gov (United States)

    Zhao, Xin; Takeshi, Yasui; Zheng, Zheng

    2016-11-01

    Optical metrology techniques based on dual optical frequency combs have emerged as a hotly studied area targeting a wide range of applications from optical spectroscopy to microwave and terahertz frequency measurement. Generating two sets of high-quality comb lines with slightly different comb-tooth spacings with high mutual coherence and stability is the key to most of the dual-comb schemes. The complexity and costs of such laser sources and the associated control systems to lock the two frequency combs hinder the wider adoption of such techniques. Here we demonstrate a very simple and rather different approach to tackle such a challenge. By employing novel laser cavity designs in a mode-locked fiber laser, a simple fiber laser setup could emit dual-comb pulse output with high stability and good coherence between the pulse trains. Based on such lasers, comb-tooth-resolved dual-comb optical spectroscopy is demonstrated. Picometer spectral resolving capability could be realized with a fiber-optic setup and a low-cost data acquisition system and standard algorithms. Besides, the frequency of microwave signals over a large range can be determined based on a simple setup. Our results show the capability of such single-fiber-laser-based dual-comb scheme to reduce the complexity and cost of dual-comb systems with excellent quality for different dual-comb applications.

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

    Science.gov (United States)

    Grimberg, Brian T

    2011-03-31

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

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

    NARCIS (Netherlands)

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

    1994-01-01

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

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

    Directory of Open Access Journals (Sweden)

    M.J. Najafzadeh

    2009-08-01

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

  17. Multi-channel imaging cytometry with a single detector

    Science.gov (United States)

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

    2018-02-01

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

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

    DEFF Research Database (Denmark)

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

    2008-01-01

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

  19. Adaptive sampling dual terahertz comb spectroscopy using dual free-running femtosecond lasers.

    Science.gov (United States)

    Yasui, Takeshi; Ichikawa, Ryuji; Hsieh, Yi-Da; Hayashi, Kenta; Cahyadi, Harsono; Hindle, Francis; Sakaguchi, Yoshiyuki; Iwata, Tetsuo; Mizutani, Yasuhiro; Yamamoto, Hirotsugu; Minoshima, Kaoru; Inaba, Hajime

    2015-06-02

    Terahertz (THz) dual comb spectroscopy (DCS) is a promising method for high-accuracy, high-resolution, broadband THz spectroscopy because the mode-resolved THz comb spectrum includes both broadband THz radiation and narrow-line CW-THz radiation characteristics. In addition, all frequency modes of a THz comb can be phase-locked to a microwave frequency standard, providing excellent traceability. However, the need for stabilization of dual femtosecond lasers has often hindered its wide use. To overcome this limitation, here we have demonstrated adaptive-sampling THz-DCS, allowing the use of free-running femtosecond lasers. To correct the fluctuation of the time and frequency scales caused by the laser timing jitter, an adaptive sampling clock is generated by dual THz-comb-referenced spectrum analysers and is used for a timing clock signal in a data acquisition board. The results not only indicated the successful implementation of THz-DCS with free-running lasers but also showed that this configuration outperforms standard THz-DCS with stabilized lasers due to the slight jitter remained in the stabilized lasers.

  20. 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

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

    Science.gov (United States)

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

    2017-11-01

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

  2. 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

  3. Measurement and Characterization of Apoptosis by Flow Cytometry.

    Science.gov (United States)

    Telford, William; Tamul, Karen; Bradford, Jolene

    2016-07-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Science.gov (United States)

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

    2007-01-01

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

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

    Science.gov (United States)

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

    2009-01-01

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

  7. 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.

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

    Directory of Open Access Journals (Sweden)

    Muzaffer Keklik

    2012-10-01

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

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

    Directory of Open Access Journals (Sweden)

    Lo Kenneth

    2009-05-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

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

    Science.gov (United States)

    Zhu, Hongying; Ozcan, Aydogan

    2013-04-11

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

  12. 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.

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

    DEFF Research Database (Denmark)

    Tanev, Stoyan; Sun, Wenbo; Pond, James

    2009-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Federica Villanova

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

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

    Science.gov (United States)

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

    2013-01-01

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

  16. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods.

    Science.gov (United States)

    Øbro, Nina F; Ryder, Lars P; Madsen, Hans O; Andersen, Mette K; Lausen, Birgitte; Hasle, Henrik; Schmiegelow, Kjeld; Marquart, Hanne V

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and/or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative immunophenotype and antigen modulation) that highlight important methodological pitfalls. These findings demonstrate that with sufficient experience, flow cytometry is reliable for minimal residual disease monitoring in B-cell precursor acute lymphoblastic leukemia, although rare cases require supplementary PCR-based monitoring.

  17. 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.

  18. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods

    DEFF Research Database (Denmark)

    Obro, Nina F; Ryder, Lars P; Madsen, Hans O

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring...... clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and....../or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative...

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

    Science.gov (United States)

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

    2016-06-01

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

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

    Science.gov (United States)

    Acosta, Maria; Pereira, José; Arroz, Maria

    2016-05-01

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

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

    Science.gov (United States)

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

    2010-04-01

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

  2. 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.

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

    Science.gov (United States)

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

    2013-06-01

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

  4. 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)

  5. Simultaneous ultrasound and photoacoustics based flow cytometry

    Science.gov (United States)

    Gnyawali, Vaskar; Strohm, Eric M.; Tsai, Scott S. H.; Kolios, Michael C.

    2018-04-01

    We have developed a flow cytometer based on simultaneous detection of ultrasound and photoacoustic waves from individual particles/cells flowing in a microfluidic channel. Our polydimethylsiloxane (PDMS) based hydrodynamic 3-dimensional (3D) flow-focusing microfluidic device contains a cross-junction channel, a micro-needle (ID 100 μm and OD 200 μm) insert, and a 3D printed frame to hold and align a high frequency (center frequency 375 MHz) ultrasound transducer. The focused flow passes through a narrow focal zone with lateral and axial focal lengths of 6-8 μm and 15-20 μm, respectively. Both the lateral and axial alignments are achieved by screwing the transducer to the frame onto the PDMS device. Individual particles pass through an interrogation zone in the microfluidic channel with a collinearly aligned ultrasound transducer and a focused 532 nm wavelength laser beam. The particles are simultaneously insonified by high-frequency ultrasound and irradiated by a laser beam. The ultrasound backscatter and laser generated photoacoustic waves are detected for each passing particle. The backscattered ultrasound and photoacoustic signal are strongly dependent on the size, morphology, mechanical properties, and material properties of the flowing particles; these parameters can be extracted by analyzing unique features in the power spectrum of the signals. Frequencies less than 100 MHz do not have these unique spectral signatures. We show that we can reliably distinguish between different particles in a sample using the acoustic-based flow cytometer. This technique, when extended to biomedical applications, allows us to rapidly analyze the spectral signatures from individual single cells of a large cell population, with applications towards label-free detection and characterization of healthy and diseased cells.

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

    NARCIS (Netherlands)

    Mengarelli, Isabella; Fryga, Andrew; Barberi, Tiziano

    2016-01-01

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

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

    Science.gov (United States)

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

    2016-02-01

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

  8. 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)

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Science.gov (United States)

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

    1990-01-01

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

  11. 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

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

    OpenAIRE

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

    2013-01-01

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

  13. 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.

  14. 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.

  15. 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

  16. Switchable dual-wavelength single-longitudinal-mode erbium fiber laser utilizing a dual-ring scheme with a saturable absorber

    Science.gov (United States)

    Yang, Zi-Qing; Huang, Tzu-Jung; Chang, Yao-Jen; Yeh, Chien-Hung; Chow, Chi-Wai; Chen, Jing-Heng; Chen, Kun-Huang

    2018-06-01

    In this work, we propose and demonstrate a switchable dual-wavelength erbium-doped fiber (EDF) ring laser with stable single-longitudinal-mode (SLM) output. Here, a dual-ring (DR) structure with an unpumped EDF of 2 m is designed to achieve SLM oscillation. Five fiber Bragg gratings (FBGs) are applied in the laser cavity serving as the reflective element to generate different dual-wavelength outputs. In the measurement, six sets of generated dual-wavelengths with various mode-spacing (Δλ) can be achieved via the five FBGs. Additionally, the stability performance of the proposed EDF DR laser is also demonstrated.

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

    Directory of Open Access Journals (Sweden)

    Carmen E Contreras

    2004-03-01

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

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

    Science.gov (United States)

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

    2001-01-01

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

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

    Science.gov (United States)

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

    2008-12-01

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

  20. Picometer-resolution dual-comb spectroscopy with a free-running fiber laser.

    Science.gov (United States)

    Zhao, Xin; Hu, Guoqing; Zhao, Bofeng; Li, Cui; Pan, Yingling; Liu, Ya; Yasui, Takeshi; Zheng, Zheng

    2016-09-19

    Dual-comb spectroscopy holds the promise as real-time, high-resolution spectroscopy tools. However, in its conventional schemes, the stringent requirement on the coherence between two lasers requires sophisticated control systems. By replacing control electronics with an all-optical dual-comb lasing scheme, a simplified dual-comb spectroscopy scheme is demonstrated using one dual-wavelength, passively mode-locked fiber laser. Pulses with a intracavity-dispersion-determined repetition-frequency difference are shown to have good mutual coherence and stability. Capability to resolve the comb teeth and a picometer-wide optical spectral resolution are demonstrated using a simple data acquisition system. Energy-efficient, free-running fiber lasers with a small comb-tooth-spacing could enable low-cost dual-comb systems.

  1. 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.

  2. Hyperspectral cytometry.

    Science.gov (United States)

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

    2014-01-01

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

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

    Science.gov (United States)

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

    2009-09-01

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

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

    NARCIS (Netherlands)

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

    2004-01-01

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

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

    Science.gov (United States)

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

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

    Directory of Open Access Journals (Sweden)

    Josep M. Gasol

    2000-06-01

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

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

    NARCIS (Netherlands)

    W.H.B.M. Levering

    2007-01-01

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

  8. Drilling of Copper Using a Dual-Pulse Femtosecond Laser

    Directory of Open Access Journals (Sweden)

    Chung-Wei Cheng

    2016-02-01

    Full Text Available The drilling of copper using a dual-pulse femtosecond laser with wavelength of 800 nm, pulse duration of 120 fs and a variable pulse separation time (0.1–150 ps is investigated theoretically. A one-dimensional two-temperature model with temperature-dependent material properties is considered, including dynamic optical properties and the thermal-physical properties. Rapid phase change and phase explosion models are incorporated to simulate the material ablation process. Numerical results show that under the same total laser fluence of 4 J/cm2, a dual-pulse femtosecond laser with a pulse separation time of 30–150 ps can increase the ablation depth, compared to the single pulse. The optimum pulse separation time is 85 ps. It is also demonstrated that a dual pulse with a suitable pulse separation time for different laser fluences can enhance the ablation rate by about 1.6 times.

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

    Science.gov (United States)

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

    2008-09-10

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

  10. flowPeaks: a fast unsupervised clustering for flow cytometry data via K-means and density peak finding.

    Science.gov (United States)

    Ge, Yongchao; Sealfon, Stuart C

    2012-08-01

    For flow cytometry data, there are two common approaches to the unsupervised clustering problem: one is based on the finite mixture model and the other on spatial exploration of the histograms. The former is computationally slow and has difficulty to identify clusters of irregular shapes. The latter approach cannot be applied directly to high-dimensional data as the computational time and memory become unmanageable and the estimated histogram is unreliable. An algorithm without these two problems would be very useful. In this article, we combine ideas from the finite mixture model and histogram spatial exploration. This new algorithm, which we call flowPeaks, can be applied directly to high-dimensional data and identify irregular shape clusters. The algorithm first uses K-means algorithm with a large K to partition the cell population into many small clusters. These partitioned data allow the generation of a smoothed density function using the finite mixture model. All local peaks are exhaustively searched by exploring the density function and the cells are clustered by the associated local peak. The algorithm flowPeaks is automatic, fast and reliable and robust to cluster shape and outliers. This algorithm has been applied to flow cytometry data and it has been compared with state of the art algorithms, including Misty Mountain, FLOCK, flowMeans, flowMerge and FLAME. The R package flowPeaks is available at https://github.com/yongchao/flowPeaks. yongchao.ge@mssm.edu Supplementary data are available at Bioinformatics online.

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

    Directory of Open Access Journals (Sweden)

    MUZAFFER KEKLIK

    2012-01-01

    Full Text Available

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

  12. Cytometry metadata in XML

    Science.gov (United States)

    Leif, Robert C.; Leif, Stephanie H.

    2016-04-01

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

  13. 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

  14. 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.

  15. 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...

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

    Science.gov (United States)

    Zhu, Hongying; Ozcan, Aydogan

    2015-01-01

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

  17. Intra-laser-cavity microparticle sensing with a dual-wavelength distributed-feedback laser

    NARCIS (Netherlands)

    Bernhardi, Edward H.; van der Werf, Kees O; Hollink, Anton J F; Wörhoff, Kerstin; de Ridder, René M; Subramaniam, Vinod; Pollnau, Markus

    An integrated intra-laser-cavity microparticle sensor based on a dual-wavelength distributed-feedback channel waveguide laser in ytterbium-doped amorphous aluminum oxide on a silicon substrate is demonstrated. Real-time detection and accurate size measurement of single micro-particles with diameters

  18. 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.

  19. Use of flow cytometry for the possible identification of radio-induced changes in DNA of animal cells

    International Nuclear Information System (INIS)

    Spano, M.; Leonardi, M.; Cordelli, E.

    1991-01-01

    Since DNA is the main cellular target of ionizing irradiation, methods fit for analyzing DNA alterations should be able to discern irradiated versus control cells. Flow cytometry allows the rapid measurement of DNA content of single chromosomes or cell nuclei at very high resolution on a statistically significant sample. Alterations of chromatine structure can also be analyzed by flow cytometry. Briefly, evaluation of in situ DNA resistance to denaturation can be evaluated by flow cytometric analysis of different staining pattern of single versus double strange regions of DNA. In the present work both approaches were used with the aim to recognize cells derived from an irradiated sample of breast chicken. Although flow cytometry has been demonstrated to be a useful tool to detect DNA alterations and has been widely used to detect damages on DNA induced by several physical and chemical agents, it was unable to detect clastogenic effects induced by electrons on DNA of chicken breast cells. Heavily irradiated nuclei, even if challenged by denaturating treatments that partially collapse chromatine organization, do not present differences from non irradiated samples after flow cytometric DNA content measurement. (16 refs)

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

    Science.gov (United States)

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

    2015-02-01

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

  1. Dual-channel amplification in a single-mode diode laser for multi-isotope laser cooling

    International Nuclear Information System (INIS)

    Booth, James L.; Van Dongen, Janelle; Lebel, Paul; Klappauf, Bruce G.; Madison, Kirk W.

    2007-01-01

    The output from two grating-stabilized external-cavity diode lasers were injected into a single-mode diode laser. Operating at a wavelength of 780 nm, this laser produced ∼50 mW of power with two main frequency components of the same spectral characteristics of the seed lasers. The power ratio of the amplified components was freely adjustable due to gain saturation, and amplification was observed for frequency differences of the two seed lasers in the range from 73 MHz to 6.6 GHz. This system was used to realize a dual isotope magneto-optic trap (MOT) for rubidium ( 85 Rb and 87 Rb). The resulting position and cloud size of the dual isotope MOT was the same as that of the single species MOTs to within ±10 and ±20 μm, respectively. We also characterized the additional spectral components produced by four wave mixing (FWM) in the diode laser amplifier and utilized a particular FWM sideband to realize hyperfine pumping and subsequent laser trapping of 85 Rb in the absence of a 'repump' laser dedicated to hyperfine pumping

  2. 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

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

    Science.gov (United States)

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

    1990-01-01

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

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

    Science.gov (United States)

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

    2018-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Muhammed Majeed

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

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

    Science.gov (United States)

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

    2018-01-01

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

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

    DEFF Research Database (Denmark)

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

    2014-01-01

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

  8. Reticulocyte analysis using flow cytometry.

    Science.gov (United States)

    Corberand, J X

    1996-12-01

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

  9. Vapor plume oscillation mechanisms in transient keyhole during tandem dual beam fiber laser welding

    Science.gov (United States)

    Chen, Xin; Zhang, Xiaosi; Pang, Shengyong; Hu, Renzhi; Xiao, Jianzhong

    2018-01-01

    Vapor plume oscillations are common physical phenomena that have an important influence on the welding process in dual beam laser welding. However, until now, the oscillation mechanisms of vapor plumes remain unclear. This is primarily because mesoscale vapor plume dynamics inside a millimeter-scale, invisible, and time-dependent keyhole are difficult to quantitatively observe. In this paper, based on a developed three-dimensional (3D) comprehensive model, the vapor plume evolutions in a dynamical keyhole are directly simulated in tandem dual beam, short-wavelength laser welding. Combined with the vapor plume behaviors outside the keyhole observed by high-speed imaging, the vapor plume oscillations in dynamical keyholes at different inter-beam distances are the first, to our knowledge, to be quantitatively analyzed. It is found that vapor plume oscillations outside the keyhole mainly result from vapor plume instabilities inside the keyhole. The ejection velocity at the keyhole opening and dynamical behaviors outside the keyhole of a vapor plume both violently oscillate with the same order of magnitude of high frequency (several kHz). Furthermore, the ejection speed at the keyhole opening and ejection area outside the keyhole both decrease as the beam distance increases, while the degree of vapor plume instability first decreases and then increases with increasing beam distance from 0.6 to 1.0 mm. Moreover, the oscillation mechanisms of a vapor plume inside the dynamical keyhole irradiated by dual laser beams are investigated by thoroughly analyzing the vapor plume occurrence and flow process. The vapor plume oscillations in the dynamical keyhole are found to mainly result from violent local evaporations and severe keyhole geometry variations. In short, the quantitative method and these findings can serve as a reference for further understanding of the physical mechanisms in dual beam laser welding and of processing optimizations in industrial applications.

  10. On-chip microparticle detection and sizing using a dual-wavelength waveguide laser

    NARCIS (Netherlands)

    Bernhardi, Edward; van der Werf, Kees; Hollink, Anton; Worhoff, Kerstin; de Ridder, R.M.; Subramaniam, Vinod; Pollnau, Markus

    An integrated intra-laser-cavity microparticle sensor based on a dual-phase-shift, dual-wavelength distributed-feedback channel waveguide laser in ytterbium-doped aluminium oxide is presented. Single micro-particles with diameters ranging between 1 μm and 20 μm are detected.

  11. A Novel Dual-Stage Hydrothermal Flow Reactor

    DEFF Research Database (Denmark)

    Hellstern, Henrik Christian; Becker, Jacob; Hald, Peter

    2015-01-01

    The dual-stage reactor is a novel continuous flow reactor with two reactors connected in series. It is designed for hydrothermal flow synthesis of nanocomposites, in which a single particle consists of multiple materials. The secondary material may protect the core nanoparticle from oxidation....... The dual-stage reactor combines the ability to produce advanced materials with an upscaled capacity in excess of 10 g/hour (dry mass). TiO2 was synthesized in the primary reactor and reproduced previous results. The dual-stage capability was succesfully demonstrated with a series of nanocomposites incl. Ti...

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

    Science.gov (United States)

    Poulton, Nicole J

    2016-01-01

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

  13. Comb-Resolved Dual-Comb Spectroscopy Stabilized by Free-Running Continuous-Wave Lasers

    Science.gov (United States)

    Kuse, Naoya; Ozawa, Akira; Kobayashi, Yohei

    2012-11-01

    We demonstrate dual-comb spectroscopy with relatively phase-locked two frequency combs, instead of frequency combs firmly fixed to the absolute frequency references. By stabilizing two beat frequencies between two mode-locked lasers at different wavelengths observed via free-running continuous-wave (CW) lasers, two combs are tightly phase locked to each other. The frequency noise of the CW lasers barely affects the performance of dual-comb spectroscopy because of the extremely fast common-mode noise rejection. Transform-limited comb-resolved dual-comb spectroscopy with a 6 Hz radio frequency linewidth is demonstrated by the use of Yb-fiber oscillators.

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

    Directory of Open Access Journals (Sweden)

    Greg Finak

    2014-08-01

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

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

    Science.gov (United States)

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

    2017-09-01

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

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

    Science.gov (United States)

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

    2016-07-01

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

  17. Phase-measuring laser holographic interferometer for use in high speed flows

    Science.gov (United States)

    Yanta, William J.; Spring, W. Charles, III; Gross, Kimberly Uhrich; McArthur, J. Craig

    Phase-measurement techniques have been applied to a dual-plate laser holographic interferometer (LHI). This interferometer has been used to determine the flowfield densities in a variety of two-dimensional and axisymmetric flows. In particular, LHI has been applied in three different experiments: flowfield measurements inside a two-dimensional scramjet inlet, flow over a blunt cone, and flow over an indented nose shape. Comparisons of experimentally determined densities with computational results indicate that, when phase-measurement techniques are used in conjunction with state-of-the-art image-processing instrumentation, holographic interferometry can be a diagnostic tool with high resolution, high accuracy, and rapid data retrieval.

  18. On-chip microparticle detection and sizing using a dual-wavelength waveguide laser

    NARCIS (Netherlands)

    Bernhardi, Edward H.; van der Werf, Kees O; Hollink, Anton J F; Worhoff, Kerstin; De Ridder, Rene M.; Subramaniam, Vinod; Pollnau, Markus

    2013-01-01

    An integrated intra-laser-cavity microparticle sensor based on a dual-phase-shift, dual-wavelength distributed-feedback channel waveguide laser in Al2O3:Yb3+ is presented. Real-time detection and accurate size measurement of single microparticles with diameters ranging between 1 μm and 20 μm are

  19. 78 FR 5186 - Clinical Flow Cytometry in Hematologic Malignancies; Public Workshop; Request for Comments

    Science.gov (United States)

    2013-01-24

    ... need for such products to assist clinical laboratories in performing this testing. FDA has been working... this workshop include: (1) Overview of Quality control and standardization issues associated with Clinical Flow Cytometry (FCM), including discussion of a NIST traceable standard; (2) Biological controls...

  20. Gigahertz dual-comb modelocked diode-pumped semiconductor and solid-state lasers

    Science.gov (United States)

    Link, S. M.; Mangold, M.; Golling, M.; Klenner, A.; Keller, U.

    2016-03-01

    We present a simple approach to generate simultaneously two gigahertz mode-locked pulse trains from a single gain element. A bi-refringent crystal in the laser cavity splits the one cavity beam into two cross-polarized and spatially separated beams. This polarization-duplexing is successfully demonstrated for both a semiconductor disk laser (i.e. MIXSEL) and a diode-pumped solid-state Nd:YAG laser. The beat between the two beams results in a microwave frequency comb, which represents a direct link between the terahertz optical frequencies and the electronically accessible microwave regime. This dual-output technique enables compact and cost-efficient dual-comb lasers for spectroscopy applications.

  1. 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.

  2. Intra-cavity decomposition of a dual-directional laser beam

    CSIR Research Space (South Africa)

    Naidoo, Darryl

    2011-01-01

    Full Text Available A method of decomposing a dual-directional laser beam into a forward propagating field and a backward propagating field for an apertured plano-concave cavity is presented. An intra-cavity aperture is a simple method of laser beam shaping as higher...

  3. Induction studies with Escherichia coli expressing recombinant interleukin-13 using multi-parameter flow cytometry

    DEFF Research Database (Denmark)

    Shitu, J. O.; Woodley, John; Wnek, R.

    2009-01-01

    The expression of interleukin-13 (IL13) following induction with IPTG in Escherichia coli results in metabolic changes as indicated by multi-parameter flow cytometry and traditional methods of fermentation profiling (O-2 uptake rate, CO2 evolution rate and optical density measurements). Induction...

  4. Novel Laser Ignition Technique Using Dual-Pulse Pre-Ionization

    Science.gov (United States)

    Dumitrache, Ciprian

    Recent advances in the development of compact high power laser sources and fiber optic delivery of giant pulses have generated a renewed interest in laser ignition. The non-intrusive nature of laser ignition gives it a set of unique characteristics over the well-established capacitive discharge devices (or spark plugs) that are currently used as ignition sources in engines. Overall, the use of laser ignition has been shown to have a positive impact on engine operation leading to a reduction in NOx emission, fuel saving and an increased operational envelope of current engines. Conventionally, laser ignition is achieved by tightly focusing a high-power q-switched laser pulse until the optical intensity at the focus is high enough to breakdown the gas molecules. This leads to the formation of a spark that serves as the ignition source in engines. However, there are certain disadvantages associated with this ignition method. This ionization approach is energetically inefficient as the medium is transparent to the laser radiation until the laser intensity is high enough to cause gas breakdown. As a consequence, very high energies are required for ignition (about an order of magnitude higher energy than capacitive plugs at stoichiometric conditions). Additionally, the fluid flow induced during the plasma recombination generates high vorticity leading to high rates of flame stretching. In this work, we are addressing some of the aforementioned disadvantages of laser ignition by developing a novel approach based on a dual-pulse pre-ionization scheme. The new technique works by decoupling the effect of the two ionization mechanisms governing plasma formation: multiphoton ionization (MPI) and electron avalanche ionization (EAI). An UV nanosecond pulse (lambda = 266 nm) is used to generate initial ionization through MPI. This is followed by an overlapped NIR nanosecond pulse (lambda = 1064 nm) that adds energy into the pre-ionized mixture into a controlled manner until the

  5. 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.  ...

  6. Utility of peripheral blood immunophenotyping by flow cytometry in the diagnosis of pediatric acute leukemia.

    Science.gov (United States)

    Metrock, Laura K; Summers, Ryan J; Park, Sunita; Gillespie, Scott; Castellino, Sharon; Lew, Glen; Keller, Frank G

    2017-10-01

    Childhood acute leukemia is traditionally diagnosed from a bone marrow aspirate (BMA). New-onset acute leukemia patients do not always have visible circulating blasts in the peripheral blood (PB) at diagnosis. While the role of bone marrow flow cytometry for the diagnosis of acute leukemia is well established, the utility of PB flow cytometry (PBFC) is unknown. We performed a single-institution retrospective analysis to compare PBFC versus BMA in establishing or excluding a diagnosis of childhood acute leukemia. We retrospectively identified 485 PBFC samples with concurrent BMA from 2008 to 2013. Results of four-color flow cytometry for immunophenotypic characterization of leukemic versus nonclonal disease were characterized. Sensitivity and specificity were calculated among patients without a known diagnosis or prior therapy. Among 485 samples eligible for analysis, 120 had negative PBFC and BMA, 359 had positive PBFC and BMA, 3 had negative PBFC and positive BMA, and 3 had positive PBFC and negative BMA. There were small but significant differences in sensitivity (100 vs. 93.8%; P = 0.002) and positive predictive value (100 vs. 93.8%; P = 0.002) favoring BMA over PBFC among those demonstrating absence of circulating morphologic blasts. PBFC has high sensitivity and specificity for the diagnosis of childhood acute leukemia. The predictive value of PBFC remains high for patients without visible circulating blasts and may enhance the diagnostic process for determining the indications for marrow testing. © 2017 Wiley Periodicals, Inc.

  7. 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.

  8. Detection of Intracellular Factor VIII Protein in Peripheral Blood Mononuclear Cells by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Gouri Shankar Pandey

    2013-01-01

    Full Text Available Flow cytometry is widely used in cancer research for diagnosis, detection of minimal residual disease, as well as immune monitoring and profiling following immunotherapy. Detection of specific host proteins for diagnosis predominantly uses quantitative PCR and western blotting assays. In this study, we optimized a flow cytometry-based detection assay for Factor VIII protein in peripheral blood mononuclear cells (PBMCs. An indirect intracellular staining (ICS method was standardized using monoclonal antibodies to different domains of human Factor VIII protein. The FVIII protein expression level was estimated by calculating the mean and median fluorescence intensities (MFI values for each monoclonal antibody. ICS staining of transiently transfected cell lines supported the method's specificity. Intracellular FVIII protein expression was also detected by the monoclonal antibodies used in the study in PBMCs of five blood donors. In summary, our data suggest that intracellular FVIII detection in PBMCs of hemophilia A patients can be a rapid and reliable method to detect intracellular FVIII levels.

  9. 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.

  10. CytometryML: a markup language for analytical cytology

    Science.gov (United States)

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

    2003-06-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

  12. Antimicrobial Activity of Rhoeo discolor Phenolic Rich Extracts Determined by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Rebeca García-Varela

    2015-10-01

    Full Text Available Traditional medicine has led to the discovery of important active substances used in several health-related areas. Phytochemicals in Rhoeo discolor extracts have proven to have important antimicrobial activity. In the present study, our group determined the antimicrobial effects of extracts of Rhoeo discolor, a plant commonly used in Mexico for both medicinal and ornamental purposes. We evaluated the in vitro activity of phenolic rich extracts against specifically chosen microorganisms of human health importance by measuring their susceptibility via agar-disc diffusion assay and flow cytometry: Gram-positive Listeria innocua and Streptococcus mutans, Gram-negative Escherichia coli and Pseudomonas aeruginosa, and lastly a fungal pathogen Candida albicans. Ten different extracts were tested in eight different doses on all the microorganisms. Analytical data revealed a high content of phenolic compounds. Both agar-disc diffusion assay and flow cytometry results demonstrated that Pseudomonas aeruginosa was the least affected by extract exposure. However, low doses of these extracts (predominantly polar, in a range from 1 to 4 μg/mL, did produce a statistically significant bacteriostatic and bactericidal effect on the rest of the microorganisms. These results suggest the addition of certain natural extracts from Rhoeo discolor could act as antibacterial and antimycotic drugs or additives for foods and cosmetics.

  13. CytometryML and other data formats

    Science.gov (United States)

    Leif, Robert C.

    2006-02-01

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

  14. Evaluation of CD307a expression patterns during normal B-cell maturation and in B-cell malignancies by flow cytometry.

    Science.gov (United States)

    Auat, Mariangeles; Cardoso, Chandra Chiappin; Santos-Pirath, Iris Mattos; Rudolf-Oliveira, Renata Cristina Messores; Matiollo, Camila; Lange, Bárbara Gil; da Silva, Jessica Pires; Dametto, Gisele Cristina; Pirolli, Mayara Marin; Colombo, Maria Daniela Holthausen Perico; Santos-Silva, Maria Claudia

    2018-02-24

    Flow cytometric immunophenotyping is deemed a fundamental tool for the diagnosis of B-cell neoplasms. Currently, the investigation of novel immunophenotypic markers has gained importance, as they can assist in the precise subclassification of B-cell malignancies by flow cytometry. Therefore, the purpose of the present study was to evaluate the expression of CD307a during normal B-cell maturation and in B-cell malignancies as well as to investigate its potential role in the differential diagnosis of these entities. CD307a expression was assessed by flow cytometry in normal precursor and mature B cells and in 115 samples collected from patients diagnosed with precursor and mature B-cell neoplasms. CD307a expression was compared between neoplastic and normal B cells. B-acute lymphoblastic leukemia cases exhibited minimal expression of CD307a, displaying a similar expression pattern to that of normal B-cell precursors. Mantle cell lymphoma (MCL) cases showed the lowest levels of CD307a among mature B-cell neoplasms. CD307a expression was statistically lower in MCL cases than in chronic B lymphocytic leukemia (CLL) and marginal zone lymphoma (MZL) cases. No statistical differences were observed between CD307a expression in neoplastic and normal plasma cells. These results indicate that the assessment of CD307a expression by flow cytometry could be helpful to distinguish CLL from MCL, and the latter from MZL. Although these results are not entirely conclusive, they provide a basis for further studies in a larger cohort of patients. © 2018 International Clinical Cytometry Society. © 2018 International Clinical Cytometry Society.

  15. Simulation of flow in dual-scale porous media

    Science.gov (United States)

    Tan, Hua

    Liquid composite molding (LCM) is one of the most effective processes for manufacturing near net-shaped parts from fiber-reinforced polymer composites. The quality of LCM products and the efficiency of the process depend strongly on the wetting of fiber preforms during the mold-filling stage of LCM. Mold-filling simulation is a very effective approach to optimize the LCM process and mold design. Recent studies have shown that the flow modeling for the single-scale fiber preforms (made from random mats) has difficulties in accurately predicting the wetting in the dual-scale fiber preforms (made from woven and stitched fabrics); the latter are characterized by the presence of unsaturated flow created due to two distinct length-scales of pores (i.e., large pores outside the tows and small pores inside the tows) in the same media. In this study, we first develop a method to evaluate the accuracy of the permeability-measuring devices for LCM, and conduct a series of 1-D mold-filling experiments for different dual-scale fabrics. The volume averaging method is then applied to derive the averaged governing equations for modeling the macroscopic flow through the dual-scale fabrics. The two sets of governing equations are coupled with each other through the sink terms representing the absorptions of mass, energy, and species (degree of resin cure) from the global flow by the local fiber tows. The finite element method (FEM) coupled with the control volume method, also known as the finite element/control volume (FE/CV) method, is employed to solve the governing equations and track the moving boundary signifying the moving liquid-front. The numerical computations are conducted with the help of an in-house developed computer program called PORE-FLOW(c). We develop the flux-corrected transport (FCT) based FEM to stabilize the convection-dominated energy and species equations. A fast methodology is proposed to simulate the dual-scale flow under isothermal conditions, where flow

  16. Dual comb generation from a mode-locked fiber laser with orthogonally polarized interlaced pulses.

    Science.gov (United States)

    Akosman, Ahmet E; Sander, Michelle Y

    2017-08-07

    Ultra-high precision dual-comb spectroscopy traditionally requires two mode-locked, fully stabilized lasers with complex feedback electronics. We present a novel mode-locked operation regime in a thulium-holmium co-doped fiber laser, a frequency-halved state with orthogonally polarized interlaced pulses, for dual comb generation from a single source. In a linear fiber laser cavity, an ultrafast pulse train composed of co-generated, equal intensity and orthogonally polarized consecutive pulses at half of the fundamental repetition rate is demonstrated based on vector solitons. Upon optical interference of the orthogonally polarized pulse trains, two stable microwave RF beat combs are formed, effectively down-converting the optical properties into the microwave regime. These co-generated, dual polarization interlaced pulse trains, from one all-fiber laser configuration with common mode suppression, thus provide an attractive compact source for dual-comb spectroscopy, optical metrology and polarization entanglement measurements.

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

    Science.gov (United States)

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

    2018-05-01

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

  18. Radial flow gas dynamic laser

    International Nuclear Information System (INIS)

    Damm, F.C.

    1975-01-01

    The unique gas dynamic laser provides outward radial supersonic flow from a toroidal shaped stacked array of a plurality of nozzles, through a diffuser having ring shaped and/or linear shaped vanes, and through a cavity which is cylindrical and concentric with the stacked array, with the resultant laser beam passing through the housing parallel to the central axis of the diffuser which is coincident with the axis of the gas dynamic laser. Therefore, greater beam extraction flexibility is attainable, because of fewer flow shock disturbances, as compared to the conventional unidirectional flow gas dynamic laser in which unidirectional supersonic flow sweeps through a rectangular cavity and is exhausted through a two-dimensional diffuser. (auth)

  19. Assessment of ploidy stability of the somatic embryogenesis process in Quercus suber L. using flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Loureiro, J.; Pinto, G.; Lopes, T.; Doležel, Jaroslav; Santos, C.

    2005-01-01

    Roč. 221, - (2005), s. 815-822 ISSN 0032-0935 Institutional research plan: CEZ:AV0Z50380511 Keywords : Flow cytometry * ploidy stability * nuclear DNA content Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.108, year: 2005

  20. [Clinical usefulness of urine-formed elements' information obtained from bacteria detection by flow cytometry method that uses nucleic acid staining].

    Science.gov (United States)

    Nakagawa, Hiroko; Yuno, Tomoji; Itho, Kiichi

    2009-03-01

    Recently, specific detection method for Bacteria, by flow cytometry method using nucleic acid staining, was developed as a function of automated urine formed elements analyzer for routine urine testing. Here, we performed a basic study on this bacteria analysis method. In addition, we also have a comparison among urine sediment analysis, urine Gram staining and urine quantitative cultivation, the conventional methods performed up to now. As a result, the bacteria analysis with flow cytometry method that uses nucleic acid staining was excellent in reproducibility, and higher sensitivity compared with microscopic urinary sediment analysis. Based on the ROC curve analysis, which settled urine culture method as standard, cut-off level of 120/microL was defined and its sensitivity = 85.7%, specificity = 88.2%. In the analysis of scattergram, accompanied with urine culture method, among 90% of rod positive samples, 80% of dots were appeared in the area of 30 degrees from axis X. In addition, one case even indicated that analysis of bacteria by flow cytometry and scattergram of time series analysis might be helpful to trace the progress of causative bacteria therefore the information supposed to be clinically significant. Reporting bacteria information with nucleic acid staining flow cytometry method is expected to contribute to a rapid diagnostics and treatment of urinary tract infections. Besides, the contribution to screening examination of microbiology and clinical chemistry, will deliver a more efficient solution to urine analysis.

  1. Laser scanning cytometry (LCS) allows detailed analysis of the cell cycle in PI stained human fibroblasts (TIG-7).

    Science.gov (United States)

    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.

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

    Science.gov (United States)

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

    2014-06-06

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

  3. Dual-signal heterodyne lock-in amplification with lasers

    NARCIS (Netherlands)

    Witteman, W.J.

    2006-01-01

    High-sensitivity heterodyne detection with lasers applied to radar and satellite communication is seriously hampered by the large electronic bandwidth due to Doppler shift and frequency instability. These drawbacks can be circumvented by dual-signal heterodyne detection. The system consists of

  4. Advantages of flow cytometry immunophenotyping for the diagnosis of central nervous system non-Hodgkin's lymphoma in AIDS patients.

    Science.gov (United States)

    Subirá, D; Górgolas, M; Castañón, S; Serrano, C; Román, A; Rivas, F; Tomás, J F

    2005-01-01

    Neurological disorders are common in HIV-infected patients. Central nervous system (CNS) lymphoma should always be considered because it is an important cause of morbidity and mortality. To investigate the clinical utility of flow cytometry immunophenotyping (FCI) in diagnosing or discarding leptomeningeal involvement in HIV-infected patients and to compare its sensitivity with that of conventional cytological methods. Fifty-six cerebrospinal fluid (CSF) samples from 29 HIV-infected patients were independently evaluated by flow cytometry and cytology. The description of an aberrant immunophenotype was the criterion used to define the malignant nature of any CSF cell population. FCI and cytology gave concordant results for 48 of the 56 CSF samples studied: 37 were negative for malignancy and 11 had evidence of CNS lymphoma. Discordant results were obtained for eight CSF samples, and the accuracy of the FCI findings could be demonstrated for four CSF samples described as positive for malignancy according to the FCI criteria. A high level of agreement was found between the results obtained using the two methods, but FCI gave at least 25% higher sensitivity than conventional cytomorphological methods for the detection of malignant cells. This advantage suggests that, in case of negative flow cytometry results, disorders other than non-Hodgkin's lymphoma should be strongly considered.

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

    Science.gov (United States)

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

    2016-08-01

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

  6. Remarkable coexistence of multiple cytotypes of the Gymnadenia conopsea aggregate (the fragrant orchid): evidence from flow cytometry

    Czech Academy of Sciences Publication Activity Database

    Trávníček, Pavel; Kubátová, B.; Čurn, V.; Rauchová, Jana; Krajníková, E.; Jersáková, J.; Suda, Jan

    2011-01-01

    Roč. 107, č. 1 (2011), s. 77-87 ISSN 0305-7364 Institutional research plan: CEZ:AV0Z6005908 Keywords : coexistence * contact zone * flow cytometry Subject RIV: EF - Botanics Impact factor: 4.030, year: 2011

  7. Correlation between flow cytometry and histologic findings: ten year experience in the investigation of lymphoproliferative diseases

    Directory of Open Access Journals (Sweden)

    Alanna Mara Pinheiro Sobreira Bezerra

    2011-06-01

    Full Text Available Objective: To demonstrate the advantages of correlatingflow cytometry immunophenotyping with the pathology/immunohistochemistry of lymph nodes or nodules in the diagnosisof lymphoproliferative diseases. Methods: A retrospective studywas carried out of 157 biopsy or fine-needle aspiration lymph nodes/nodule specimens taken from 142 patients, from 1999 and 2009.The specimens were simultaneously studied with flow cytometryand pathology at Hospital Israelita Albert Einstein. The specimenswere prepared in hematoxylin/eosin, Giemsa, or monoclonal antibodystained slides for detecting specific antibodies for the purposesof pathology/immunohistochemical analysis. The samples werehemolyzed and marked with different monoclonal antibody panels fordifferent antigens in flow cytometry immunophenotyping. Results:The diagnostic results of pathology/immunohistochemical studiesand flow cytometry immunophenotyping agreed in 115 patients(81%, corresponding to 127 specimens, as follows according tothe pathologic diagnosis: 63 patients with non-Hodgkin’s B-celllymphoma; 26 patients with reactive lymphoid hyperplasia; 5 patientswith non-Hodgkin’s T-cell lymphoma; 4 patients with atypical lymphoidproliferation; 5 patients with a chronic granulomatous inflammatoryprocess; 5 patients with a non-hematologic diagnosis; 2 patientswith granulocytic sarcoma; 2 patients with thymoma; 1 patientwith byphenotypic leukemia; 1 patient with kappa plasmocytoma;1 patient with Hodgkin’s lymphoma. Subtypes of lymphomas couldbe classified by associating the two techniques: 19 patients withfollicular lymphoma; 15 patients with diffuse large B-cell lymphoma; 7patients with small lymphocytic B-cell lymphoma/chronic lymphocyticleukemia; 3 patients with mantle cell lymphoma; 1 patient withBurkitt’s lymphoma; 1 patient with MALT type lymphoma; 1 patientwith post-transplant lymphoproliferative disease; 2 patients with highgrade non-Hodgkin’s B-cell lymphoma; 1 patient with low grade

  8. Dual-comb spectroscopy of water vapor with a free-running semiconductor disk laser.

    Science.gov (United States)

    Link, S M; Maas, D J H C; Waldburger, D; Keller, U

    2017-06-16

    Dual-comb spectroscopy offers the potential for high accuracy combined with fast data acquisition. Applications are often limited, however, by the complexity of optical comb systems. Here we present dual-comb spectroscopy of water vapor using a substantially simplified single-laser system. Very good spectroscopy measurements with fast sampling rates are achieved with a free-running dual-comb mode-locked semiconductor disk laser. The absolute stability of the optical comb modes is characterized both for free-running operation and with simple microwave stabilization. This approach drastically reduces the complexity for dual-comb spectroscopy. Band-gap engineering to tune the center wavelength from the ultraviolet to the mid-infrared could optimize frequency combs for specific gas targets, further enabling dual-comb spectroscopy for a wider range of industrial applications. Copyright © 2017, American Association for the Advancement of Science.

  9. Highly reproducible LENR experiments using dual laser stimulation

    International Nuclear Information System (INIS)

    Letts, Dennis

    2015-01-01

    The present article reports a summary of results relating to 170 on-protocol tests performed on five deuterated palladium bulk cathodes stimulated by dual lasers at 8, 15 and 20 THz over the period March 2007 to May 2008. These frequencies were not measured but assumed to be effective based on a calculated difference beat frequency. Excess power was observed in 161 tests, giving a success rate of approximately 95%. The cathode fabrication, loading and laser application protocols are discussed. (author)

  10. Resonant laser techniques for combustion and flow diagnostics

    Energy Technology Data Exchange (ETDEWEB)

    Fritzon, Rolf

    1998-05-01

    This thesis presents results from two areas of research. Firstly, the resonant coherent laser techniques polarization spectroscopy (PS), degenerate four-wave mixing (DFWM) and stimulated emission (SE) have been developed in the general field of combustion diagnostics. Secondly, laser induced fluorescence (LIF) has been developed and applied for the visualization of mixture fractions in turbulent non reacting flows. PS was developed for instantaneous two-dimensional imaging of minor species in flames, the technique being demonstrated on OH and NO. Various aspects of imaging and of detection in general were investigated. Two-photon induced PS was demonstrated for the detection of NH{sub 3}, CO and N{sub 2} molecules. LIF was monitored simultaneously to allow a quantitative comparison between the techniques. Furthermore, PS and DFWM were developed for instantaneous two-dimensional OH temperature imaging. Through a novel experimental approach based on the use of a dual-wavelength dye laser and a diffraction grating the temperature imaging measurements were performed using only one laser and one CCD camera. A comparison between the two techniques was made. SE was through a crossed-beam arrangement developed for spatially resolved detection of flame species. Two-dimensional LIF was developed and applied for measuring mixture fractions in the shear layer between two co-flowing turbulent gaseous jets. The technique was further applied in a study of the mixing of a turbulent water jet impinging orthogonally onto a flat surface. Average concentration fields in the center-plane of the jet was compared with results from large eddy simulations and with data from the literature 221 refs, 48 figs, 5 tabs

  11. Analysis of Cellular DNA Content by Flow Cytometry.

    Science.gov (United States)

    Darzynkiewicz, Zbigniew; Huang, Xuan; Zhao, Hong

    2017-11-01

    Cellular DNA content can be measured by flow cytometry with the aim of : (1) revealing cell distribution within the major phases of the cell cycle, (2) estimating frequency of apoptotic cells with fractional DNA content, and/or (3) disclosing DNA ploidy of the measured cell population. In this unit, simple and universally applicable methods for staining fixed cells are presented, as are methods that utilize detergents and/or proteolytic treatment to permeabilize cells and make DNA accessible to fluorochrome. Additionally, supravital cell staining with Hoechst 33342, which is primarily used for sorting live cells based on DNA-content differences for their subsequent culturing, is described. Also presented are methods for staining cell nuclei isolated from paraffin-embedded tissues. Available algorithms are listed for deconvolution of DNA-content-frequency histograms to estimate percentage of cells in major phases of the cell cycle and frequency of apoptotic cells with fractional DNA content. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley and Sons, Inc.

  12. Laser manipulation of atomic and molecular flows

    Science.gov (United States)

    Lilly, Taylor C.

    The continuing advance of laser technology enables a range of broadly applicable, laser-based flow manipulation techniques. The characteristics of these laser-based flow manipulations suggest that they may augment, or be superior to, such traditional electro-mechanical methods as ionic flow control, shock tubes, and small scale wind tunnels. In this study, methodology was developed for investigating laser flow manipulation techniques, and testing their feasibility for a number of aerospace, basic physics, and micro technology applications. Theories for laser-atom and laser-molecule interactions have been under development since the advent of laser technology. The theories have yet to be adequately integrated into kinetic flow solvers. Realizing this integration would greatly enhance the scaling of laser-species interactions beyond the realm of ultra-cold atomic physics. This goal was realized in the present study. A representative numerical investigation, of laser-based neutral atomic and molecular flow manipulations, was conducted using near-resonant and non-resonant laser fields. To simulate the laser interactions over a range of laser and flow conditions, the following tools were employed: a custom collisionless gas particle trajectory code and a specifically modified version of the Direct Simulation Monte Carlo statistical kinetic solver known as SMILE. In addition to the numerical investigations, a validating experiment was conducted. The experimental results showed good agreement with the numerical simulations when experimental parameters, such as finite laser line width, were taken into account. Several areas of interest were addressed: laser induced neutral flow steering, collimation, direct flow acceleration, and neutral gas heating. Near-resonant continuous wave laser, and non-resonant pulsed laser, interactions with cesium and nitrogen were simulated. These simulations showed trends and some limitations associated with these interactions, used for flow

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

    Science.gov (United States)

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

    2017-10-01

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

  14. A fast spectrum dual path flow cermet reactor

    International Nuclear Information System (INIS)

    Anghaie, S.; Feller, G.J.; Peery, S.D.; Parsley, R.C.

    1993-01-01

    A cermet fueled, dual path fast reactor for space nuclear propulsion applications is conceptually designed. The reactor utilizes an outer annulus core and an inner cylindrical core with radial and axial reflector. The dual path flow minimizes the impact of power peaking near the radial reflector. Basic neutronics and core design aspects of the reactor are discussed. The dual path reactor is integrated into a 25000 lbf thrust nuclear rocket

  15. flowAI: automatic and interactive anomaly discerning tools for flow cytometry data.

    Science.gov (United States)

    Monaco, Gianni; Chen, Hao; Poidinger, Michael; Chen, Jinmiao; de Magalhães, João Pedro; Larbi, Anis

    2016-08-15

    Flow cytometry (FCM) is widely used in both clinical and basic research to characterize cell phenotypes and functions. The latest FCM instruments analyze up to 20 markers of individual cells, producing high-dimensional data. This requires the use of the latest clustering and dimensionality reduction techniques to automatically segregate cell sub-populations in an unbiased manner. However, automated analyses may lead to false discoveries due to inter-sample differences in quality and properties. We present an R package, flowAI, containing two methods to clean FCM files from unwanted events: (i) an automatic method that adopts algorithms for the detection of anomalies and (ii) an interactive method with a graphical user interface implemented into an R shiny application. The general approach behind the two methods consists of three key steps to check and remove suspected anomalies that derive from (i) abrupt changes in the flow rate, (ii) instability of signal acquisition and (iii) outliers in the lower limit and margin events in the upper limit of the dynamic range. For each file analyzed our software generates a summary of the quality assessment from the aforementioned steps. The software presented is an intuitive solution seeking to improve the results not only of manual but also and in particular of automatic analysis on FCM data. R source code available through Bioconductor: http://bioconductor.org/packages/flowAI/ CONTACTS: mongianni1@gmail.com or Anis_Larbi@immunol.a-star.edu.sg Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  16. Dual-wavelength laser transmission photoscanner for breast cancer detection

    International Nuclear Information System (INIS)

    Kaneko, M.; He, P.; Tanaka, H.; Takahashi, M.; Takai, M.; Baba, K.; Yamashita, Y.; Ohta, K.

    1989-01-01

    This paper reports on the prototype of a laser transmission photoscanner (LTPS) constructed and used for the detection of breast cancer and compared with x-ray mammography. LTPS has been improved to enable spectroanalysis and application in breast cancer screening. The new type is introduced. In order to obtain higher sensitivity, the output of lasers was increased in intensity. The signal integration time was increased 10-fold, and the width of the detector area was doubled. The gated operation of the detector enables the good throughput. Simultaneous scanning in the dual wavelengths of 630 and 830 nm makes it possible to differentiate hemoglobin (Hb) and oxyhemoglobin (HbO 2 ) in spectroanalysis by means of Lambert--Beer's law. Clinical application of dual-wavelength LTPS shows good correlation with pathology

  17. Flow cytometry as a rapid test for detection of penicillin resistance directly in bacterial cells in Enterococcus faecalis and Staphylococcus aureus.

    Science.gov (United States)

    Jarzembowski, T; Wiśniewska, K; Józwik, A; Bryl, E; Witkowski, J

    2008-08-01

    We studied the usefulness of flow cytometry for detection of penicillin resistance in E. faecalis and S. aureus by direct binding of commercially available fluorescent penicillin, Bocillin FL, to cells obtained from culture. There were significantly lower percentages of fluorescent cells and median and mean fluorescence values per particle in penicillin-resistant than in penicillin-sensitive strains of both species observed. The method allows rapid detection of penicillin resistance in S. aureus and E. faecalis. The results encourage further investigations on the detection of antibiotic resistance in bacteria using flow cytometry.

  18. Coherent laser radar with dual-frequency Doppler estimation and interferometric range detection

    NARCIS (Netherlands)

    Onori, D.; Scotti, F.; Laghezza, F.; Scaffardi, M.; Bogoni, A.

    2016-01-01

    The concept of a coherent interferometric dual frequency laser radar, that measures both the target range and velocity, is presented and experimentally demonstrated. The innovative architecture combines the dual frequency lidar concept, allowing a precise and robust Doppler estimation, with the

  19. The cut-off point of dual energy X-ray and laser of calcaneus osteoporosis diagnosis in postmenopausal women

    International Nuclear Information System (INIS)

    Salimzadeh, A.; Forough, B.; Olia, B.; Alishiri, G. H.; Ghasemzadeh, A.

    2005-01-01

    Dual X-Ray Absorptiometry is a method which can extensively be used for bone mineral densitometry . Another more recent method is dual energy X-ray and laser, which associate with dual X ray absorptiometry, assisted by laser measure heel thickness. In this study the cut off points for dual energy X-ray and laser of calcaneus in the diagnosis of osteoporosis in different bone regions in postmenopausal women had been determined. Materials and Methods: In 268 postmenopausal women, BMD of the spinal and femoral regions was measured by DM, and the value for the calcaneous was measured by dual energy X-ray and laser. The agreement of the two methods in the diagnosis of osteoporosis and optimal cut-off point for dual energy X-ray and laser in defining osteoporosis was obtained. What obtained was the agreement of the two methods in the diagnosis of osteoporosis, as well as the optimal cut-off point for dual energy X-ray and laser in defining osteoporosis. Results: Dual X-Ray Absorptiometry showed osteoporosis in 40.7% of cases with 35.2% in L2-L4, 16.2% in the femoral neck, and 11.7% for the femoral total region. The dual energy X-ray and laser found osteoporosis, considering -2.5 SD as a threshold, in 26.1% of cases. Agreement of the two methods in the diagnosis of osteoporosis (Kappa score) was 0.443 for the lumbar region, 0.464 for the neck, and, 0.421 for total femur regions (all P values were significant). Using Receiver Operating Characteristic curves, it was found that a T-score of -2.1, -2.6 and -2.4 as the optimal cut-off point of dual energy X-ray and laser in the diagnosis of osteoporosis in the lumbar spine, the neck and total region of femur, respectively. Conclusion: The results of this study showed a moderate agreement between the two methods in the diagnosis of osteoporosis. It seems that the dual energy X-ray and laser cannot be used as a substitute for the DM method, but it can be used as a screening method to find (to diagnose) osteoporosis

  20. Polyploidy in the Olive Complex (Olea europaea): Evidence from Flow Cytometry and Nuclear Microsatellite Analyses

    Science.gov (United States)

    Besnard, G.; Garcia-Verdugo, C.; Rubio De Casas, R.; Treier, U. A.; Galland, N.; Vargas, P.

    2008-01-01

    Background Phylogenetic and phylogeographic investigations have been previously performed to study the evolution of the olive tree complex (Olea europaea). A particularly high genomic diversity has been found in north-west Africa. However, to date no exhaustive study has been addressed to infer putative polyploidization events and their evolutionary significance in the diversification of the olive tree and its relatives. Methods Representatives of the six olive subspecies were investigated using (a) flow cytometry to estimate genome content, and (b) six highly variable nuclear microsatellites to assess the presence of multiple alleles at co-dominant loci. In addition, nine individuals from a controlled cross between two individuals of O. europaea subsp. maroccana were characterized with microsatellites to check for chromosome inheritance. Key Results Based on flow cytometry and genetic analyses, strong evidence for polyploidy was obtained in subspp. cerasiformis (tetraploid) and maroccana (hexaploid), whereas the other subspecies appeared to be diploids. Agreement between flow cytometry and genetic analyses gives an alternative approach to chromosome counting to determine ploidy level of trees. Lastly, abnormalities in chromosomes inheritance leading to aneuploid formation were revealed using microsatellite analyses in the offspring from the controlled cross in subsp. maroccana. Conclusions This study constitutes the first report for multiple polyploidy in olive tree relatives. Formation of tetraploids and hexaploids may have played a major role in the diversification of the olive complex in north-west Africa. The fact that polyploidy is found in narrow endemic subspecies from Madeira (subsp. cerasiformis) and the Agadir Mountains (subsp. maroccana) suggests that polyploidization has been favoured to overcome inbreeding depression. Lastly, based on previous phylogenetic analyses, we hypothesize that subsp. cerasiformis resulted from hybridization between ancestors

  1. Tracking by flow cytometry antigen-specific follicular helper T cells in wild-type animals after protein vaccination.

    Science.gov (United States)

    Chakarov, Svetoslav; Fazilleau, Nicolas

    2015-01-01

    Flow cytometry is a valuable technology used in immunology to characterize and enumerate the different cell subpopulations specific for a nonself-antigen in the context of an ongoing immune response. Among them, follicular helper T cells are the cognate regulators of B cells in secondary lymphoid tissues. Thus, tracking them is of high interest especially in the context of protein vaccination. For this purpose, transgenic antigen-receptor mouse models have been largely used. It is now clear that transgenic models are not always the best means to study the dynamics of the immune response since they can modify the response. In this chapter, we describe how to track endogenous antigen-specific follicular helper T cells by flow cytometry after protein vaccination in nonmodified wild-type animals, which ultimately provides a comprehensive way to enumerate, characterize, and isolate these particular cells in vivo.

  2. Identification and Characterization of Plasma Cells in Normal Human Bone Marrow by High-Resolution Flow Cytometry

    NARCIS (Netherlands)

    Terstappen, Leonardus Wendelinus Mathias Marie; Johnsen, Steen; Segers-Nolten, Gezina M.J.; Loken, Michael R.

    1990-01-01

    The low frequency of plasma cells and the lack of specific cell surface markers has been a major obstacle for a detailed characterization of plasma cells in normal human bone marrow. Multiparameter flow cytometry enabled the identification of plasma cells in normal bone marrow aspirates. The plasma

  3. Mutant spectra of irradiated CHO AL cells determined with multiple markers analyzed by flow cytometry

    International Nuclear Information System (INIS)

    Ross, Carley D.; French, C. Tenley; Keysar, Stephen B.; Fox, Michael H.

    2007-01-01

    We have previously developed a sensitive and rapid mammalian cell mutation assay which is based on a Chinese hamster ovary cell line that stably incorporates human chromosome 11 (CHO A L ) and uses flow cytometry to measure mutations in CD59. We now show that multiparameter flow cytometry may be used to simultaneously analyze irradiated CHO A L cells for mutations in five CD genes along chromosome 11 (CD59, CD44, CD90, CD98, CD151) and also a GPI-anchor gene. Using this approach, 19 different mutant clones derived from individual sorted mutant cells were analyzed to determine the mutant spectrum induced by ionizing radiation. All clones analyzed were negative for CD59 expression and PCR confirmed that at least CD59 exon 4 was also absent. As expected, ionizing radiation frequently caused large deletions along chromosome 11. This technology can readily be used to rapidly analyze the mutant yield as well as the spectrum of mutations caused by a variety of genotoxic agents and provide greater insight into the mechanisms of mutagenesis

  4. 2006 Bethesda International Consensus recommendations on the immunophenotypic analysis of hematolymphoid neoplasia by flow cytometry: optimal reagents and reporting for the flow cytometric diagnosis of hematopoietic neoplasia.

    Science.gov (United States)

    Wood, Brent L; Arroz, Maria; Barnett, David; DiGiuseppe, Joseph; Greig, Bruce; Kussick, Steven J; Oldaker, Teri; Shenkin, Mark; Stone, Elizabeth; Wallace, Paul

    2007-01-01

    Immunophenotyping by flow cytometry has become standard practice in the evaluation and monitoring of patients with hematopoietic neoplasia. However, despite its widespread use, considerable variability continues to exist in the reagents used for evaluation and the format in which results are reported. As part of the 2006 Bethesda Consensus conference, a committee was formed to attempt to define a consensus set of reagents suitable for general use in the diagnosis and monitoring of hematopoietic neoplasms. The committee included laboratory professionals from private, public, and university hospitals as well as large reference laboratories that routinely operate clinical flow cytometry laboratories with an emphasis on lymphoma and leukemia immunophenotyping. A survey of participants successfully identified the cell lineage(s) to be evaluated for each of a variety of specific medical indications and defined a set of consensus reagents suitable for the initial evaluation of each cell lineage. Elements to be included in the reporting of clinical flow cytometric results for leukemia and lymphoma evaluation were also refined and are comprehensively listed. The 2006 Bethesda Consensus conference represents the first successful attempt to define a set of consensus reagents suitable for the initial evaluation of hematopoietic neoplasia. Copyright 2007 Clinical Cytometry Society.

  5. Rapid Flow Cytometry-Based Test for the Diagnosis of Lipopolysaccharide Responsive Beige-Like Anchor (LRBA Deficiency

    Directory of Open Access Journals (Sweden)

    Laura Gámez-Díaz

    2018-04-01

    Full Text Available The diagnosis of lipopolysaccharide-responsive beige-like-anchor-protein (LRBA deficiency currently relies on gene sequencing approaches that do not support a timely diagnosis and clinical management. We developed a rapid and sensitive test for clinical implementation based on the detection of LRBA protein by flow cytometry in peripheral blood cells after stimulation. LRBA protein was assessed in a prospective cohort of 54 healthy donors and 57 patients suspected of LRBA deficiency. Receiver operating characteristics analysis suggested an LRBA:MFI ratio cutoff point of 2.6 to identify LRBA-deficient patients by FACS with 94% sensitivity and 80% specificity and to discriminate them from patients with a similar clinical picture but other disease-causing mutations. This easy flow cytometry-based assay allows a fast screening of patients with suspicion of LRBA deficiency reducing therefore the number of patients requiring LRBA sequencing and accelerating the treatment implementation. Detection of biallelic mutations in LRBA is however required for a definitive diagnosis.

  6. Rapid Flow Cytometry-Based Test for the Diagnosis of Lipopolysaccharide Responsive Beige-Like Anchor (LRBA) Deficiency.

    Science.gov (United States)

    Gámez-Díaz, Laura; Sigmund, Elena C; Reiser, Veronika; Vach, Werner; Jung, Sophie; Grimbacher, Bodo

    2018-01-01

    The diagnosis of lipopolysaccharide-responsive beige-like-anchor-protein (LRBA) deficiency currently relies on gene sequencing approaches that do not support a timely diagnosis and clinical management. We developed a rapid and sensitive test for clinical implementation based on the detection of LRBA protein by flow cytometry in peripheral blood cells after stimulation. LRBA protein was assessed in a prospective cohort of 54 healthy donors and 57 patients suspected of LRBA deficiency. Receiver operating characteristics analysis suggested an LRBA:MFI ratio cutoff point of 2.6 to identify LRBA-deficient patients by FACS with 94% sensitivity and 80% specificity and to discriminate them from patients with a similar clinical picture but other disease-causing mutations. This easy flow cytometry-based assay allows a fast screening of patients with suspicion of LRBA deficiency reducing therefore the number of patients requiring LRBA sequencing and accelerating the treatment implementation. Detection of biallelic mutations in LRBA is however required for a definitive diagnosis.

  7. Investigations of a Dual Seeded 1178 nm Raman Laser System

    Science.gov (United States)

    2016-01-14

    was obtained by Raman amplification of a distributed feedback diode laser in a variably strained polarization- maintaining fiber with a record-high...Calia, D.B., “50W CW visible laser source at 589 nm obtained via frequency doubling of three coherently combined narrow-band Raman fiber amplifiers...AFRL-RD-PS- TP-2016-0009 AFRL-RD-PS- TP-2016-0009 INVESTIGATIONS OF A DUAL SEEDED 1178 NM RAMAN LASER SYSTEM Leanne Henry, et al. 14 January

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

    DEFF Research Database (Denmark)

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

    2018-01-01

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

  9. Accuracy of Automated Flow Cytometry-Based Leukocyte Counts To Rule Out Urinary Tract Infection in Febrile Children: a Prospective Cross-Sectional Study

    Science.gov (United States)

    Duong, Hong Phuoc; Wissing, Karl Martin; Tram, Nathalie; Mascart, Georges; Lepage, Philippe

    2016-01-01

    Automated flow cytometry of urine remains an incompletely validated method to rule out urinary tract infection (UTI) in children. This cross-sectional analytical study was performed to compare the predictive values of flow cytometry and a dipstick test as initial diagnostic tests for UTI in febrile children and prospectively included 1,106 children (1,247 episodes). Urine culture was used as the gold standard test for diagnosing UTI. The performance of screening tests to diagnose UTI were established using receiver operating characteristic (ROC) analysis. Among these 1,247 febrile episodes, 221 UTIs were diagnosed (17.7% [95% confidence interval {CI}, 15.6 to 19.8%]). The area under the ROC curve for flow cytometry white blood cell (WBC) counts (0.99 [95% CI, 0.98 to 0.99]) was significantly superior to that for red blood cell (0.74 [95% CI, 0.70 to 0.78]) and bacterial counts (0.89 [95% CI, 0.87 to 0.92]) (P UTI in febrile children. PMID:27682127

  10. Dual-wavelength erbium-doped fiber laser with asymmetric fiber Bragg grating Fabry-Perot cavity

    Science.gov (United States)

    Chen, Cong; Xu, Zhi-wei; Wang, Meng; Chen, Hai-yan

    2014-11-01

    A novel dual-wavelength fiber laser with asymmetric fiber Bragg grating (FBG) Fabry-Perot (FP) cavity is proposed and experimentally demonstrated. A couple of uniform FBGs are used as the cavity mirrors, and the third FBG is used as intracavity wavelength selector by changing its operation temperature. Experimental results show that by adjusting the operation temperature of the intracavity wavelength selector, a tunable dual-wavelength laser emission can be achieved. The results demonstrate the new concept of dual-wavelength lasing with asymmetric FBG FP resonator and its technical feasibility.

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

    Science.gov (United States)

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

    2012-10-01

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

  12. Velocimetry of fast microscopic liquid jets by nanosecond dual-pulse laser illumination for megahertz X-ray free-electron lasers.

    Science.gov (United States)

    Grünbein, Marie Luise; Shoeman, Robert L; Doak, R Bruce

    2018-03-19

    To conduct X-ray Free-Electron Laser (XFEL) measurements at megahertz (MHz) repetition rates, sample solution must be delivered in a micron-sized liquid free-jet moving at up to 100 m/s. This exceeds by over a factor of two the jet speeds measurable with current high-speed camera techniques. Accordingly we have developed and describe herein an alternative jet velocimetry based on dual-pulse nanosecond laser illumination. Three separate implementations are described, including a small laser-diode system that is inexpensive and highly portable. We have also developed and describe analysis techniques to automatically and rapidly extract jet speed from dual-pulse images.

  13. Automated Analysis of Flow Cytometry Data to Reduce Inter-Lab Variation in the Detection of Major Histocompatibility Complex Multimer-Binding T Cells

    DEFF Research Database (Denmark)

    Pedersen, Natasja Wulff; Chandran, P. Anoop; Qian, Yu

    2017-01-01

    Manual analysis of flow cytometry data and subjective gate-border decisions taken by individuals continue to be a source of variation in the assessment of antigen-specific T cells when comparing data across laboratories, and also over time in individual labs. Therefore, strategies to provide...... automated analysis of major histocompatibility complex (MHC) multimer-binding T cells represent an attractive solution to decrease subjectivity and technical variation. The challenge of using an automated analysis approach is that MHC multimer-binding T cell populations are often rare and therefore...... laboratories. We used three different methods, FLOw Clustering without K (FLOCK), Scalable Weighted Iterative Flow-clustering Technique (SWIFT), and ReFlow to analyze flow cytometry data files from 28 laboratories. Each laboratory screened for antigen-responsive T cell populations with frequency ranging from 0...

  14. Implementation of laser speckle contrast analysis as connection kit for mobile phone for assessment of skin blood flow

    Science.gov (United States)

    Jakovels, Dainis; Saknite, Inga; Spigulis, Janis

    2014-05-01

    Laser speckle contrast analysis (LASCA) offers a non-contact, full-field, and real-time mapping of capillary blood flow and can be considered as an alternative method to Laser Doppler perfusion imaging. LASCA technique has been implemented in several commercial instruments. However, these systems are still too expensive and bulky to be widely available. Several optical techniques have found new implementations as connection kits for mobile phones thus offering low cost screening devices. In this work we demonstrate simple implementation of LASCA imaging technique as connection kit for mobile phone for primary low-cost assessment of skin blood flow. Stabilized 650 nm and 532 nm laser diode modules were used for LASCA illumination. Dual wavelength illumination could provide additional information about skin hemoglobin and oxygenation level. The proposed approach was tested for arterial occlusion and heat test. Besides, blood flow maps of injured and provoked skin were demonstrated.

  15. Digitally tunable dual wavelength emission from semiconductor ring lasers with filtered optical feedback

    International Nuclear Information System (INIS)

    Khoder, Mulham; Verschaffelt, Guy; Nguimdo, Romain Modeste; Danckaert, Jan; Leijtens, Xaveer; Bolk, Jeroen

    2013-01-01

    We report on a novel integrated approach to obtain dual wavelength emission from a semiconductor laser based on on-chip filtered optical feedback. Using this approach, we show experiments and numerical simulations of dual wavelength emission of a semiconductor ring laser. The filtered optical feedback is realized on-chip by employing two arrayed waveguide gratings to split/recombine light into different wavelength channels. Semiconductor optical amplifiers are placed in the feedback loop in order to control the feedback strength of each wavelength channel independently. By tuning the current injected into each of the amplifiers, we can effectively cancel the gain difference between the wavelength channels due to fabrication and material dichroism, thus resulting in stable dual wavelength emission. We also explore the accuracy needed in the operational parameters to maintain this dual wavelength emission. (letter)

  16. Real-time dual-comb spectroscopy with a free-running bidirectionally mode-locked fiber laser

    Science.gov (United States)

    Mehravar, S.; Norwood, R. A.; Peyghambarian, N.; Kieu, K.

    2016-06-01

    Dual-comb technique has enabled exciting applications in high resolution spectroscopy, precision distance measurements, and 3D imaging. Major advantages over traditional methods can be achieved with dual-comb technique. For example, dual-comb spectroscopy provides orders of magnitude improvement in acquisition speed over standard Fourier-transform spectroscopy while still preserving the high resolution capability. Wider adoption of the technique has, however, been hindered by the need for complex and expensive ultrafast laser systems. Here, we present a simple and robust dual-comb system that employs a free-running bidirectionally mode-locked fiber laser operating at telecommunication wavelength. Two femtosecond frequency combs (with a small difference in repetition rates) are generated from a single laser cavity to ensure mutual coherent properties and common noise cancellation. As the result, we have achieved real-time absorption spectroscopy measurements without the need for complex servo locking with accurate frequency referencing, and relatively high signal-to-noise ratio.

  17. Flow cytometry for intracellular SPION quantification: specificity and sensitivity in comparison with spectroscopic methods

    Directory of Open Access Journals (Sweden)

    Friedrich RP

    2015-06-01

    Full Text Available Ralf P Friedrich,1 Christina Janko,1 Marina Poettler,1 Philipp Tripal,1 Jan Zaloga,1 Iwona Cicha,1 Stephan Dürr,1,2 Johannes Nowak,3 Stefan Odenbach,3 Ioana Slabu,4 Maik Liebl,4 Lutz Trahms,4 Marcus Stapf,5 Ingrid Hilger,5 Stefan Lyer,1 Christoph Alexiou1 1Department of Otorhinolaryngology, Head and Neck Surgery, Section of Experimental Oncology and Nanomedicine, University hospital Erlangen, 2Department of Otorhinolaryngology, Head and Neck Surgery, Section of Phoniatrics and Pediatric Audiology, University hospital Erlangen, Erlangen, 3Technische Universität Dresden, Chair of Magnetofluiddynamics, Measuring and Automation Technology, Dresden, 4Physikalisch-Technische Bundesanstalt Berlin, Berlin, 5Department of Radiology, Division of Diagnostic and Interventional Radiology, Experimental Radiology, University hospital Jena, Jena, Germany Abstract: Due to their special physicochemical properties, iron nanoparticles offer new promising possibilities for biomedical applications. For bench to bedside translation of superparamagnetic iron oxide nanoparticles (SPIONs, safety issues have to be comprehensively clarified. To understand concentration-dependent nanoparticle-mediated toxicity, the exact quantification of intracellular SPIONs by reliable methods is of great importance. In the present study, we compared three different SPION quantification methods (ultraviolet spectrophotometry, magnetic particle spectroscopy, atomic adsorption spectroscopy and discussed the shortcomings and advantages of each method. Moreover, we used those results to evaluate the possibility to use flow cytometric technique to determine the cellular SPION content. For this purpose, we correlated the side scatter data received from flow cytometry with the actual cellular SPION amount. We showed that flow cytometry provides a rapid and reliable method to assess the cellular SPION content. Our data also demonstrate that internalization of iron oxide nanoparticles in human

  18. Dual Fine Tracking Control of a Satellite Laser Communication Uplink

    National Research Council Canada - National Science Library

    Noble, Louis A

    2006-01-01

    A dual fine tracking control system (FTCS) is developed for a single aperture optical communication receiver to compensate for high frequency disturbances affecting tracking of two incident laser communication beams...

  19. Dual-pump CARS measurements in a hydrogen diffusion flame in cross-flow with AC dielectric barrier discharge

    Science.gov (United States)

    Nishihara, Munetake; Freund, Jonathan B.; Glumac, Nick G.; Elliott, Gregory S.

    2018-03-01

    This paper presents dual-pump coherent anti-Stokes Raman scattering (CARS) measurements for simultaneous detection of flow temperature and relative concentration, applied to the characterization of a discharge-coupled reacting jet in a cross flow. The diagnostic is hydrogen Q-branch based, providing a much wider dynamic range compared to detection in the S-branch. For a previously developed dielectric barrier discharge, aligned co-axially with the fuel jet, OH planar laser induced fluorescence measurements show that the disturbance in the flame boundary leads to mixing enhancement. The H2-N2 dual-pump CARS measurement was used to map two-dimensional temperature distributions. The increase of the maximum temperature was up to 300 K, with 50% more H2 consumption, providing the reason for the decrease in the flame length by 25%. The increase of the relative H2O-H2 fraction was accompanied with a temperature increase, which indicates local equivalence ratios of below 1. The H2-O2 dual-pump measurements confirmed that the fuel-oxidizer ratios remain in the fuel-lean side at most of the probed locations.

  20. Laser streaming: Turning a laser beam into a flow of liquid.

    Science.gov (United States)

    Wang, Yanan; Zhang, Qiuhui; Zhu, Zhuan; Lin, Feng; Deng, Jiangdong; Ku, Geng; Dong, Suchuan; Song, Shuo; Alam, Md Kamrul; Liu, Dong; Wang, Zhiming; Bao, Jiming

    2017-09-01

    Transforming a laser beam into a mass flow has been a challenge both scientifically and technologically. We report the discovery of a new optofluidic principle and demonstrate the generation of a steady-state water flow by a pulsed laser beam through a glass window. To generate a flow or stream in the same path as the refracted laser beam in pure water from an arbitrary spot on the window, we first fill a glass cuvette with an aqueous solution of Au nanoparticles. A flow will emerge from the focused laser spot on the window after the laser is turned on for a few to tens of minutes; the flow remains after the colloidal solution is completely replaced by pure water. Microscopically, this transformation is made possible by an underlying plasmonic nanoparticle-decorated cavity, which is self-fabricated on the glass by nanoparticle-assisted laser etching and exhibits size and shape uniquely tailored to the incident beam profile. Hydrophone signals indicate that the flow is driven via acoustic streaming by a long-lasting ultrasound wave that is resonantly generated by the laser and the cavity through the photoacoustic effect. The principle of this light-driven flow via ultrasound, that is, photoacoustic streaming by coupling photoacoustics to acoustic streaming, is general and can be applied to any liquid, opening up new research and applications in optofluidics as well as traditional photoacoustics and acoustic streaming.

  1. Flow and mixing characteristics in a stirred tank with dual wide paddles; 2 dan waido padoruyoku tsuki kakuhan sonai no ryudo{center{underscore}dot}kongo tokusei

    Energy Technology Data Exchange (ETDEWEB)

    Takata, Kazutaka; Ito, Hisayoshi; Kikuchi, Masahiko; Okamoto, Yukimichi [Shinko Pantec Corp., Hyogo (Japan)

    1999-03-10

    Flow structure and mixing characteristics in a stirred tank with dual wide paddle impeller were examined using computational fluid dynamics (CFD). CFD was conducted using analysis code for fluid flow, and velocity measured by laser doppler velocimeter, power consumption and mixing time were used for evaluating computed results. The computed flow field and power consumption agreed well with themeasured values within 5 % mixing process well agreed with the observations. Since the computed flow pattern and mixing process agreed well with the measured values, computed results are useful for evaluating complex flow field in a stirred tank. A detailed investigation using computed results are useful for evaluating complex flow field in a stirred tank A detaile investigation using computed results shows that dual cross-installed wide paddle impellers lead to superior mixing performance in the stirred tank, and pressure gradient between upper and lower paddles is found to be the factor that promotes fluid transport in the tank, which is never the case when the dual wide paddles are installed in the same plane. (author)

  2. Use of flow cytometry for high-throughput cell population estimates in fixed brain tissue

    Directory of Open Access Journals (Sweden)

    Nicole A Young

    2012-07-01

    Full Text Available The numbers and types of cells in an area of cortex define its function. Therefore it is essential to characterize the numbers and distributions of total cells in areas of the cortex, as well as to identify numbers of subclasses of neurons and glial cells. To date, the large size of the primate brain and the lack of innovation in cell counting methods have been a roadblock to obtaining high-resolution maps of cell and neuron density across the cortex in humans and non-human primates. Stereological counting methods and the isotropic fractionator are valuable tools for estimating cell numbers, but are better suited to smaller, well-defined brain structures or to cortex as a whole. In the present study, we have extended our flow-cytometry based counting method, the flow fractionator (Collins et al., 2010a, to include high-throughput total cell population estimates in homogenized cortical samples. We demonstrate that our method produces consistent, accurate and repeatable cell estimates quickly. The estimates we report are in excellent agreement with estimates for the same samples obtained using a Neubauer chamber and a fluorescence microscope. We show that our flow cytometry-based method for total cell estimation in homogenized brain tissue is more efficient and more precise than manual counting methods. The addition of automated nuclei counting to our flow fractionator method allows for a fully automated, rapid characterization of total cells and neuronal and non-neuronal populations in human and non-human primate brains, providing valuable data to further our understanding of the functional organization of normal, aging and diseased brains.

  3. Global phenotypic characterisation of human platelet lysate expanded MSCs by high-throughput flow cytometry.

    Science.gov (United States)

    Reis, Monica; McDonald, David; Nicholson, Lindsay; Godthardt, Kathrin; Knobel, Sebastian; Dickinson, Anne M; Filby, Andrew; Wang, Xiao-Nong

    2018-03-02

    Mesenchymal stromal cells (MSCs) are a promising cell source to develop cell therapy for many diseases. Human platelet lysate (PLT) is increasingly used as an alternative to foetal calf serum (FCS) for clinical-scale MSC production. To date, the global surface protein expression of PLT-expended MSCs (MSC-PLT) is not known. To investigate this, paired MSC-PLT and MSC-FCS were analysed in parallel using high-throughput flow cytometry for the expression of 356 cell surface proteins. MSC-PLT showed differential surface protein expression compared to their MSC-FCS counterpart. Higher percentage of positive cells was observed in MSC-PLT for 48 surface proteins, of which 13 were significantly enriched on MSC-PLT. This finding was validated using multiparameter flow cytometry and further confirmed by quantitative staining intensity analysis. The enriched surface proteins are relevant to increased proliferation and migration capacity, as well as enhanced chondrogenic and osteogenic differentiation properties. In silico network analysis revealed that these enriched surface proteins are involved in three distinct networks that are associated with inflammatory responses, carbohydrate metabolism and cellular motility. This is the first study reporting differential cell surface protein expression between MSC-PLT and MSC-FSC. Further studies are required to uncover the impact of those enriched proteins on biological functions of MSC-PLT.

  4. Quantitative analysis of gold and carbon nanoparticles in mammalian cells by flow cytometry light scattering

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Gang [Nanjing University, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences (China); Liu, Naicheng; Wang, Zhenheng [Nanjing University, Department of Orthopedics, Jinling Hospital, School of Medicine (China); Shi, Tongguo; Gan, Jingjing; Wang, Zhenzhen; Zhang, Junfeng, E-mail: jfzhang@nju.edu.cn [Nanjing University, State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences (China)

    2017-02-15

    Nanoparticle-based applications for diagnostics and therapeutics have been extensively studied. These applications require a profound understanding of the fate of nanoparticles (NPs) in cellular environments. However, until now, few analytical methods are available and most of them rely on fluorescent properties or special elements of NPs; therefore, for NPs without observable optical properties or special elements, the existing methods are hardly applicable. In this study, we introduce a flow cytometry light scattering (FCLS)-based approach that quantifies in situ NPs accurately in mammalian cells. Continuous cells of heterogeneous human epithelial colorectal adenocarcinoma (Caco-2 cells), mouse peritoneal macrophages (MPM), and human adenocarcinomic alveolar basal epithelia (A549 cells) were cultured with NPs with certain concentrations and size. The intensity of the flow cytometric side scattered light, which indicates the quantity of NPs in the cells, was analyzed. The result shows an accurate size- and dose-dependent uptake of Au NPs (5, 30, 250 nm) in Caco-2 cells. The size- and dose- dependence of Au NPs (5, 30, 250 nm) and carbon NPs (50, 500 nm) in cells was validated by transmission electron microscope (TEM). This paper demonstrates the great potential of flow cytometry light scattering in the quantitative study of the size and dose effect on in situ metallic or non-metallic NPs in mammalian cells.

  5. Quantitative analysis of gold and carbon nanoparticles in mammalian cells by flow cytometry light scattering

    Science.gov (United States)

    Zhou, Gang; Liu, Naicheng; Wang, Zhenheng; Shi, Tongguo; Gan, Jingjing; Wang, Zhenzhen; Zhang, Junfeng

    2017-02-01

    Nanoparticle-based applications for diagnostics and therapeutics have been extensively studied. These applications require a profound understanding of the fate of nanoparticles (NPs) in cellular environments. However, until now, few analytical methods are available and most of them rely on fluorescent properties or special elements of NPs; therefore, for NPs without observable optical properties or special elements, the existing methods are hardly applicable. In this study, we introduce a flow cytometry light scattering (FCLS)-based approach that quantifies in situ NPs accurately in mammalian cells. Continuous cells of heterogeneous human epithelial colorectal adenocarcinoma (Caco-2 cells), mouse peritoneal macrophages (MPM), and human adenocarcinomic alveolar basal epithelia (A549 cells) were cultured with NPs with certain concentrations and size. The intensity of the flow cytometric side scattered light, which indicates the quantity of NPs in the cells, was analyzed. The result shows an accurate size- and dose-dependent uptake of Au NPs (5, 30, 250 nm) in Caco-2 cells. The size- and dose- dependence of Au NPs (5, 30, 250 nm) and carbon NPs (50, 500 nm) in cells was validated by transmission electron microscope (TEM). This paper demonstrates the great potential of flow cytometry light scattering in the quantitative study of the size and dose effect on in situ metallic or non-metallic NPs in mammalian cells.

  6. 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.

  7. Zinc fixation preserves flow cytometry scatter and fluorescence parameters and allows simultaneous analysis of DNA content and synthesis, and intracellular and surface epitopes

    DEFF Research Database (Denmark)

    Jensen, Uffe Birk; Owens, David; Pedersen, Søren

    2010-01-01

    Zinc salt-based fixation (ZBF) has proved advantageous in histochemical analyses conducted on intact tissues but has not been exploited in flow cytometry procedures that focus on quantitative analysis of individual cells. Here, we show that ZBF performs equally well to paraformaldehyde in the pre......Zinc salt-based fixation (ZBF) has proved advantageous in histochemical analyses conducted on intact tissues but has not been exploited in flow cytometry procedures that focus on quantitative analysis of individual cells. Here, we show that ZBF performs equally well to paraformaldehyde...... allowing subsequent quantitative PCR analysis or labeling for incorporation of the thymidine analog EdU following surface and intracellular epitope staining. Finally, ZBF treatment allows for long-term storage of labeled cells with little change in these parameters. Thus, we present a protocol for zinc...... salt fixation of cells that allows for the simultaneous analysis of DNA and intracellular and cell surface proteins by flow cytometry....

  8. Development of an unbiased, semi-automated approach for classifying plasma cell immunophenotype following multicolor flow cytometry of bone marrow aspirates.

    Science.gov (United States)

    Post, Steven R; Post, Ginell R; Nikolic, Dejan; Owens, Rebecca; Insuasti-Beltran, Giovanni

    2018-03-24

    Despite increased usage of multiparameter flow cytometry (MFC) to assess diagnosis, prognosis, and therapeutic efficacy (minimal residual disease, MRD) in plasma cell neoplasms (PCNs), standardization of methodology and data analysis is suboptimal. We investigated the utility of using the mean and median fluorescence intensities (FI) obtained from MFC to objectively describe parameters that distinguish plasma cell (PC) phenotypes. In this retrospective study, flow cytometry results from bone marrow aspirate specimens from 570 patients referred to the Myeloma Institute at UAMS were evaluated. Mean and median FI data were obtained from 8-color MFC of non-neoplastic, malignant, and mixed PC populations using antibodies to CD38, CD138, CD19, CD20, CD27, CD45, CD56, and CD81. Of 570 cases, 252 cases showed only non-neoplastic PCs, 168 showed only malignant PCs, and 150 showed mixed PC populations. Statistical analysis of median FI data for each CD marker showed no difference in expression intensity on non-neoplastic and malignant PCs, between pure and mixed PC populations. ROC analysis of the median FI of CD expression in non-neoplastic and malignant PCs was used to develop an algorithm to convert quantitative FI values to qualitative assessments including "negative," "positive," "dim," and "heterogeneous" expression. FI data derived from 8-color MFC can be used to define marker expression on PCs. Translation of FI data from Infinicyt software to an Excel worksheet streamlines workflow and eliminates transcriptional errors when generating flow reports. © 2018 International Clinical Cytometry Society. © 2018 International Clinical Cytometry Society.

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

    Directory of Open Access Journals (Sweden)

    Tomoyuki Kaneko

    2011-06-01

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

  10. Flow cytometry: design, development and experimental validation

    International Nuclear Information System (INIS)

    Seigneur, Alain

    1987-01-01

    The flow cytometry techniques allow the analysis and sorting of living biologic cells at rates above five to ten thousand events per second. After a short review, we present in this report the design and development of a 'high-tech' apparatus intended for research laboratories and the experimental results. The first part deals with the physical principles allowing morphologic and functional analysis of cells or cellular components. The measured parameters are as follows: electrical resistance pulse sizing, light scattering and fluorescence. Hydrodynamic centering is used, and in the same way, the division of a water-stream into droplets leading to electrostatic sorting of particles. The second part deals with the apparatus designed by the 'Commissariat a l'Energie Atomique' (C.E.A.) and industrialised by 'ODAM' (ATC 3000). The last part of this thesis work is the performance evaluations of this cyto-meter. The difference between the two size measurement methods are analyzed: electrical resistance pulse sizing versus small-angle light scattering. By an original optics design, high sensitivity has been reached in the fluorescence measurement: the equivalent noise corresponds to six hundred fluorescein isothiocyanate (FITC) molecules. The sorting performances have also been analyzed and the cell viability proven. (author) [fr

  11. Operational characteristics of dual gain single cavity Nd:YVO 4 laser

    Indian Academy of Sciences (India)

    Operational characteristics of a dual gain single cavity Nd:YVO4 laser have been investigated. With semiconductor diode laser pump power of 2 W, 800 mW output was obtained with a slope efficiency of 49%. Further, by changing the relative orientation of the two crystals the polarization characteristics of the output could be ...

  12. Dual Solutions for Nonlinear Flow Using Lie Group Analysis.

    Directory of Open Access Journals (Sweden)

    Muhammad Awais

    Full Text Available `The aim of this analysis is to investigate the existence of the dual solutions for magnetohydrodynamic (MHD flow of an upper-convected Maxwell (UCM fluid over a porous shrinking wall. We have employed the Lie group analysis for the simplification of the nonlinear differential system and computed the absolute invariants explicitly. An efficient numerical technique namely the shooting method has been employed for the constructions of solutions. Dual solutions are computed for velocity profile of an upper-convected Maxwell (UCM fluid flow. Plots reflecting the impact of dual solutions for the variations of Deborah number, Hartman number, wall mass transfer are presented and analyzed. Streamlines are also plotted for the wall mass transfer effects when suction and blowing situations are considered.

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

    Science.gov (United States)

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

    2013-05-01

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

  14. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry

    Directory of Open Access Journals (Sweden)

    L.P.S. Alves

    Full Text Available The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i cell permeabilization, ii Nile red staining, and iii analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99 compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots.

  15. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry.

    Science.gov (United States)

    Alves, L P S; Almeida, A T; Cruz, L M; Pedrosa, F O; de Souza, E M; Chubatsu, L S; Müller-Santos, M; Valdameri, G

    2017-01-16

    The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots.

  16. A simple and efficient method for poly-3-hydroxybutyrate quantification in diazotrophic bacteria within 5 minutes using flow cytometry

    Science.gov (United States)

    Alves, L.P.S.; Almeida, A.T.; Cruz, L.M.; Pedrosa, F.O.; de Souza, E.M.; Chubatsu, L.S.; Müller-Santos, M.; Valdameri, G.

    2017-01-01

    The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method was successfully applied to the quantification of PHB in epiphytic bacteria isolated from rice roots. PMID:28099582

  17. Improved graft survival in highly sensitized patients undergoing renal transplantation after the introduction of a clinically validated flow cytometry crossmatch.

    LENUS (Irish Health Repository)

    Limaye, Sandhya

    2009-04-15

    Flow cytometric techniques are increasingly used in pretransplant crossmatching, although there remains debate regarding the clinical significance and predictive value of donor-specific antibodies detected by flow cytometry. At least some of the discrepancies between published studies may arise from differences in cutoffs used and lack of standardization of the test.

  18. Multiparameter flow cytometry reveals myelodysplasia-related aberrant antigen expression in myelodysplastic/myeloproliferative neoplasms.

    Science.gov (United States)

    Kern, Wolfgang; Bacher, Ulrike; Schnittger, Susanne; Alpermann, Tamara; Haferlach, Claudia; Haferlach, Torsten

    2013-05-01

    Within the myelodysplastic/myeloproliferative neoplasm (MDS/MPN) category of the WHO (2008), only chronic myelomonocytic leukemia was so far evaluated by multiparameter flow cytometry (MFC). To investigate the potential of MFC for MDS/MPNs, unclassifiable (MDS/MPNu), and refractory anemia associated with ring sideroblasts and marked thrombocytosis (RARS-T), we studied 91 patients with these entities (60 males/31 females; 35.3-87.4 years) for MDS-related aberrant immunophenotypes (≥ 2 different cell lineages with ≥ 3 aberrantly expressed antigens). Data were correlated with cytomorphology and cytogenetics. MFC identified MDS-related immunophenotypes in 54/91 (59.3%) of patients. Patients with or without MDS-related immunophenotype did not differ significantly by demographic characteristics, blood values, or median overall survival. MDS-related immunophenotype cases showed a higher number of aberrantly expressed antigens (mean ± SD, 4.9 ± 2.4 vs. 2.0 ± 1.4; P MPNu and RARS-T. MFC therefore may be helpful to separate cases into more "MDS-like" or "MPN-like" subgroups. Copyright © 2012 International Clinical Cytometry Society.

  19. Synthetic aperture flow imaging using dual stage beamforming

    DEFF Research Database (Denmark)

    Li, Ye; Jensen, Jørgen Arendt

    2013-01-01

    A method for synthetic aperture flow imaging using dual stage beamforming has been developed. The main motivation is to increase the frame rate and still maintain a beamforming quality sufficient for flow estimation that is possible to implement in a commercial scanner. This method can generate...

  20. Blood group antigen studies using CdTe quantum dots and flow cytometry

    Directory of Open Access Journals (Sweden)

    Cabral Filho PE

    2015-07-01

    Full Text Available Paulo E Cabral Filho,1 Maria IA Pereira,1 Heloise P Fernandes,2 Andre A de Thomaz,3 Carlos L Cesar,3 Beate S Santos,4 Maria L Barjas-Castro,2 Adriana Fontes1 1Departamento de Biofísica e Radiobiologia, Universidade Federal de Pernambuco, Recife, Pernambuco, 2Centro de Hematologia e Hemoterapia, Universidade Estadual de Campinas, Instituto Nacional de Ciência e Tecnologia do Sangue, Campinas, São Paulo, 3Departamento de Eletrônica Quântica, Instituto de Física Gleb Wataghin, Universidade Estadual de Campinas, Campinas, São Paulo, 4Departamento de Ciências Farmacêuticas, Universidade Federal de Pernambuco, Recife, PE, Brazil Abstract: New methods of analysis involving semiconductor nanocrystals (quantum dots [QDs] as fluorescent probes have been highlighted in life science. QDs present some advantages when compared to organic dyes, such as size-tunable emission spectra, broad absorption bands, and principally exceptional resistance to photobleaching. Methods applying QDs can be simple, not laborious, and can present high sensibility, allowing biomolecule identification and quantification with high specificity. In this context, the aim of this work was to apply dual-color CdTe QDs to quantify red blood cell (RBC antigen expression on cell surface by flow cytometric analysis. QDs were conjugated to anti-A or anti-B monoclonal antibodies, as well as to the anti-H (Ulex europaeus I lectin, to investigate RBCs of A1, B, A1B, O, A2, and Aweak donors. Bioconjugates were capable of distinguishing the different expressions of RBC antigens, both by labeling efficiency and by flow cytometry histogram profile. Furthermore, results showed that RBCs from Aweak donors present fewer amounts of A antigens and higher amounts of H, when compared to A1 RBCs. In the A group, the amount of A antigens decreased as A1 > A3 > AX = Ael, while H antigens were AX = Ael > A1. Bioconjugates presented stability and remained active for at least 6 months. In conclusion

  1. Unidirectional, dual-comb lasing under multiple pulse formation mechanisms in a passively mode-locked fiber ring laser

    OpenAIRE

    Liu, Ya; Zhao, Xin; Hu, Guoqing; Li, Cui; Zhao, Bofeng; Zheng, Zheng

    2016-01-01

    Dual-comb lasers from which asynchronous ultrashort pulses can be simultaneously generated have recently become an interesting research subject. They could be an intriguing alternative to the current dual-laser optical-frequency-comb source with highly sophisticated electronic control systems. If generated through a common light path traveled by all pulses, the common-mode noises between the spectral lines of different pulse trains could be significantly reduced. Therefore, coherent dual-comb...

  2. Standardization of 8-color flow cytometry across different flow cytometer instruments: A feasibility study in clinical laboratories in Switzerland.

    Science.gov (United States)

    Glier, Hana; Heijnen, Ingmar; Hauwel, Mathieu; Dirks, Jan; Quarroz, Stéphane; Lehmann, Thomas; Rovo, Alicia; Arn, Kornelius; Matthes, Thomas; Hogan, Cassandra; Keller, Peter; Dudkiewicz, Ewa; Stüssi, Georg; Fernandez, Paula

    2017-07-29

    The EuroFlow Consortium developed a fully standardized flow cytometric approach from instrument settings, through antibody panel, reagents and sample preparation protocols, to data acquisition and analysis. The Swiss Cytometry Society (SCS) promoted a study to evaluate the feasibility of using such standardized measurements of 8-color data across two different flow cytometry platforms - Becton Dickinson (BD) FACSCanto II and Beckman Coulter (BC) Navios, aiming at increasing reproducibility and inter-laboratory comparability of immunophenotypic data in clinical laboratories in Switzerland. The study was performed in two phases, i.e. a learning phase (round 1) and an analytical phase (rounds 2 and 3) consisting of a total of three rounds. Overall, 10 laboratories using BD FACSCanto II (n=6) or BC Navios (n=4) flow cytometers participated. Each laboratory measured peripheral blood samples from healthy donors stained with a uniform antibody panel of reagents - EuroFlow Lymphoid Screening Tube (LST) - applying the EuroFlow standardized protocols for instrument setup and sample preparation (www.EuroFlow.org). All data files were analyzed centrally and median fluorescence intensity (MedFI) values for individual markers on defined lymphocyte subsets were recorded; variability from reference MedFI values was assessed using performance scores. Data troubleshooting and discussion of the results with the participants followed after each round at SCS meetings. The results of the learning phase demonstrated that standardized instrument setup and data acquisition are feasible in routine clinical laboratories without previous experience with EuroFlow. During the analytical phase, highly comparable data were obtained at the different laboratories using either BD FACSCanto II or BC Navios. The coefficient of variation of MedFI for 7 of 11 markers performed repeatedly below 30%. In the last study round, 89% of participants scored over 90% MedFI values within the acceptance criteria

  3. Flow cytometry, microsatellites and niche models reveal the origins and geographical structure of Alnus glutinosa populations in Europe

    Czech Academy of Sciences Publication Activity Database

    Mandák, Bohumil; Vít, Petr; Krak, Karol; Trávníček, Pavel; Havrdová, Alena; Hadincová, Věroslava; Zákravský, Petr; Jarolímová, Vlasta; Bacles, C. F. E.; Douda, Jan

    2016-01-01

    Roč. 117, č. 1 (2016), s. 107-120 ISSN 0305-7364 R&D Projects: GA ČR(CZ) GAP504/11/0402 Institutional support: RVO:67985939 Keywords : flow cytometry * microsatellites * glacial refugia Subject RIV: EF - Botanics Impact factor: 4.041, year: 2016

  4. B-ALL minimal residual disease flow cytometry: an application of a novel method for optimization of a single-tube model.

    Science.gov (United States)

    Shaver, Aaron C; Greig, Bruce W; Mosse, Claudio A; Seegmiller, Adam C

    2015-05-01

    Optimizing a clinical flow cytometry panel can be a subjective process dependent on experience. We develop a quantitative method to make this process more rigorous and apply it to B lymphoblastic leukemia/lymphoma (B-ALL) minimal residual disease (MRD) testing. We retrospectively analyzed our existing three-tube, seven-color B-ALL MRD panel and used our novel method to develop an optimized one-tube, eight-color panel, which was tested prospectively. The optimized one-tube, eight-color panel resulted in greater efficiency of time and resources with no loss in diagnostic power. Constructing a flow cytometry panel using a rigorous, objective, quantitative method permits optimization and avoids problems of interdependence and redundancy in a large, multiantigen panel. Copyright© by the American Society for Clinical Pathology.

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

    Science.gov (United States)

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

    2018-06-01

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

  6. Applications of ultra-short pulsed laser ablation: thin films deposition and fs/ns dual-pulse laser-induced breakdown spectroscopy

    International Nuclear Information System (INIS)

    Teghil, R; De Bonis, A; Galasso, A; Santagata, A; Albano, G; Villani, P; Spera, D; Parisi, G P

    2008-01-01

    In this paper, we report a survey of two of the large number of possible practical applications of the laser ablation performed by an ultra-short pulse laser, namely pulsed laser deposition (PLD) and fs/ns dual-pulse laser-induced breakdown spectroscopy (DP-LIBS). These applications differ from those using just longer pulsed lasers as a consequence of the distinctive characteristics of the plasma produced by ultra-short laser beams. The most important feature of this plasma is the large presence of particles with nanometric size which plays a fundamental role in both applications.

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

    NARCIS (Netherlands)

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

    1993-01-01

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

  8. Enhancement of the static extinction ratio by using a dual-section distributed feedback laser integrated with an electro-absorption modulator

    Science.gov (United States)

    Cho, Chun-Hyung; Kim, Jongseong; Sung, Hyuk-Kee

    2016-09-01

    We report on the enhancement of the static extinction ratio by using a dual-section distributed feedback laser diode integrated with an electro-absorption modulator. A directly- modulated dual-section laser can provide improved modulation performance under a low bias level ( i.e., below the threshold level) compared with a standard directly-modulated laser. By combining the extinction ratio from a dual-section laser with that from an electro-absorption modulator section, a total extinction ratio of 49.6. dB are successfully achieved.

  9. Flow cytometry analysis of inflammatory cells isolated from the sciatic nerve and DRG after chronic constriction injury in mice.

    Science.gov (United States)

    Liu, Liping; Yin, Yan; Li, Fei; Malhotra, Charvi; Cheng, Jianguo

    2017-06-01

    Cellular responses to nerve injury play a central role in the pathogenesis of neuropathic pain. However, the analysis of site specific cellular responses to nerve injury and neuropathic pain is limited to immunohistochemistry staining with numerous limitations. We proposed to apply flow cytometry to overcome some of the limitations and developed two protocols for isolation of cells from small specimens of the sciatic nerve and dorsal root ganglion (DRG) in mice. RESULTS AND COMPARASION WITH EXISTING: methods We found that both the non-enzymatic and enzymatic approaches were highly effective in harvesting a sufficient number of cells for flow cytometry analysis in normal and pathological conditions. The total number of cells in the injury site of the sciatic and its DRGs increased significantly 14days after chronic constriction injury (CCI) of the sciatic nerve, compared to sham surgery control or the contralateral control. The enzymatic approach yielded a significantly higher total number of cells and CD45 negative cells, suggesting that this approach allows for harvest of more resident cells, compared to the non-enzymatic method. The percentage of CD45 + /CD11b + cells was significantly increased in the sciatic nerve but not in the DRG. These results were consistent with both protocols. We thus offer two simple and effective protocols that allow for application of flow cytometry to the investigation of cellular and molecular mechanisms of neuropathic pain. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Effects of laser wavelengths and pulse energy ratio on the emission enhancement in dual pulse LIBS

    International Nuclear Information System (INIS)

    Ahmed, Rizwan; Iqbal, Javed; Baig, M Aslam

    2015-01-01

    We present new studies on the effects of laser wavelengths, pulse energy ratio and interpulse delay between two laser pulses in the collinear dual pulse configuration of laser-induced breakdown spectroscopy (LIBS) on an iron sample in air using the fundamental (1064 nm) and the second harmonics (532 nm) of Nd:YAG lasers. In the dual pulse LIBS, an optimum value of interpulse delay with an appropriate combination of laser wavelengths, and laser pulse energy ratio, yields a 30 times signal intensity enhancement in the neutral iron lines as compared with single pulse LIBS. A comparison in the spatial variations of electron temperature along the axis of the plume expansion in single and double pulse LIBS has also been studied. (letter)

  11. Dual-beam operation of the Astra Gemini laser facility

    International Nuclear Information System (INIS)

    Bryan Parry; Nicola Booth; Oleg Chekhlov; John Collier; Edwin Divall; Klaus Ertel; Peta Foster; Steve Hawkes; Chris Hooker; Victoria Marshall

    2010-01-01

    Complete text of publication follows. Gemini is a Petawatt class Ti:Sapphire laser system at the Rutherford Appleton Laboratory, UK. It was designed as a dual beam laser, with two independently configurable 800 nm beams delivering 15 J to target in 30 fs pulse duration, giving 0.5 PW peak power per beam. It is capable of reaching intensities over 10 22 W/cm 2 . Gemini can achieve a maximum repetition rate of one shot every 20 seconds, allowing it to deliver hundreds of shots per day; a feature which makes it unique among PW lasers. Already this has proved valuable in experiments involving electron acceleration in gas jets. The first Gemini beamline became operational in 2008. Commissioning of the second beam was deferred to allow earlier access to the facility by experimental scientists, and to develop operational experience. In this mode, Gemini has already produced significant results from a number of advanced plasma physics experiments. The second beam of Gemini is now coming online, with the first dual beam experiment starting in June 2010. The flexibility offered by two short pulse, ultra high intensity beams is another aspect that makes this laser system unique. The dual beams enable versatile configurations and illumination geometries, facilitating a wider range of experiments than is possible with only a single beam. Operationally however, it introduces additional factors which must be monitored and controlled in order to achieve experimental success. The beams must be timed with respect to each other with accuracy less than the pulse duration. The beam foci must also be overlapped spatially, and the stability of both these factors maintained over extended periods. We report on the second beam commissioning process, including the latest results on the characteristics, stability and spatio-temporal overlap of the two beams. We present details of amplifier performance, along with measurements of beam quality, focal spot, pulse duration and contrast, to give a

  12. Asynchronous and synchronous dual-wavelength pulse generation in a passively mode-locked fiber laser with a mode-locker.

    Science.gov (United States)

    Hu, Guoqing; Pan, Yingling; Zhao, Xin; Yin, Siyao; Zhang, Meng; Zheng, Zheng

    2017-12-01

    The evolution from asynchronous to synchronous dual-wavelength pulse generation in a passively mode-locked fiber laser is experimentally investigated by tailoring the intracavity dispersion. Through tuning the intracavity-loss-dependent gain profile and the birefringence-induced filter effect, asynchronous dual-wavelength soliton pulses can be generated until the intracavity anomalous dispersion is reduced to ∼8  fs/nm. The transition from asynchronous to synchronous pulse generation is then observed at an elevated pump power in the presence of residual anomalous dispersion, and it is shown that pulses are temporally synchronized at the mode-locker in the cavity. Spectral sidelobes are observed and could be attributed to the four-wave-mixing effect between dual-wavelength pulses at the carbon nanotube mode-locker. These results could provide further insight into the design and realization of such dual-wavelength ultrafast lasers for different applications such as dual-comb metrology as well as better understanding of the inter-pulse interactions in such dual-comb lasers.

  13. Numerical simulation of air flow through turbocharger compressors with dual volute design

    Energy Technology Data Exchange (ETDEWEB)

    Jiao, Kui; Li, Xianguo; Wu, Hao [Department of Mechanical and Mechatronics Engineering, University of Waterloo, Waterloo, ON (Canada); Sun, Harold; Schram, Tim [Ford Motor Company, Dearborn, MI 48126 (United States); Krivitzky, Eric; Larosiliere, Louis M. [Concepts NREC, White River Junction, VT 05001 (United States)

    2009-11-15

    In this paper, turbocharger centrifugal compressors with dual volute design were investigated by using Computational Fluid Dynamics (CFD) method. The numerical simulation focused on the air flow from compressor impeller inlet to volute exit, and the overall performance level and range are predicted. The numerical investigation revealed that the dual volute design could separate the compressor into two operating regions: ''high efficiency'' and ''low efficiency'' regions with different air flow characteristics, and treating these two regions separately with dual diffuser design showed extended stable operating range and improved efficiency by comparing with conventional single volute design. The ''dual sequential volute'' concept also showed the potential to further extend the stable operating range by closing one of the volutes at low air flow rates. Furthermore, by comparing with other alternate designs such as variable diffuser vanes and variable inlet guide vanes, the operation of the dual sequential volute also features relatively simple control and calibration. (author)

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

    Science.gov (United States)

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

    2012-01-01

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

  15. Multiparametric flow cytometry in the diagnosis and characterization of low-grade pulmonary mucosa-associated lymphoid tissue lymphomas.

    Science.gov (United States)

    Zaer, F S; Braylan, R C; Zander, D S; Iturraspe, J A; Almasri, N M

    1998-06-01

    Primary mucosa associated lymphoid tissue (MALT) lymphomas are rare neoplasms that seem to have a better prognosis than nodal lymphomas. Morphologic diagnosis of these lesions may be difficult because of features that overlap with those of benign lymphoid infiltrates. In this study, we assessed the contribution of multi-parametric flow cytometry in demonstrating clonality and further characterizing pulmonary MALT lymphomas. Based on a clinical or pathologic suspicion of MALT-lymphoma, 3 transbronchial biopsies, 4 fine needle aspirates, 1 core needle biopsy, and 13 wedge excisions of lung were submitted fresh (unfixed) to our laboratory for evaluation. Among the 13 cases diagnosed as MALT lymphomas, B-cell monoclonality was established by identifying expression of a single immunoglobulin light chain on CD20 or CD19-positive cells in 12 cases. One case lacked expression of both light chains on B-cells. Of 11 lymphoma cases in which CD5 and CD10 surface antigens were assessed, no cases expressed CD10, and 1 case demonstrated weak CD5 expression. Nine of 10 cases studied were diploid and 1 case was hyperdiploid. All of the lymphomas displayed low (< or = 3%) S-phase fractions consistent with low grade processes. In 10 patients with short follow-up, none died of their disease and the majority had no evidence of lymphoma dissemination. In seven of the remaining eight cases, B-cells were polyclonal consistent with reactive processes. In one morphologically reactive case, flow cytometric analysis was unsuccessful because of poor cell viability. The pulmonary MALT lymphomas in this study represent a group of B-cell tumors with distinctive morphologic, immunophenotypic, and cell kinetic characteristics. Multi-parametric flow cytometry is useful for confirming B-cell monoclonality and illustrating an antigenic profile compatible with this diagnosis. Flow cytometry can be particularly helpful when working with small biopsies and cytologic samples with limited diagnostic

  16. Dual-wavelength external cavity laser device for fluorescence suppression in Raman spectroscopy

    Science.gov (United States)

    Zhang, Xuting; Cai, Zhijian; Wu, Jianhong

    2017-10-01

    Raman spectroscopy has been widely used in the detection of drugs, pesticides, explosives, food additives and environmental pollutants, for its characteristics of fast measurement, easy sample preparation, and molecular structure analyzing capability. However, fluorescence disturbance brings a big trouble to these applications, with strong fluorescence background covering up the weak Raman signals. Recently shifted excitation Raman difference spectroscopy (SERDS) not only can completely remove the fluorescence background, but also can be easily integrated into portable Raman spectrometers. Usually, SERDS uses two lasers with small wavelength gap to excite the sample, then acquires two spectra, and subtracts one to the other to get the difference spectrum, where the fluorescence background will be rejected. So, one key aspects of successfully applying SERDS method is to obtain a dual-wavelength laser source. In this paper, a dual-wavelength laser device design based on the principles of external cavity diode laser (ECDL) is proposed, which is low-cost and compact. In addition, it has good mechanical stability because of no moving parts. These features make it an ideal laser source for SERDS technique. The experiment results showed that the device can emit narrow-spectral-width lasers of two wavelengths, with the gap smaller than 2 nanometers. The laser power corresponding to each wavelength can be up to 100mW.

  17. Immunological Tools: Engaging Students in the Use and Analysis of Flow Cytometry and Enzyme-linked Immunosorbent Assay (ELISA)

    Science.gov (United States)

    Ott, Laura E.; Carson, Susan

    2014-01-01

    Flow cytometry and enzyme-linked immunosorbent assay (ELISA) are commonly used techniques associated with clinical and research applications within the immunology and medical fields. The use of these techniques is becoming increasingly valuable in many life science and engineering disciplines as well. Herein, we report the development and…

  18. Dual-beam laser welding of AZ31B magnesium alloy in zero-gap lap joint configuration

    Science.gov (United States)

    Harooni, Masoud; Carlson, Blair; Kovacevic, Radovan

    2014-03-01

    Porosity within laser welds of magnesium alloys is one of the main roadblocks to achieving high quality joints. One of the causes of pore formation is the presence of pre-existing coatings on the surface of magnesium alloy such as oxide or chromate layers. In this study, single-beam and dual-beam laser heat sources are investigated in relation to mitigation of pores resulting from the presence of the as-received oxide layer on the surface of AZ31B-H24 magnesium alloy during the laser welding process. A fiber laser with a power of up to 4 kW is used to weld samples in a zero-gap lap joint configuration. The effect of dual-beam laser welding with different beam energy ratios is studied on the quality of the weld bead. The purpose of this paper is to identify the beam ratio that best mitigates pore formation in the weld bead. The laser molten pool and the keyhole condition, as well as laser-induced plasma plume are monitored in real-time by use of a high speed charge-coupled device (CCD) camera assisted with a green laser as an illumination source. Tensile and microhardness tests were used to measure the mechanical properties of the laser welded samples. Results showed that a dual-beam laser configuration can effectively mitigate pore formation in the weld bead by a preheating-welding mechanism.

  19. Instrument for real-time pulse-shape analysis of slit-scan flow cytometry signals

    NARCIS (Netherlands)

    van Oven, C.; Aten, J. A.

    1990-01-01

    An instrument is described which analyses shapes of fluorescence profiles generated by particles passing through the focussed laser beam of a flow cytometer. The output signal of this pulse-shape analyzer is used as input for the signal processing electronics of a commercial flow cytometer system.

  20. Detection of microparticles from human red blood cells by multiparametric flow cytometry

    Science.gov (United States)

    Grisendi, Giulia; Finetti, Elena; Manganaro, Daniele; Cordova, Nicoletta; Montagnani, Giuliano; Spano, Carlotta; Prapa, Malvina; Guarneri, Valentina; Otsuru, Satoru; Horwitz, Edwin M.; Mari, Giorgio; Dominici, Massimo

    2015-01-01

    Background During storage, red blood cells (RBC) undergo chemical and biochemical changes referred to as “storage lesions”. These events determine the loss of RBC integrity, resulting in lysis and release of microparticles. There is growing evidence of the clinical importance of microparticles and their role in blood transfusion-related side effects and pathogen transmission. Flow cytometry is currently one of the most common techniques used to quantify and characterise microparticles. Here we propose multiparametric staining to monitor and quantify the dynamic release of microparticles by stored human RBC. Material and methods RBC units (n=10) were stored under blood bank conditions for up to 42 days. Samples were tested at different time points to detect microparticles and determine the haemolysis rate (HR%). Microparticles were identified by flow cytometry combining carboxyfluorescein diacetate succinimidyl ester (CFSE) dye, annexin V and anti-glycophorin A antibody. Results We demonstrated that CFSE can be successfully used to label closed vesicles with an intact membrane. The combination of CFSE and glycophorin A antibody was effective for monitoring and quantifying the dynamic release of microparticles from RBC during storage. Double staining with CFSE/glycophorin A was a more precise approach, increasing vesicle detection up to 4.7-fold vs the use of glycophorin A/annexin V alone. Moreover, at all the time points tested, we found a robust correlation (R=0.625; p=0.0001) between HR% and number of microparticles detected. Discussion Multiparametric staining, based on a combination of CFSE, glycophorin A antibody and annexin V, was able to detect, characterise and monitor the release of microparticles from RBC units during storage, providing a sensitive approach to labelling and identifying microparticles for transfusion medicine and, more broadly, for cell-based therapies. PMID:25369588

  1. The value of calcaneal bone mass measurement using a dual X-ray laser calscan device in risk screening for osteoporosis

    Directory of Open Access Journals (Sweden)

    Gulseren Kayalar

    2009-01-01

    Full Text Available OBJECTIVE: To evaluate how bone mineral density in the calcaneus measured by a dual energy X-ray laser (DXL correlates with bone mineral density in the spine and hip in Turkish women over 40 years of age and to determine whether calcaneal dual energy X-ray laser variables are associated with clinical risk factors to the same extent as axial bone mineral density measurements obtained using dual energy x-ray absorbtiometry (DXA. MATERIALS AND METHODS: A total of 2,884 Turkish women, aged 40-90 years, living in Ankara were randomly selected. Calcaneal bone mineral density was evaluated using a dual energy X-ray laser Calscan device. Subjects exhibiting a calcaneal dual energy X-ray laser T- score <-2.5 received a referral for DXA of the spine and hip. Besides dual energy X-ray laser measurements, all subjects were questioned about their medical history and the most relevant risk factors for osteoporosis. RESULTS: Using a T-score threshold of -2.5, which is recommended by the World Health Organization (WHO, dual energy X-ray laser calcaneal measurements showed that 13% of the subjects had osteoporosis, while another 56% had osteopenia. The mean calcaneal dual energy X-ray laser T-score of postmenopausal subjects who were smokers with a positive history of fracture, hormone replacement therapy (HRT, covered dressing style, lower educational level, no regular exercise habits, and low tea consumption was significantly lower than that obtained for the other group (p<0.05. A significant correlation was observed between the calcaneal dual energy X-ray laser T-score and age (r=-0.465, p=0.001, body mass index (BMI (r=0.223, p=0.001, number of live births (r=-0.229, p=0.001, breast feeding time (r=-0.064, p=0.001, and age at menarche (r=-0.050, p=0.008. The correlations between calcaneal DXL and DXA T-scores (r=0.340, p=0.001 and calcaneal DXL and DXA Z-scores (r=0.360, p=0.001 at the spine, and calcaneal DXL and DXA T- scores (r=0.28, p=0.001 and calcaneal

  2. Tunable and stable single-longitudinal-mode dual-wavelength erbium fiber laser with 1.3 nm mode spacing output

    International Nuclear Information System (INIS)

    Yeh, C H; Shih, F Y; Wang, C H; Chow, C W; Chi, S

    2008-01-01

    In this investigation, we propose and investigate a stable and tunable dual-wavelength erbium-doped fiber (EDF) ring laser with self-injected Fabry-Perot laser diode (FP-LD) scheme. By using an FP-LD incorporated with a tunable bandpass filter (TBF) within the gain cavity, the fiber laser can lase at two single-longitudinal-mode (SLM) wavelengths simultaneously due to the self-injected operation. The proposed dual-wavelength laser has a good performance of the output power and optical side-mode suppression ratio (SMSR). The laser also shows a wide tuning range from 1523.08 to 1562.26 nm. Besides, the output stabilities of the fiber laser are also discussed

  3. Unidirectional, dual-comb lasing under multiple pulse formation mechanisms in a passively mode-locked fiber ring laser

    Science.gov (United States)

    Liu, Ya; Zhao, Xin; Hu, Guoqing; Li, Cui; Zhao, Bofeng; Zheng, Zheng

    2016-09-01

    Dual-comb lasers from which asynchronous ultrashort pulses can be simultaneously generated have recently become an interesting research subject. They could be an intriguing alternative to the current dual-laser optical-frequency-comb source with highly sophisticated electronic control systems. If generated through a common light path traveled by all pulses, the common-mode noises between the spectral lines of different pulse trains could be significantly reduced. Therefore, coherent dual-comb generation from a completely common-path, unidirectional lasing cavity would be an interesting territory to explore. In this paper, we demonstrate such a dual-comb lasing scheme based on a nanomaterial saturable absorber with additional pulse narrowing and broadening mechanisms concurrently introduced into a mode-locked fiber laser. The interactions between multiple soliton formation mechanisms result in unusual bifurcation into two-pulse states with quite different characteristics. Simultaneous oscillation of pulses with four-fold difference in pulsewidths and tens of Hz repetition rate difference is observed. The coherence between these spectral-overlapped, picosecond and femtosecond pulses is further verified by the corresponding asynchronous cross-sampling and dual-comb spectroscopy measurements.

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

    NARCIS (Netherlands)

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

    2004-01-01

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

  5. Detection of mycoplasmas in goat milk by flow cytometry.

    Science.gov (United States)

    Assunção, Patricia; Davey, Hazel M; Rosales, Ruben S; Antunes, Nuno T; de la Fe, Christian; Ramirez, Ana S; de Galarreta, Carlos M Ruiz; Poveda, Jose B

    2007-12-01

    The detection of mycoplasma in milk can be performed by either culture techniques or polymerase chain reaction (PCR) based methods. Although PCR can reduce the average diagnostic time to 5 h in comparison with the several days for the isolation of the agent, there is still a need to develop methods, which could give earlier results. For this purpose, we tested the ability of flow cytometry (FC) to detect mycoplasmas in milk samples. Milk samples inoculated with four different mycoplasmas, Mycoplasma agalactiae, Mycoplasma putrefaciens, Mycoplasma capricolum subsp. Capricolum, or Mycoplasma mycoides subsp. mycoides large-colony type, known to cause contagious agalactia in goats, were stained with the DNA stain SYBR Green I and analyzed by FC. Three goat milk samples, from which mycoplasmas have been isolated in broth medium were also analyzed. All mycoplasmas were easily distinguished from debris of milk samples, but it was not possible to distinguish between the different mycoplasma species. In our conditions, the detection limit of the technique was of the order of 10(3)-10(4) cells ml(-1). Furthermore, mycoplasmas were also distinguished from Staphylococcus aureus. FC together with SYBR Green I was able to distinguish between mycoplasma cells and debris present in milk samples and gave results in 20-30 min. This is an important first step in developing a robust, routine flow cytometric method for the detection of mycoplasmas in milk samples. (c) 2007 International Society for Analytical Cytology

  6. Development of an Immunomagnetic Separation Method for Viable Salmonella Typhimurium Detected by Flow Cytometry

    DEFF Research Database (Denmark)

    Ahmed, Shakil; Rubahn, Horst-Günter; Erdmann, Helmut

    2016-01-01

    for detection of food-related bacteria. In this study, a flow cytometry based immunomagnetic separation (IMS) method for the isolation and enrichment of Salmonella Typhimurium from liquid samples was developed and optimized. Both polyclonal and monoclonal antibodies have been used to couple with 1 micron sized...... and bacteria, immunocapture time, staining and buffering conditions for the viability assays were optimized. The capture efficiency of IMS was>98% for a range of Salmonella Typhimurium cell concentrations from 103 to 105/mL using 108/mL bead concentration. The method proved to have high (98%) specificity...

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

    Science.gov (United States)

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

    2017-01-01

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

  8. 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.

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

    NARCIS (Netherlands)

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

    2004-01-01

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

  10. Highly multiparametric analysis by mass cytometry.

    Science.gov (United States)

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

    2010-09-30

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

  11. Analysis of basophil activation by flow cytometry in pediatric house dust mite allergy.

    Science.gov (United States)

    González-Muñoz, Miguel; Villota, Julian; Moneo, Ignacio

    2008-06-01

    Detection of allergen-induced basophil activation by flow cytometry has been shown to be a useful tool for allergy diagnosis. The aim of this study was to assess the potential of this technique for the diagnosis of pediatric house dust mite allergy. Quantification of total and specific IgE and basophil activation test were performed to evaluate mite allergic (n = 24), atopic (n = 23), and non-allergic children (n = 9). Allergen-induced basophil activation was detected as a CD63-upregulation. Receiver operating characteristics (ROC) curve analysis was performed to calculate the optimal cut-off value of activated basophils discriminating mite allergic and non-allergic children. ROC curve analysis yielded a threshold value of 18% activated basophils when mite-sensitized and atopic children were studied [area under the curve (AUC) = 0.99, 95% confidence interval (CI) = 0.97-1.01, p 43 kU/l) values for Dermatophagoides pteronyssinus allergen. They also showed positive prick (wheal diameter >1.0 cm) and basophil activation (>87%) tests and high specific IgE (>100 kU/l) with shrimp allergen. Shrimp sensitization was demonstrated by high levels of Pen a 1-specific IgE (>100 kU/l). Cross-reactivity between mite and shrimp was confirmed by fluorescence enzyme immunoassay (FEIA-CAP) inhibition study in these two cases. This study demonstrated that the analysis of allergen-induced CD63 upregulation by flow cytometry is a reliable tool for diagnosis of mite allergy in pediatric patients, with sensitivity similar to routine diagnostic tests and a higher specificity. Furthermore, this method can provide additional information in case of disagreement between in vivo and in vitro test results.

  12. A Dual-Plane PIV Study of Turbulent Heat Transfer Flows

    Science.gov (United States)

    Wernet, Mark P.; Wroblewski, Adam C.; Locke, Randy J.

    2016-01-01

    Thin film cooling is a widely used technique in turbomachinery and rocket propulsion applications, where cool injection air protects a surface from hot combustion gases. The injected air typically has a different velocity and temperature from the free stream combustion flow, yielding a flow field with high turbulence and large temperature differences. These thin film cooling flows provide a good test case for evaluating computational model prediction capabilities. The goal of this work is to provide a database of flow field measurements for validating computational flow prediction models applied to turbulent heat transfer flows. In this work we describe the application of a Dual-Plane Particle Image Velocimetry (PIV) technique in a thin film cooling wind tunnel facility where the injection air stream velocity and temperatures are varied in order to provide benchmark turbulent heat transfer flow field measurements. The Dual-Plane PIV data collected include all three components of velocity and all three components of vorticity, spanning the width of the tunnel at multiple axial measurement planes.

  13. Ploidy Levels among Species in the ‘Oxalis tuberosa Alliance’ as Inferred by Flow Cytometry

    OpenAIRE

    EMSHWILLER, EVE

    2002-01-01

    The ‘Oxalis tuberosa alliance’ is a group of Andean Oxalis species allied to the Andean tuber crop O. tuberosa Molina (Oxalidaceae), commonly known as ‘oca’. As part of a larger project studying the origins of polyploidy and domestication of cultivated oca, flow cytometry was used to survey DNA ploidy levels among Bolivian and Peruvian accessions of alliance members. In addition, this study provided a first assessment of C‐values in the alliance by estimating nuclear DNA contents of these acc...

  14. Effect of acetic acid on Saccharomyces carlsbergensis ATCC 6269 batch ethanol production monitored by flow cytometry.

    Science.gov (United States)

    Freitas, Cláudia; Neves, Elisabete; Reis, Alberto; Passarinho, Paula C; da Silva, Teresa Lopes

    2012-11-01

    Bioethanol produced from lignocellulosic materials has been considered a sustainable alternative fuel. Such type of raw materials have a huge potential, but their hydrolysis into mono-sugars releases toxic compounds such as weak acids, which affect the microorganisms' physiology, inhibiting the growth and ethanol production. Acetic acid (HAc) is the most abundant weak acid in the lignocellulosic materials hydrolysates. In order to understand the physiological changes of Saccharomyces carlsbergensis when fermenting in the presence of different acetic acid (HAc) concentrations, the yeast growth was monitored by multi-parameter flow cytometry at same time that the ethanol production was assessed. The membrane potential stain DiOC(6)(3) fluorescence intensity decreased as the HAc concentration increased, which was attributed to the plasmic membrane potential reduction as a result of the toxic effect of the HAc undissociated form. Nevertheless, the proportion of cells with permeabilized membrane did not increase with the HAc concentration increase. Fermentations ending at lower external pH and higher ethanol concentrations depicted the highest proportions of permeabilized cells and cells with increased reactive oxygen species levels. Flow cytometry allowed monitoring, near real time (at-line), the physiological states of the yeast during the fermentations. The information obtained can be used to optimize culture conditions to improve bioethanol production.

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

    Science.gov (United States)

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

    2018-02-01

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

  16. A dual-wavelength tunable laser with superimposed fiber Bragg gratings

    International Nuclear Information System (INIS)

    Álvarez-Tamayo, R I; Durán-Sánchez, M; Pottiez, O; Ibarra-Escamilla, B; Kuzin, E A; Cruz, J L; Andrés, M V

    2013-01-01

    We report a dual-wavelength tunable fiber laser. The cavity is formed by two superimposed fiber Bragg gratings (FBGs) and a temperature tunable high-birefringence fiber optical loop mirror (FOLM). FBGs with wavelengths of 1548.5 and 1538.5 nm were printed in the same section of a fiber using two different masks. The superimposed FBGs were placed on a mechanical mount that allows stretch or compression of the FBGs. As a result of the FBG strain both lines are shifted simultaneously. Dual-wavelength generation requires a fine adjustment of the cavity loss for both wavelengths. (paper)

  17. Autologous Pure Platelet-Rich Plasma Dermal Injections for Facial Skin Rejuvenation: Clinical, Instrumental, and Flow Cytometry Assessment.

    Science.gov (United States)

    Cameli, Norma; Mariano, Maria; Cordone, Iole; Abril, Elva; Masi, Serena; Foddai, Maria Laura

    2017-06-01

    Platelet-rich plasma (PRP) is an emerging treatment in dermatology recently proposed for skin rejuvenation. To evaluate the efficacy and safety of autologous pure PRP dermal injections on facial skin rejuvenation, investigating the cellularity of PRP samples. Twelve patients underwent 3 sessions of PRP injection at 1-month intervals. The clinical and instrumental outcomes were evaluated before (T0) and 1 month (T1) after the end of treatment by means of transepidermal water loss, corneometry, Cutometer, Visioscan, and Visioface. A flow cytometry characterization on PRP and peripheral blood (PB) samples was performed. Clinical and patient evaluation showed improvement of skin texture. Skin gross elasticity, skin smoothness parameters, skin barrier function, and capacitance were significantly improved. No difference between PRP and PB lymphocyte immunological asset was observed. A leukocyte population (mainly CD3) and neutrophils depletion were documented in all the PRP samples. This instrumental study demonstrated that PRP poor in leukocytes can provide objective improvements in skin biostimulation. Flow cytometry showed no variability among the PRP samples using a reproducible separation system and a low content in proinflammatory cells. Although a pilot study, it may be helpful for future investigations on PRP cellularity.

  18. Design of the Flow Plates for a Dual Cooled Fuel Assembly

    International Nuclear Information System (INIS)

    Kim, Jae Yong; Yoon, Kyung Ho; Lee, Young Ho; Lee, Kang Hee; Kim, Hyung Kyu

    2009-01-01

    In a dual cooled fuel assembly, the array and position of fuels are changed from those of a conventional PWR fuel assembly to achieve a power uprating. The flow plate provides flow holes to direct the heated coolant into/out of the fuel assembly and structural intensity to insure that the fuel rod is axially restrained within the spacer grids. So, flow plates of top/bottom end pieces (TEP/BEP) have to be modified into proper shape. Because the flow holes' area of a flow plate affects pressure drop, the flow holes' area must be larger than/equal to that of conventional flow plates. And design criterion of the TEP/BEP says that the flow plate should withstand a 22.241 kN axial load during handling lest a calculated stress intensity should exceed the Condition I allowable stress. In this paper, newly designed flow plates of a TEP/BEP are suggested and stress analysis is conducted to evaluate strength robustness of the flow plates for the dual cooled fuel assembly

  19. Formation of novel reactive intermediate by electron-laser dual beam irradiation

    International Nuclear Information System (INIS)

    Ishida, Akito; Takamuku, Setsuo

    1992-01-01

    The pulse radiolysis system of the Institute of Scientific and Industrial Research, Osaka University, (ISIR) has been progressed to observe a highly reactive species, which is produced by successive irradiation of electron and laser or of CW-UV-light and electron. The dual beam irradiation system, which consists of the beam synchronization system, the optical alignment, and the measurement system, is described in detail. Dual beam irradiation studies on 2-methylbenzophenone and some compounds with a C=N bond have been carried out by use of this system. Pulse radiolysis of 2-methylbenzophenone in benzene induced formation of an unstable photoenol via the triplet state, which was irradiated by a visible laser pulse to give dihydroanthrone. Pulse radiolysis of syn-benzalaniline and a nitrileylide in 2-methyltetrahydrofuran, which were produced by steady state photoirradiation at low temperature, enabled us to observe their very unstable radical anions. (author)

  20. Microfabrication and Test of a Three-Dimensional Polymer Hydro-focusing Unit for Flow Cytometry Applications

    Science.gov (United States)

    Yang, Ren; Feeback, Daniel L.; Wang, Wan-Jun

    2005-01-01

    This paper details a novel three-dimensional (3D) hydro-focusing micro cell sorter for micro flow cytometry applications. The unit was microfabricated by means of SU-8 3D lithography. The 3D microstructure for coaxial sheathing was designed, microfabricated, and tested. Three-dimensional hydrofocusing capability was demonstrated with an experiment to sort labeled tanned sheep erythrocytes (red blood cells). This polymer hydro-focusing microstructure is easily microfabricated and integrated with other polymer microfluidic structures. Keywords: SU-8, three-dimensional hydro-focusing, microfluidic, microchannel, cytometer

  1. Flow cytometry total cell counts : A field study assessing microbiological water quality and growth in unchlorinated drinking water distribution systems

    NARCIS (Netherlands)

    Liu, G.; Van der Mark, E.J.; Verberk, J.Q.; Van Dijk, J.C.

    2013-01-01

    e objective of this study was to evaluate the application of flow cytometry total cell counts (TCCs) as a parameter to assess microbial growth in drinking water distribution systems and to determine the relationships between different parameters describing the biostability of treated water. A

  2. Single- and dual-wavelength laser pulses induced modification in 10×(Al/Ti)/Si multilayer system

    Energy Technology Data Exchange (ETDEWEB)

    Salatić, B. [University of Belgrade, Institute of Physics Belgrade, Pregrevica 118, 11080 Belgrade (Serbia); Petrović, S., E-mail: spetro@vinca.rs [University of Belgrade, Institute of Nuclear Science-Vinča, POB 522, 11001 Belgrade (Serbia); Peruško, D. [University of Belgrade, Institute of Nuclear Science-Vinča, POB 522, 11001 Belgrade (Serbia); Čekada, M.; Panjan, P. [Jožef Stefan Institute, Jamova 39, 1000 Ljubljana (Slovenia); Pantelić, D.; Jelenković, B. [University of Belgrade, Institute of Physics Belgrade, Pregrevica 118, 11080 Belgrade (Serbia)

    2016-01-01

    Graphical abstract: - Highlights: • Experimental and numerical study of laser-induced ablation and micro-sized crater formation. • Dual-wavelength pulses induce creation of wider and deeper craters due to synergies of two processes. • Sunflower-like structure formed by dual-wavelength pulses at low irradiance. • Numerical model of nanosecond pulsed laser ablation for complex (Al/Ti)/Si system has been developed. - Abstract: The surface morphology of the ablation craters created in the multilayer 10×(Al/Ti)/Si system by nanosecond laser pulses at single- and dual wavelength has been studied experimentally and numerically. A complex multilayer thin film including ten (Al/Ti) bilayers deposited by ion sputtering on Si(1 0 0) substrate to a total thickness of 260 nm were illuminated at different laser irradiance in the range 0.25–3.5 × 10{sup 9} W cm{sup −2}. Single pulse laser irradiation was done at normal incidence in air, with the single wavelength, either at 532 nm or 1064 nm or with both laser light simultaneously in the ratio of 1:10 for energy per pulse between second harmonic and 1064 nm. Most of the absorbed laser energy was rapidly transformed into heat, producing intensive modifications of composition and morphology on the sample surface. The results show an increase in surface roughness, formation of specific nanostructures, appearance of hydrodynamic features and ablation of surface material with crater formation. Applying a small fraction (10%) of the second harmonic in dual-wavelength pulses, a modification of the 10×(Al/Ti)/Si system by a single laser pulse was reflected in the formation of wider and/or deeper craters. Numerical calculations show that the main physical mechanism in ablation process is normal evaporation without phase explosion. The calculated and experimental results agree relatively well for the whole irradiance range, what makes the model applicable to complex Al/Ti multilayer systems.

  3. Measurement of Quasi-periodic Oscillating Flow Motion in Simulated Dual-cooled Annular Fuel Bundle

    International Nuclear Information System (INIS)

    Lee, Chi Young; Shin, Chang Hwan; Park, Ju Yong; Oh, Dong Seok; Chun, Tae Hyun; In, Wang Kee

    2012-01-01

    In order to increase a significant amount of reactor power in OPR1000, KAERI (Korea Atomic Energy Research Institute) has been developing a dual-cooled annular fuel. The dual-cooled annular fuel is simultaneously cooled by the water flow through the inner and the outer channels. KAERI proposed the 12x12 dual-cooled annular fuel array which was designed to be structurally compatible with the 16x16 cylindrical solid fuel array by maintaining the same array size and the guide tubes in the same locations, as shown in Fig. 1. In such a case, due to larger outer diameter of dual-cooled annular fuel than conventional solid fuel, a P/D (Pitch-to-Diameter ratio) of dual cooled annular fuel assembly becomes smaller than that of cylindrical solid fuel. A change in P/D of fuel bundle can cause a difference in the flow mixing phenomena between the dual-cooled annular and conventional cylindrical solid fuel assemblies. In this study, the rod bundle flow motion appearing in a small P/D case is investigated preliminarily using PIV (Particle Image Velocimetry) for dual-cooled annular fuel application

  4. Distinction between asymptomatic monoclonal B-cell lymphocytosis with cyclin D1 overexpression and mantle cell lymphoma: from molecular profiling to flow cytometry.

    Science.gov (United States)

    Espinet, Blanca; Ferrer, Ana; Bellosillo, Beatriz; Nonell, Lara; Salar, Antonio; Fernández-Rodríguez, Concepción; Puigdecanet, Eulàlia; Gimeno, Javier; Garcia-Garcia, Mar; Vela, Maria Carmen; Luño, Elisa; Collado, Rosa; Navarro, José Tomás; de la Banda, Esmeralda; Abrisqueta, Pau; Arenillas, Leonor; Serrano, Cristina; Lloreta, Josep; Miñana, Belén; Cerutti, Andrea; Florensa, Lourdes; Orfao, Alberto; Sanz, Ferran; Solé, Francesc; Dominguez-Sola, David; Serrano, Sergio

    2014-02-15

    According to current diagnostic criteria, mantle cell lymphoma (MCL) encompasses the usual, aggressive variants and rare, nonnodal cases with monoclonal asymptomatic lymphocytosis, cyclin D1-positive (MALD1). We aimed to understand the biology behind this clinical heterogeneity and to identify markers for adequate identification of MALD1 cases. We compared 17 typical MCL cases with a homogeneous group of 13 untreated MALD1 cases (median follow-up, 71 months). We conducted gene expression profiling with functional analysis in five MCL and five MALD1. Results were validated in 12 MCL and 8 MALD1 additional cases by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and in 24 MCL and 13 MALD1 cases by flow cytometry. Classification and regression trees strategy was used to generate an algorithm based on CD38 and CD200 expression by flow cytometry. We found 171 differentially expressed genes with enrichment of neoplastic behavior and cell proliferation signatures in MCL. Conversely, MALD1 was enriched in gene sets related to immune activation and inflammatory responses. CD38 and CD200 were differentially expressed between MCL and MALD1 and confirmed by flow cytometry (median CD38, 89% vs. 14%; median CD200, 0% vs. 24%, respectively). Assessment of both proteins allowed classifying 85% (11 of 13) of MALD1 cases whereas 15% remained unclassified. SOX11 expression by qRT-PCR was significantly different between MCL and MALD1 groups but did not improve the classification. We show for the first time that MALD1, in contrast to MCL, is characterized by immune activation and driven by inflammatory cues. Assessment of CD38/CD200 by flow cytometry is useful to distinguish most cases of MALD1 from MCL in the clinical setting. MALD1 should be identified and segregated from the current MCL category to avoid overdiagnosis and unnecessary treatment. ©2013 AACR

  5. Enumeration of antigen-specific CD8+ T lymphocytes by single-platform, HLA tetramer-based flow cytometry: a European multicenter evaluation.

    NARCIS (Netherlands)

    Heijnen, I.; Barnett, D.; Arroz, M.J.; Barry, S.M.; Bonneville, M.; Brando, B.; D'Hautcourt, J.L.; Kern, F.; Totterman, T.H.; Marijt, E.W.; Bossy, D.; Preijers, F.W.M.B.; Rothe, G.; Gratama, J.W.

    2004-01-01

    BACKGROUND: HLA class I peptide tetramers represent powerful diagnostic tools for detection and monitoring of antigen-specific CD8(+) T cells. The impetus for the current multicenter study is the critical need to standardize tetramer flow cytometry if it is to be implemented as a routine diagnostic

  6. Use of a Whole-Cell Biosensor and Flow Cytometry to Detect AHL Production by an Indigenous Soil Community During Decomposition

    DEFF Research Database (Denmark)

    Burmølle, Mette; Hansen, Lars Hestbjerg; Sørensen, Søren Johannes

    2005-01-01

    originating from Vibrio fischeri. This resulted in a whole-cell biosensor, responding to the presence of AHL compounds. The biosensor was introduced to compost soil microcosms amended with nettle leaves. After 3 days of incubation, cells were extracted and analyzed by flow cytometry. All microcosms contained...

  7. High-power dual-wavelength external-Cavity diode laser based on tapered amplifier with tunable terahertz frequency difference

    DEFF Research Database (Denmark)

    Chi, Mingjun; Jensen, Ole Bjarlin; Petersen, Paul Michael

    2011-01-01

    Tunable dual-wavelength operation of a diode laser system based on a tapered diode amplifier with double-Littrow external-cavity feedback is demonstrated around 800nm. The two wavelengths can be tuned individually, and the frequency difference of the two wavelengths is tunable from 0.5 to 5:0 THz......, this is the highest output power from a dual-wavelength diode laser system operating with tunable terahertz frequency difference. © 2011 Optical Society of America....

  8. A dual far-infrared laser diagnostic of magnetized plasmas

    International Nuclear Information System (INIS)

    Darrow, D.S.; Park, H.K.

    1988-02-01

    A dual far-infrared laser has been constructed and its properties have been exploited to probe tokamak-like discharges in the CDX toroidal device. Thermal variation of the difference frequency between the two far-infrared cavities is slow, though the cavities lack thermal stabilization, simply because their assembly on the same chassis exposes them to virtually identical temperature changes. The optical arrangement beyond the laser permits conversion within minutes between interferometry and density fluctuation observation, and within an hour between different operating wavelengths. Line-average densities of 2 /times/ 10 13 cm -3 and coherent fluctuations in the neighborhood of 20 kHz have been measured with this diagnostic. 15 refs., 5 figs., 2 tabs

  9. New mononuclear leukocyte-like populations within the granulocyte scatter gate detected by flow cytometry (Conference Presentation)

    Science.gov (United States)

    Melzer, Susanne; Löffler, Markus; Kautzner, Marlene; Tárnok, Attila

    2017-02-01

    Granulocytes are the major players in innate immunity and are prognostic markers in diseases. An in-depth phenotypic characterization of granulocyte subtypes and correlation with biometry or lifestyle is so far lacking. The reason is, that either preparation of mononuclear cells was analyzed or that cells in the neutrophil window were neglected in the analysis. Here we show for the first time lymphocyte- (LL) and monocyte-like (ML) cells within the granulocyte scatter gate as new, previously unknown cell subpopulation. Immunophenotyping of 905 healthy German adults from the LIFE study [1] was performed by 10-color flow cytometry [2]. Age of men (n=420): 56.5±14.0 years, women (n=485): 56.7±13.6 y (range of 18-81 y). Data analyzed by FlowJo v10.0.6. Values compared by Mann-Whitney-U test: men vs women, young (18-49 y) vs. elderly (50-81 y.) men, and young (19-49 y.) vs. elderly (50-81 y.) women; significance: page, except LL2 in women. In conclusion, new lymphocyte like cell types with the neutrophil scatter characteristics are reported. Counts correlate with age and gender. We plan to sort these new subtypes for further functional characterization and aim to establish them as cellular biomarkers for the early detection of various diseases. [1] BMC Public Health. 2015;15:691; [2] Cytometry A. 2014;85(9):781

  10. Development by flow cytometry of bioassays based on chlorella for environmental monitoring

    Directory of Open Access Journals (Sweden)

    Petrescu C-M,

    2016-05-01

    Full Text Available In ecotoxicological assessments, bioassays (ecotoxicity tests or biotests are one of the main tools, defined as methods which use living cells, tissues, organism or communities to assess exposure-related effects of chemicals. The increasing complexity of environmental degradation requires an increase in the capacity of scientific approach in monitoring and notification as early as possible risks. Our own objective concerns the detection of aquatic environment pollution in Romania and particularly in the Danube basin. For assessing aquatic environment pollution degree or for assessing cytotoxicity or ecotoxicity of pollutants (heavy metals, nanoparticles, pesticides, etc. we developed news experimental bioassays based on the use of viability and apoptosis biomarkers of Chlorella cells by flow cytometry. Our proposed bioassays could be rapid and very sensitive tests for in laboratory aquatic risk assessment and biomonitoring.

  11. Flow cytometry protocol to evaluate ionizing radiation effects on P-glycoprotein activity

    International Nuclear Information System (INIS)

    Santos, Neyliane Goncalves dos; Amaral, Ademir; Cavalcanti, Mariana Brayner . E-mail; Neves, Maria Amelia Batista; Machado, Cintia Gonsalves de Faria

    2008-01-01

    The aim of this work was to establish a protocol to evaluate ionizing radiation effects on P-glycoprotein (P-gp) activity. For this, human peripheral blood samples were irradiated in vitro with different doses and P-gp activity was analyzed for CD4 and CD8 T lymphocytes through rhodamine123-efflux assay by flow cytometry. By simultaneous employment of percentage and mean fluorescence index parameters, subject-by-subject analysis pointed out changes in P-gp activity for some individuals and irradiated samples. Based on this work, the proposed protocol was considered adequate for evaluating P-gp activity on cells after radioactive stress. Besides, this research suggests that P-gp activity could be an important factor to define patient-specific protocols in combined chemo- and radiotherapy, particularly when radiation exposure precedes chemical treatment. (author)

  12. 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.

  13. 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.)

  14. Flow cytometry protocol to evaluate ionizing radiation effects on P-glycoprotein activity

    Energy Technology Data Exchange (ETDEWEB)

    Santos, Neyliane Goncalves dos; Amaral, Ademir; Cavalcanti, Mariana Brayner [Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil). Dept. de Energia Nuclear]. E-mail; neylisantos@yahoo.com.br; Neves, Maria Amelia Batista; Machado, Cintia Gonsalves de Faria [Fundacao de Hematologia e Hemoterapia de Pernambuco, Recife, PE (Brazil). Unidade de Laboratorios Especializados. Lab. de Imunofenotipagem

    2008-12-15

    The aim of this work was to establish a protocol to evaluate ionizing radiation effects on P-glycoprotein (P-gp) activity. For this, human peripheral blood samples were irradiated in vitro with different doses and P-gp activity was analyzed for CD4 and CD8 T lymphocytes through rhodamine123-efflux assay by flow cytometry. By simultaneous employment of percentage and mean fluorescence index parameters, subject-by-subject analysis pointed out changes in P-gp activity for some individuals and irradiated samples. Based on this work, the proposed protocol was considered adequate for evaluating P-gp activity on cells after radioactive stress. Besides, this research suggests that P-gp activity could be an important factor to define patient-specific protocols in combined chemo- and radiotherapy, particularly when radiation exposure precedes chemical treatment. (author)

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

    Science.gov (United States)

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

    2018-02-01

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

  16. Optimization of CO2 Laser Cutting Process using Taguchi and Dual Response Surface Methodology

    Directory of Open Access Journals (Sweden)

    M. Madić

    2014-09-01

    Full Text Available Selection of optimal cutting parameter settings for obtaining high cut quality in CO2 laser cutting process is of great importance. Among various analytical and experimental optimization methods, the application of Taguchi and response surface methodology is one of most commonly used for laser cutting process optimization. Although the concept of dual response surface methodology for process optimization has been used with success, till date, no experimental study has been reported in the field of laser cutting. In this paper an approach for optimization of CO2 laser cutting process using Taguchi and dual response surface methodology is presented. The goal was to determine the near optimal laser cutting parameter values in order to ensure robust condition for minimization of average surface roughness. To obtain experimental database for development of response surface models, Taguchi’s L25 orthogonal array was implemented for experimental plan. Three cutting parameters, the cutting speed (3, 4, 5, 6, 7 m/min, the laser power (0.7, 0.9, 1.1, 1.3, 1.5 kW, and the assist gas pressure (3, 4, 5, 6, 7 bar, were used in the experiment. To obtain near optimal cutting parameters settings, multi-stage Monte Carlo simulation procedure was performed on the developed response surface models.

  17. Applications of flow cytometry in plant pathology for genome size determination, detection and physiological status.

    Science.gov (United States)

    D'Hondt, Liesbet; Höfte, Monica; Van Bockstaele, Erik; Leus, Leen

    2011-10-01

    Flow cytometers are probably the most multipurpose laboratory devices available. They can analyse a vast and very diverse range of cell parameters. This technique has left its mark on cancer, human immunodeficiency virus and immunology research, and is indispensable in routine clinical diagnostics. Flow cytometry (FCM) is also a well-known tool for the detection and physiological status assessment of microorganisms in drinking water, marine environments, food and fermentation processes. However, flow cytometers are seldom used in plant pathology, despite FCM's major advantages as both a detection method and a research tool. Potential uses of FCM include the characterization of genome sizes of fungal and oomycete populations, multiplexed pathogen detection and the monitoring of the viability, culturability and gene expression of plant pathogens, and many others. This review provides an overview of the history, advantages and disadvantages of FCM, and focuses on the current applications and future possibilities of FCM in plant pathology. © 2011 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2011 BSPP AND BLACKWELL PUBLISHING LTD.

  18. A comparative evaluation of Eosin-5'-maleimide flow cytometry reveals a high diagnostic efficacy for hereditary spherocytosis.

    Science.gov (United States)

    Joshi, P; Aggarwal, A; Jamwal, M; Sachdeva, M U S; Bansal, D; Malhotra, P; Sharma, P; Das, R

    2016-10-01

    Laboratory diagnosis of hereditary spherocytosis (HS) relies on increased incubated red cell osmotic fragility test for screening. We evaluated the diagnostic role of eosin-5'-maleimide (EMA) binding test by flow cytometry in spherocytic and microcytic hypochromic hematological disorders in North Indians. EMA binding test using flow cytometry was performed on 55 HS (40 families), 26 iron deficiency anemia (IDA), 32 β-thalassemia trait (βTT), and 10 autoimmune hemolytic anemia (AIHA) cases and 121 normals. Mean channel fluorescence (MCF) and coefficient of variation (CV) were studied. Different MCF parameters (MCF, MCF ratio, percent decrease MCF) and percent increase in CV were analyzed. Receiver operating characteristics analysis was performed to determine best cutoff values, sensitivity, and specificity for discriminating HS from other red cell disorders. MCF ratio of HS group was significantly lower than normals (0.67 ± 0.07 vs. 1.01 ± 0.05, P < 0.001) and other cases. All patients with HS showed MCF ratio to be ≤0.79. Four postsplenectomy cases with near-normal hemograms also revealed low MCF ratio, showing the specificity of the test. EMA assay was efficient to diagnose cases of HS including postsplenectomy cases and shows no overlap with IDA, βTT, and AIHA. © 2016 John Wiley & Sons Ltd.

  19. A UV pre-ionized dual-wavelength short-pulse high-power CO{sub 2} laser facility for laser particle acceleration research

    Energy Technology Data Exchange (ETDEWEB)

    Ebrahim, N A; Mouris, J F; Davis, R W

    1994-12-01

    In this report we describe the Chalk River dual-wavelength, short-pulse, single-mode, high-power CO{sub 2} laser facility for research in laser particle acceleration and CANDU materials modifications. The facility is designed and built around UV-preionized transversely-excited atmospheric-pressure (TEA) Lumonics CO{sub 2} laser discharge modules. Peak focussed power densities of up to 2 x 10{sup 14} W/cm{sup 2} in 500 ps pulses have been obtained. (author). 10 refs., 9 figs.

  20. Analysis of dual-mode lasing characteristics in a 1310-nm optically injected quantum dot distributed feedback laser

    Science.gov (United States)

    Raghunathan, R.; Olinger, J.; Hurtado, A.; Grillot, F.; Kovanis, V.; Lester, L. F.

    2015-03-01

    Recent work has shown the Quantum Dot (QD) material system to be well-suited to support dual-mode lasing. In particular, optical injection from a master laser (ML) into the residual Fabry-Perot (FP) modes of a 1310 nm Quantum Dot Distributed Feedback (QD-DFB) laser has been recently demonstrated to offer a highly reliable platform for stable dual-mode lasing operation. External controls on the ML, such as operating temperature and bias current, can be used to precisely adjust the spacing between the two lasing modes. This tunability of modeseparation is very promising for a range of applications requiring the generation of microwave, millimeter wave and terahertz signals. Considering the versatility and utility of such a scheme, it is imperative to acquire a deeper understanding of the factors that influence the dual-mode lasing process, in order to optimize performance. Toward this end, this paper seeks to further our understanding of the optically-injected dual-mode lasing mechanism. For fixed values of optical power injected into each FP residual mode and wavelength detuning, the dual-mode lasing characteristics are analyzed with regard to important system parameters such as the position and the intensity of the injected residual mode (relative to the Bragg and the other residual FP modes of the device) for two similarly-fabricated QD-DFBs. Results indicate that for dual mode lasing spaced less than 5 nm apart, the relative intensity of the injected FP mode and intracavity noise levels are critical factors in determining dual mode lasing behavior. Insight into the dual-mode lasing characteristics could provide an important design guideline for the master and QD-DFB slave laser cavities.

  1. Femtosecond laser ablation of polymethylmethacrylate via dual-color synthesized waveform

    International Nuclear Information System (INIS)

    Yang, Chan-Shan; Zaytsev, Alexey; Lin, Chih-Hsuan; Teng, Kuei-Chung; Her, Tsing-Hua; Pan, Ci-Ling

    2015-01-01

    We have demonstrated the laser ablation of PMMA using dual-color waveform synthesis of the fundamental (ω) and its second-harmonic (2ω) of a femtosecond Ti:Sapphire laser. A modest and yet clear modulation (∼22%) in ablated area versus relative phase between the 2ω and ω beams with a power-ratio of 15% (28/183 mW) is revealed. This is explained qualitatively by the dependence of ablation on multiphoton ionization of which the rate is related to the relative phase of the synthesized waveform. At higher peak power ratios, the modulation decreases rapidly, as the two-photon-ionization rate of the 2ω dominates over that of the three- to four- photon ionization of the ω beam. This technique demonstrates the feasibility of phase-controlled laser processing of materials

  2. Contribution of flow cytometry to the diagnosis of gastric lymphomas in endoscopic biopsy specimens.

    Science.gov (United States)

    Almasri, N M; Zaer, F S; Iturraspe, J A; Braylan, R C

    1997-07-01

    Gastric lymphomas seem to have unique clinical, pathologic, and immunophenotypic features that set them apart from nodal lymphomas. Microscopic examination of endoscopic biopsy specimens is the most frequent procedure used to diagnose gastric tumors, but it is very difficult, and sometimes impossible, to recognize lymphomas in endoscopic samples by histologic or even immunohistologic methods. Because most gastric lymphomas are of B-cell origin, we used flow cytometry to assess B-cell clonality in gastric biopsy specimens containing dense lymphocytic infiltrates thought to represent lymphoma. We prepared viable cell suspensions from unfixed specimens obtained from 29 consecutive patients who had a previous microscopic diagnosis of suspicious gastric lymphoid infiltrates. We performed immunophenotypic studies with multicolor flow cytometry, and we assessed clonality by examination of immunoglobulin (Ig) light-chain expression analyzed exclusively on B cells identified by anti-CD20 or CD19 antibodies. The mean number of cells recovered was 1.04 x 10(6), from an average of 5.5 gastric biopsy fragments per patient. In 26 of the 29 patients, the number of cells was adequate for analysis. We detected B-cell monoclonality in 16 cases, including 5 in which the percentage of clonal B cells was less than 5%. Of the 16 cases, only 8 could be diagnosed as lymphomas on morphologic grounds alone; the remaining 8 patients had either suspicious lymphoid infiltrates or chronic gastritis. The three cases with an insufficient number of cells were considered non-neoplastic either on histologic grounds alone or in conjunction with Southern analysis of Ig genes. We conclude that flow cytometric immunophenotypic analysis of freshly prepared cell suspensions obtained from endoscopic biopsy specimens can be used to evaluate gastric lymphocytic infiltrates. Specifically, the analysis of surface Ig light-chain expression on B cells distinguishes between monoclonal (lymphoma) and polyclonal

  3. Tunable single and dual mode operation of an external cavity quantum-dot injection laser

    International Nuclear Information System (INIS)

    Biebersdorf, A; Lingk, C; De Giorgi, M; Feldmann, J; Sacher, J; Arzberger, M; Ulbrich, C; Boehm, G; Amann, M-C; Abstreiter, G

    2003-01-01

    We investigate quantum-dot (QD) lasers in an external cavity using Littrow and Littman configurations. Here, we report on a continuously tunable QD laser with a broad tuning range from 1047 to 1130 nm with high stability and efficient side mode suppression. The full-width at half-maximum of the laser line is 0.85 nm determined mainly by the quality of the external grating. This laser can be operated in a dual-mode modus, where the mode-spacing can be tuned continuously between 1.1 and 34 nm. Simultaneous emission of the two laser modes is shown by sum frequency generation experiments

  4. Highly stable microwave carrier generation using a dual-frequency distributed feedback laser

    NARCIS (Netherlands)

    Khan, M.R.H.; Bernhardi, Edward; Marpaung, D.A.I.; Burla, M.; de Ridder, R.M.; Worhoff, Kerstin; Pollnau, Markus; Roeloffzen, C.G.H.

    2012-01-01

    Photonic generation of microwave carriers by using a dual-frequency distributed feedback waveguide laser in ytterbium-doped aluminum oxide is demonstrated. A highperformance optical frequency locked loop is implemented to stabilize the microwave carrier. This approach results in a microwave

  5. Rapid identification of Staphylococcus aureus and methicillin resistance by flow cytometry using a peptide nucleic acid probe.

    Science.gov (United States)

    Shrestha, Nabin K; Scalera, Nikole M; Wilson, Deborah A; Brehm-Stecher, Byron; Procop, Gary W

    2011-09-01

    A total of 56 Staphylococcus aureus isolates incubated for 2 h in the presence or absence of oxacillin were analyzed by flow cytometry after labeling with an S. aureus-specific peptide nucleic acid (PNA) probe. Two defined ratios, the paired signal count ratio (PSCR) and the gate signal count ratio (GSCR), differentiated methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) with sensitivities of 100% each and specificities of 96% and 100%, respectively.

  6. A rapid, reliable method of evaluating growth and viability of intraerythrocytic protozoan hemoparasites using fluorescence flow cytometry

    OpenAIRE

    Davis,W. C.; Wyatt,C. R.; Hamilton,M. J.; Goff,W. L.

    1992-01-01

    Fluorescence flow cytometry was employed to assess the potential of a vital dye, hydroethiedine, for use in the detection and monitoring of the viability of hemoparasites in infected erythrocytes, using Babesia bovis as a model parasite. The studies demonstrated that hydroethidine is taken up by B. bovis and metabolically converted to the DNA binding fluorochrone, ethidium. Following uptake of the dye, erythrocytes contamine viable parasites were readily distinguished and quantitated. Timed s...

  7. Broadband multi-wavelength Brillouin lasers with an operating wavelength range of 1500–1600 nm generated by four-wave mixing in a dual wavelength Brillouin fiber laser cavity

    Science.gov (United States)

    Li, Q.; Jia, Z. X.; Weng, H. Z.; Li, Z. R.; Yang, Y. D.; Xiao, J. L.; Chen, S. W.; Huang, Y. Z.; Qin, W. P.; Qin, G. S.

    2018-05-01

    We demonstrate broadband multi-wavelength Brillouin lasers with an operating wavelength range of 1500–1600 nm and a frequency separation of ~9.28 GHz generated by four-wave mixing in a dual wavelength Brillouin fiber laser cavity. By using one continuous-wave laser as the pump source, multi-wavelength Brillouin lasers with an operating wavelength range of 1554–1574 nm were generated via cascaded Brillouin scattering and four-wave mixing. Interestingly, when pumped by two continuous-wave lasers with an appropriate frequency separation, the operating wavelength range of the multi-wavelength Brillouin lasers was increased to 1500–1600 nm due to cavity-enhanced cascaded four-wave mixing among the frequency components generated by two pump lasers in the dual wavelength Brillouin laser cavity.

  8. Use of flow cytometry to identify monoclonal antibodies that recognize conserved epitopes on orthologous leukocyte differentiation antigens in goats, llamas, and rabbits

    Czech Academy of Sciences Publication Activity Database

    Davis, W. C.; Drbal, Karel; El-Aziz, A.; Mosaad, A.E.; Elbagory, A.R.M.; TIbary, A.; Barrington, G.M.; Park, Y.H.; Hamilton, M.J.

    2007-01-01

    Roč. 119, 1-2 (2007), s. 123-130 ISSN 0165-2427 Institutional research plan: CEZ:AV0Z50520514 Keywords : flow cytometry * monoclonal antibodies * leukocytes Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.957, year: 2007

  9. Usefullness of IGH/TCR PCR studies in lymphoproliferative disorders with inconclusive clonality by flow cytometry.

    Science.gov (United States)

    Ribera, Jordi; Zamora, Lurdes; Juncà, Jordi; Rodríguez, Inés; Marcé, Silvia; Cabezón, Marta; Millá, Fuensanta

    2013-07-25

    In up to 5-15% of studies of lymphoproliferative disorders (LPD) flow cytometry (FCM) or immunomorphologic methods cannot discriminate malignant from reactive processes. The aim of this work was to determine the usefulness of PCR for solving these diagnostic uncertainties. We analyzed IGH and TCRγ genes by PCR in 106 samples with inconclusive FCM results. A clonal result was registered in 36/106 studies, with a LPD being confirmed in 27 (75%) of these cases. Specifically, 9/9 IGH clonal and 16/25 TCRγ clonal results were finally diagnosed with LPD. Additionally, 2 clonal TCRγ samples with suspicion of undefined LPD were finally diagnosed with T LPD. Although polyclonal results were obtained in 47 of the cases studied (38 IGH and 9 TCRγ), hematologic neoplasms were diagnosed in 4/38 IGH polyclonal and in 1/9 TCRγ polyclonal studies. There were also 14 PCR polyclonal results (4 IGH, 10 TCRγ), albeit non-conclusive. Of these, 2/4 were eventually diagnosed with B-cell lymphoma and 3/10 with T-cell LPD. In 8 IGH samples the results of PCR techniques were non-informative but in 3/8 cases a B lymphoma was finally confirmed. We concluded that PCR is a useful technique to identify LPD when FCM is inconclusive. A PCR clonal B result is indicative of malignancy but IGH polyclonal and non-conclusive results do not exclude lymphoid neoplasms. Interpretation of T-cell clonality should be based on all the available clinical and analytical data. © 2013 Clinical Cytometry Society. Copyright © 2013 Clinical Cytometry Society.

  10. Detecting Lactococcus lactis Prophages by Mitomycin C-Mediated Induction Coupled to Flow Cytometry Analysis

    Directory of Open Access Journals (Sweden)

    Joana Oliveira

    2017-07-01

    Full Text Available Most analyzed Lactococcus lactis strains are predicted to harbor one or more prophage genomes within their chromosome; however, the true extent of the inducibility and functionality of such prophages cannot easily be deduced from sequence analysis alone. Chemical treatment of lysogenic strains with Mitomycin C is known to cause induction of temperate phages, though it is not always easy to clearly identify a lysogenic strain or to measure the number of released phage particles. Here, we report the application of flow cytometry as a reliable tool for the detection and enumeration of released lactococcal prophages using the green dye SYTO-9.

  11. Strong optical feedback in birefringent dual frequency laser

    Institute of Scientific and Technical Information of China (English)

    Mao Wei; Zhang Shu-Lian

    2006-01-01

    Strong optical feedback in a birefringent dual frequency He-Ne laser with a high reflectivity feedback mirror has been investigated for the first time. The output characteristics of two orthogonally polarized modes are demonstrated in two different optical feedback cases: one is for both modes being fed back and the other is for only one of the modes being fed back. Strong mode competition can be observed between the two modes with strong optical feedback. And when one mode's intensity is near its maximum, the other mode is nearly extinguished. When both modes are fed back into the laser cavity, the mode competition is stronger than when only one mode is fed back. The difference in initial intensity between the two orthogonally polarized modes plays an important role in the mode competition, which has been experimentally and theoretically demonstrated.

  12. Prostate extracellular vesicles in patient plasma as a liquid biopsy platform for prostate cancer using nanoscale flow cytometry

    Science.gov (United States)

    Al-Zahrani, Ali A.; Pardhan, Siddika; Brett, Sabine I.; Guo, Qiu Q.; Yang, Jun; Wolf, Philipp; Power, Nicholas E.; Durfee, Paul N.; MacMillan, Connor D.; Townson, Jason L.; Brinker, Jeffrey C.; Fleshner, Neil E.; Izawa, Jonathan I.; Chambers, Ann F.; Chin, Joseph L.; Leong, Hon S.

    2016-01-01

    Background Extracellular vesicles released by prostate cancer present in seminal fluid, urine, and blood may represent a non-invasive means to identify and prioritize patients with intermediate risk and high risk of prostate cancer. We hypothesize that enumeration of circulating prostate microparticles (PMPs), a type of extracellular vesicle (EV), can identify patients with Gleason Score≥4+4 prostate cancer (PCa) in a manner independent of PSA. Patients and Methods Plasmas from healthy volunteers, benign prostatic hyperplasia patients, and PCa patients with various Gleason score patterns were analyzed for PMPs. We used nanoscale flow cytometry to enumerate PMPs which were defined as submicron events (100-1000nm) immunoreactive to anti-PSMA mAb when compared to isotype control labeled samples. Levels of PMPs (counts/μL of plasma) were also compared to CellSearch CTC Subclasses in various PCa metastatic disease subtypes (treatment naïve, castration resistant prostate cancer) and in serially collected plasma sets from patients undergoing radical prostatectomy. Results PMP levels in plasma as enumerated by nanoscale flow cytometry are effective in distinguishing PCa patients with Gleason Score≥8 disease, a high-risk prognostic factor, from patients with Gleason Score≤7 PCa, which carries an intermediate risk of PCa recurrence. PMP levels were independent of PSA and significantly decreased after surgical resection of the prostate, demonstrating its prognostic potential for clinical follow-up. CTC subclasses did not decrease after prostatectomy and were not effective in distinguishing localized PCa patients from metastatic PCa patients. Conclusions PMP enumeration was able to identify patients with Gleason Score ≥8 PCa but not patients with Gleason Score 4+3 PCa, but offers greater confidence than CTC counts in identifying patients with metastatic prostate cancer. CTC Subclass analysis was also not effective for post-prostatectomy follow up and for

  13. Comparison of Tomo-PIV Versus Dual Plane PIV on a Synthetic Jet Flow

    Science.gov (United States)

    Wernet, Mark P.

    2017-01-01

    Particle Imaging Velocimetry (PIV) is a planar velocity measurement technique that has found widespread use across a wide class of engineering disciplines. Tomographic PIV (tomoPIV) is an extension of the traditional PIV technique whereby the velocity across a volume of fluid is measured. TomoPIV provides additional fluid mechanical properties of the flow due to the adjacent planes of velocity information that are extracted. Dual Plane PIV is another approach for providing cross-plane flow field properties. Dual Plane PIV and tomoPIV provide all of the same flow properties, albeit through very different routes with significantly different levels of effort, hence a comparison of their application and performance would prove beneficial in a well-known, highly three dimensional flow field. A synthetic jet flow which has a wide range of flow field features including high velocity gradients and regions of high vorticity was used as a rigorous test bed to determine the capabilities limitations of the Dual Plane PIV and tomoPIV techniques. The results show that compressing 3D particle field information down to a limited number of views does not permit the accurate reconstruction of the flow field. The traditional thin sheet techniques are the best approach for accurate flow field measurements.

  14. Comparative analysis of minimal residual disease detection using four-color flow cytometry, consensus IgH-PCR, and quantitative IgH PCR in CLL after allogeneic and autologous stem cell transplantation.

    Science.gov (United States)

    Böttcher, S; Ritgen, M; Pott, C; Brüggemann, M; Raff, T; Stilgenbauer, S; Döhner, H; Dreger, P; Kneba, M

    2004-10-01

    The clinically most suitable method for minimal residual disease (MRD) detection in chronic lymphocytic leukemia is still controversial. We prospectively compared MRD assessment in 158 blood samples of 74 patients with CLL after stem cell transplantation (SCT) using four-color flow cytometry (MRD flow) in parallel with consensus IgH-PCR and ASO IgH real-time PCR (ASO IgH RQ-PCR). In 25 out of 106 samples (23.6%) with a polyclonal consensus IgH-PCR pattern, MRD flow still detected CLL cells, proving higher sensitivity of flow cytometry over PCR-genescanning with consensus IgH-primers. Of 92 samples, 14 (15.2%) analyzed in parallel by MRD flow and by ASO IgH RQ-PCR were negative by our flow cytometric assay but positive by PCR, thus demonstrating superior sensitivity of RQ-PCR with ASO primers. Quantitative MRD levels measured by both methods correlated well (r=0.93). MRD detection by flow and ASO IgH RQ-PCR were equally suitable to monitor MRD kinetics after allogeneic SCT, but the PCR method detected impending relapses after autologous SCT earlier. An analysis of factors that influence sensitivity and specificity of flow cytometry for MRD detection allowed to devise further improvements of this technique.

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

    Science.gov (United States)

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

    2018-02-01

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

  16. Dual-wavelength high-power diode laser system based on an external-cavity tapered amplifier with tunable frequency difference

    DEFF Research Database (Denmark)

    Chi, Mingjun; Jensen, Ole Bjarlin; Petersen, Paul Michael

    2012-01-01

    knowledge, this is the broadest tuning range of the frequency difference from a dual-wavelength diode laser system. The spectrum, output power, and beam quality of the diode laser system are characterized. The power stability of each wavelength is measured, and the power fluctuations of the two wavelengths......A dual-wavelength high-power semiconductor laser system based on a tapered amplifier with double-Littrow external cavity is demonstrated around 800 nm. The two wavelengths can be tuned individually, and the frequency difference of the two wavelengths is tunable from 0.5 to 10.0 THz. To our...

  17. CytometryML with DICOM and FCS

    Science.gov (United States)

    Leif, Robert C.

    2018-02-01

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

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

    Science.gov (United States)

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

    2014-06-10

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

  19. Flow cytometry detection of planktonic cells with polycyclic aromatic hydrocarbons sorbed to cell surfaces

    KAUST Repository

    Cerezo, Maria I.

    2017-02-17

    Polycyclic aromatic hydrocarbons are very important components of oil pollution. These pollutants tend to sorb to cell surfaces, exerting toxic effects on organisms. Our study developed a flow cytometric method for the detection of PAHs sorbed to phytoplankton by exploiting their spectral characteristics. We discriminated between cells with PAHs from cells free of PAHs. Clear discrimination was observed with flow cytometer provided with 375 or 405nm lasers in addition to the standard 488nm laser necessary to identify phytoplankton. Using this method, we measured the relationship between the percentages of phytoplankton organisms with PAHs, with the decrease in the growth rate. Moreover, the development of this method could be extended to facilitate the study of PAHs impact on cell cultures from a large variety of organisms.

  20. Dual-beam, second-derivative tunable diode-laser infrared spectroscopy applied to trace-gas measurement

    International Nuclear Information System (INIS)

    Tallant, D.R.; Jungst, R.G.

    1981-04-01

    A dual beam diode laser spectrometer has been constructed using off-axis reflective optics. The spectrometer can be amplitude modulated for direct absorption measurements or frequency modulated to obtain derivative spectra. The spectrometer has high throughput, is easy to operate and align, provides good dual beam compensation, and has no evidence of the interference effects that have been observed in diode laser spectrometers using refractive optics. Unpurged, using second derivative techniques, the instrument has measured 108 parts-per-million CO (10 cm absorption cell, atmospheric pressure-broadened) with good signal/noise. With the replacement of marginal instrumental components, the signal/noise should be substantially increased. This instrument was developed to monitor the evolution of decomposition gases in sealed containers of small volume at atmospheric pressure

  1. Hierarchical Bayesian mixture modelling for antigen-specific T-cell subtyping in combinatorially encoded flow cytometry studies

    DEFF Research Database (Denmark)

    Lin, Lin; Chan, Cliburn; Hadrup, Sine R

    2013-01-01

    subtype identification in this novel, general model framework, and provide a detailed example using simulated data. We then describe application to a data set from an experimental study of antigen-specific T-cell subtyping using combinatorially encoded assays in human blood samples. Summary comments...... profiling in many biological areas, traditional flow cytometry measures relative levels of abundance of marker proteins using fluorescently labeled tags that identify specific markers by a single-color. One specific and important recent development in this area is the use of combinatorial marker assays...

  2. The use of bead beating to prepare suspensions of nuclei for flow cytometry from fresh leaves, herbarium leaves, petals and pollen.

    Science.gov (United States)

    Roberts, Andy V

    2007-12-01

    "Bead beating" is commonly used to release DNA from cells for genomic studies but it was used here to prepare suspensions of plant nuclei for measurement of DNA amounts by flow cytometry. Plant material was placed in 2-ml screw-capped tubes containing beads of zirconia/silica (2.5 mm diameter) or glass (2.5 or 1.0 mm diameter) and 1 ml of lysis buffer. The tubes were mechanically shaken with an FP120 FastPrep Cell Disrupter to release intact nuclei from plant tissue by the impact of the beads. The nuclei were then stained with propidium iodide (PI) and analyzed by flow cytometry. The method was tested using fresh leaves, fresh petals and herbarium leaves of Rosa canina, leaves and pollen of R. rugosa, and fresh leaves of Petroselinum crispum, Nicotiana tabacum, and Allium cepa. Batches of 12 samples of fresh leaves were prepared, simultaneously, in 45 s by bead beating in the Cell Disrupter. In flow cytometry histograms, nuclei of fresh leaves gave G(1)/G(0) peaks with CVs of less than 3.0% and nuclei from fresh petals and herbarium leaves of R. canina, and pollen of the generative nuclei of R. rugosa gave peaks with coefficients of variation (CVs) of less than 4.0%. DNA amounts estimated from 24-month-old herbarium leaves, using P. crispum as an internal standard, were less than those of fresh leaves by a small but significant amount. Suspensions of nuclei can be prepared rapidly and conveniently from a diversity of tissues by bead beating. Exposure of laboratory workers to harmful substances in the lysis buffer is minimized. (c) 2007 International Society for Analytical Cytology

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

    Science.gov (United States)

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

    2017-04-26

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

  4. Research on the speed of light transmission in a dual-frequency laser pumped single fiber with two directions

    Science.gov (United States)

    Qiu, Wei; Liu, Jianjun; Wang, Yuda; Yang, Yujing; Gao, Yuan; Lv, Pin; Jiang, Qiuli

    2018-01-01

    In this article a general theory of the coherent population oscillation effect in an erbium-doped fiber at room temperature is presented. We use dual pumping light waves with a simplified two-level system. Thus the time delay equations can be calculated from rate equations and the transmission equation. Using numerical simulation, in the case of dual-frequency pump light waves (1480 nm and 980 nm) with two directions, we analyze the influence of the pump power ratio on the group speed of light propagation. In addition, we compare slow light propagation with a single-pumping light and slow light propagation with a dual-pumping light at room temperature. The discussion shows that a larger time delay of slow light propagation can be obtained with a dual-frequency pumping laser. Compared to previous research methods, a dual-frequency laser pumped fiber with two directions is more controllable. Moreover, we conclude that the group velocity of light can be varied by changing the pump ratio.

  5. Effects of fluorescence excitation geometry on the accuracy of DNA fragment sizing by flow cytometry

    Energy Technology Data Exchange (ETDEWEB)

    Werner, James H. [Division of Bioscience, Los Alamos National Laboratory, Mail Stop M888, Los Alamos, New Mexico 87545-0001 (United States); Larson, Erica J. [Division of Bioscience, Los Alamos National Laboratory, Mail Stop M888, Los Alamos, New Mexico 87545-0001 (United States); Goodwin, Peter M. [Division of Bioscience, Los Alamos National Laboratory, Mail Stop M888, Los Alamos, New Mexico 87545-0001 (United States); Ambrose, W. Patrick [Division of Bioscience, Los Alamos National Laboratory, Mail Stop M888, Los Alamos, New Mexico 87545-0001 (United States); Keller, Richard A. [Division of Bioscience, Los Alamos National Laboratory, Mail Stop M888, Los Alamos, New Mexico 87545-0001 (United States)

    2000-06-01

    We report on various excitation geometries used in ultrasensitive flow cytometry that yield a linear relation between the fluorescence intensity measured from individual strained DNA fragments and the lengths of the fragments (in base pairs). This linearity holds for DNA samples that exhibit a wide range of conformations. The variety of DNA conformations leads to a distribution of dipole moment orientations for the dye molecules intercalated into the DNA. It is consequently important to use an excitation geometry such that all dye molecules are detected with similar efficiency. To estimate the conformation and the extent of elongation of the strained fragments in the flow, fluorescence polarization anisotropy and autocorrelation measurements were performed. Significant extension was observed for DNA fragments under the flow conditions frequently used for DNA fragment sizing. Classical calculations of the fluorescence emission collected over a finite solid angle are in agreement with the experimental measurements and have confirmed the relative insensitivity to DNA conformation of an orthogonal excitation geometry. Furthermore, the calculations suggested a modified excitation geometry that has increased our sizing resolution. (c) 2000 Optical Society of America.

  6. Effects of fluorescence excitation geometry on the accuracy of DNA fragment sizing by flow cytometry

    International Nuclear Information System (INIS)

    Werner, James H.; Larson, Erica J.; Goodwin, Peter M.; Ambrose, W. Patrick; Keller, Richard A.

    2000-01-01

    We report on various excitation geometries used in ultrasensitive flow cytometry that yield a linear relation between the fluorescence intensity measured from individual strained DNA fragments and the lengths of the fragments (in base pairs). This linearity holds for DNA samples that exhibit a wide range of conformations. The variety of DNA conformations leads to a distribution of dipole moment orientations for the dye molecules intercalated into the DNA. It is consequently important to use an excitation geometry such that all dye molecules are detected with similar efficiency. To estimate the conformation and the extent of elongation of the strained fragments in the flow, fluorescence polarization anisotropy and autocorrelation measurements were performed. Significant extension was observed for DNA fragments under the flow conditions frequently used for DNA fragment sizing. Classical calculations of the fluorescence emission collected over a finite solid angle are in agreement with the experimental measurements and have confirmed the relative insensitivity to DNA conformation of an orthogonal excitation geometry. Furthermore, the calculations suggested a modified excitation geometry that has increased our sizing resolution. (c) 2000 Optical Society of America

  7. Dual states estimation of a subsurface flow-transport coupled model using ensemble Kalman filtering

    KAUST Repository

    El Gharamti, Mohamad

    2013-10-01

    Modeling the spread of subsurface contaminants requires coupling a groundwater flow model with a contaminant transport model. Such coupling may provide accurate estimates of future subsurface hydrologic states if essential flow and contaminant data are assimilated in the model. Assuming perfect flow, an ensemble Kalman filter (EnKF) can be used for direct data assimilation into the transport model. This is, however, a crude assumption as flow models can be subject to many sources of uncertainty. If the flow is not accurately simulated, contaminant predictions will likely be inaccurate even after successive Kalman updates of the contaminant model with the data. The problem is better handled when both flow and contaminant states are concurrently estimated using the traditional joint state augmentation approach. In this paper, we introduce a dual estimation strategy for data assimilation into a one-way coupled system by treating the flow and the contaminant models separately while intertwining a pair of distinct EnKFs, one for each model. The presented strategy only deals with the estimation of state variables but it can also be used for state and parameter estimation problems. This EnKF-based dual state-state estimation procedure presents a number of novel features: (i) it allows for simultaneous estimation of both flow and contaminant states in parallel; (ii) it provides a time consistent sequential updating scheme between the two models (first flow, then transport); (iii) it simplifies the implementation of the filtering system; and (iv) it yields more stable and accurate solutions than does the standard joint approach. We conducted synthetic numerical experiments based on various time stepping and observation strategies to evaluate the dual EnKF approach and compare its performance with the joint state augmentation approach. Experimental results show that on average, the dual strategy could reduce the estimation error of the coupled states by 15% compared with the

  8. KINETIC MODEL OF ELECTRIC-DISCHARGE СО2-LASER WITH FAST FLOW

    Directory of Open Access Journals (Sweden)

    V. V. Nevdakh

    2013-01-01

    Full Text Available The paper presents a kinetic model of CW electric-discharge CO2-laser with fast flow. Expressions linking a non-saturated gain ratio, saturation intensity and output power of the fast-flow laser with excitation rates and relaxation times of laser levels have been obtained in the paper. The paper demonstrates that the higher excitation and flow rates or higher saturation intensity provide considerably higher specific output power of the fast-flow CO2-laser in comparison with a sealed-off CO2-laser. While maintaining a steady discharge the same output power of the fast-flow CO2-laser may be obtained under various discharge conditions and combinations of fast flow rate, gas mixture composition and active media temperature.

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

    Science.gov (United States)

    2014-10-01

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

  10. Phospho-specific flow cytometry identifies aberrant signaling in indolent B-cell lymphoma

    Directory of Open Access Journals (Sweden)

    Blix Egil S

    2012-10-01

    Full Text Available Abstract Background Knowledge about signaling pathways in malignant cells may provide prognostic and diagnostic information in addition to identify potential molecular targets for therapy. B-cell receptor (BCR and co-receptor CD40 signaling is essential for normal B cells, and there is increasing evidence that signaling via BCR and CD40 plays an important role in the pathogenesis of B-cell lymphoma. The aim of this study was to investigate basal and induced signaling in lymphoma B cells and infiltrating T cells in single-cell suspensions of biopsies from small cell lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL and marginal zone lymphoma (MZL patients. Methods Samples from untreated SLL/CLL and MZL patients were examined for basal and activation induced signaling by phospho-specific flow cytometry. A panel of 9 stimulation conditions targeting B and T cells, including crosslinking of the B cell receptor (BCR, CD40 ligand and interleukins in combination with 12 matching phospho-protein readouts was used to study signaling. Results Malignant B cells from SLL/CLL patients had higher basal levels of phosphorylated (p-SFKs, p-PLCγ, p-ERK, p-p38, p-p65 (NF-κB, p-STAT5 and p-STAT6, compared to healthy donor B cells. In contrast, anti-BCR induced signaling was highly impaired in SLL/CLL and MZL B cells as determined by low p-SFK, p-SYK and p-PLCγ levels. Impaired anti-BCR-induced p-PLCγ was associated with reduced surface expression of IgM and CD79b. Similarly, CD40L-induced p-ERK and p-p38 were also significantly reduced in lymphoma B cells, whereas p-p65 (NF-κB was equal to that of normal B cells. In contrast, IL-2, IL-7 and IL-15 induced p-STAT5 in tumor-infiltrating T cells were not different from normal T cells. Conclusions BCR signaling and CD40L-induced p-p38 was suppressed in malignant B cells from SLL/CLL and MZL patients. Single-cell phospho-specific flow cytometry for detection of basal as well as activation

  11. Experimental Development of Dual Phase Steel Laser-arc Hybrid Welding and its Comparison to Laser and Gas Metal Arc Welding

    Directory of Open Access Journals (Sweden)

    Wagner Duarte Antunes

    Full Text Available Abstract Dual phase DP600 steels have been used in many automobile structures and laser welding has been the standard method for the joining of different sections. This work proposed a comparison between laser welding with arc welding (GMAW and with hybrid laser-arc welding in order to access the microstructures and the mechanical behavior. The laser and hybrid welds are competitive in terms of microstructure and mechanical behavior, presenting both acceptable and tough welds. The maximum ductility of the laser and hybrid welds are very similar, around 14%, and near to the values observed in the base material. The GMAW presents low ductility due to the softening caused by tampering of the martensite, and thus is unacceptable as the welding procedure.

  12. A Dual-Crystal Cavity Ho,Tm:GdVO4 Laser

    International Nuclear Information System (INIS)

    Zhu Guo-Li; Ju You-Lun; Yao Bao-Quan; Wang Yue-Zhu

    2012-01-01

    We report a 31.2 W cw diode-pumped cryogenic Ho(0.4at.%),Tm(4at.%):GdVO 4 laser in a dual-crystal cavity. The pumping sources are two fiber-coupled laser diodes with a fiber core diameter of 0.4 mm, both of which can supply 42 W near 802 nm. With an incident pump power of 70.3 W at 802.4 nm, a cw output power of 31.2 W at 2.05 μm is attained, corresponding to an optical-to-optical conversion efficiency of 44.4%. The M 2 factor is measured as ∼1.3 under an output power of 20 W. (fundamental areas of phenomenology(including applications))

  13. Power-scalable, polarization-stable, dual-colour DFB fibre laser system for CW terahertz imaging

    DEFF Research Database (Denmark)

    Eichhorn, Finn; Pedersen, Jens Engholm; Jepsen, Peter Uhd

    Imaging with electromagnetic radiation in the terahertz (THz) range has received a large amount of attention during recent years. THz imaging systems have diverse potential application areas such as security screening, medical diagnostics and non-destructive testing. We will discuss a power......-scalable, dual-colour, polarization-maintaining distributed feedback (DFB) fibre laser system with an inherent narrow linewidth from the DFB fibre laser oscillators. The laser system can be used as source in CW THz systems employing photomixing (optical heterodyning) for generation and detection...

  14. Impact of cryopreservation on tetramer, cytokine flow cytometry, and ELISPOT

    Directory of Open Access Journals (Sweden)

    Morse Michael A

    2005-07-01

    Full Text Available Abstract Background Cryopreservation of PBMC and/or overnight shipping of samples are required for many clinical trials, despite their potentially adverse effects upon immune monitoring assays such as MHC-peptide tetramer staining, cytokine flow cytometry (CFC, and ELISPOT. In this study, we compared the performance of these assays on leukapheresed PBMC shipped overnight in medium versus cryopreserved PBMC from matched donors. Results Using CMV pp65 peptide pool stimulation or pp65 HLA-A2 tetramer staining, there was significant correlation between shipped and cryopreserved samples for each assay (p ≤ 0.001. The differences in response magnitude between cryopreserved and shipped PBMC specimens were not significant for most antigens and assays. There was significant correlation between CFC and ELISPOT assay using pp65 peptide pool stimulation, in both shipped and cryopreserved samples (p ≤ 0.001. Strong correlation was observed between CFC (using HLA-A2-restricted pp65 peptide stimulation and tetramer staining (p Conclusion We conclude that all three assays show concordant results on shipped versus cryopreserved specimens, when using a peptide-based readout. The assays are also concordant with each other in pair wise comparisons using equivalent antigen systems.

  15. Inert Carbon Nanoparticles for the Assessment of Preferential Flow in Saturated Dual-Permeability Porous Media

    KAUST Repository

    Yao, Chuanjin

    2017-06-07

    Knowledge of preferential flow in heterogeneous environments is essential for enhanced hydrocarbon recovery, geothermal energy extraction, and successful sequestration of chemical waste and carbon dioxide. Dual tracer tests using nanoparticles with a chemical tracer could indicate the preferential flow. A dual-permeability model with a high permeable core channel surrounded by a low permeable annulus was constructed and used to determine the viability of an inert carbon nanoparticle tracer for this application. A series of column experiments were conducted to demonstrate how this nanoparticle tracer can be used to implement the dual tracer tests in heterogeneous environments. The results indicate that, with the injection rate selected and controlled appropriately, nanoparticles together with a chemical tracer can assess the preferential flow in heterogeneous environments. The results also implement the dual tracer tests in heterogeneous environments by simultaneously injecting chemical and nanoparticle tracers.

  16. Rapid flow cytometry analysis of antimicrobial properties of nettle powder and cranberry powder

    Science.gov (United States)

    Hattuniemi, Maarit; Korhonen, Johanna; Jaakkola, Mari; Räty, Jarkko; Virtanen, Vesa

    2010-11-01

    Both nettle (Urtica dioica) and cranberry (Vaccinium oxycoccus) are widely known to have good influence on health. The aim of this study was to investigate antimicrobial properties of nettle powder and cranberry powder against Escherichia coli (E. coli) and monitor the growth of the bacteria by a rapid flow cytometry (FCM) method. For FCM measurements samples were stained with fluorescent dyes. The inhibitory effects of plant material on growth of E. coli were estimated by comparing the results of control sample (E. coli) to E. coli samples with plant material. FCM offers both a brilliant tool to investigate the kinetics of the growth of bacterium, since subsamples can be taken from the same liquid medium during the growing period and with fluorescent dyes a rapid method to investigate viability of the bacterium.

  17. Flow cytometry of duodenal intraepithelial lymphocytes improves diagnosis of celiac disease in difficult cases.

    Science.gov (United States)

    Valle, Julio; Morgado, José Mario T; Ruiz-Martín, Juan; Guardiola, Antonio; Lopes-Nogueras, Miriam; García-Vela, Almudena; Martín-Sacristán, Beatriz; Sánchez-Muñoz, Laura

    2017-10-01

    Diagnosis of celiac disease is difficult when the combined results of serology and histology are inconclusive. Studies using flow cytometry of intraepithelial lymphocytes (IELs) have found that celiac patients have increased numbers of γδ IELs, along with a decrease in CD3-CD103 + IELs. The objective of this article is to assess the role of flow cytometric analysis of IELs in the diagnosis of celiac disease in difficult cases. A total of 312 patients with suspicion of celiac disease were included in the study. Duodenal biopsy samples were used for histological assessment and for flow cytometric analysis of IELs. In 46 out of 312 cases (14.7%) the combination of serology and histology did not allow the confirmation or exclusion of celiac disease. HLA typing had been performed in 42 of these difficult cases. Taking into account HLA typing and the response to a gluten-free diet, celiac disease was excluded in 30 of these cases and confirmed in the remaining 12. Flow cytometric analysis of IELs allowed a correct diagnosis in 39 out of 42 difficult cases (92.8%) and had a sensitivity of 91.7% (95% CI: 61.5% to 99.8%) and a specificity of 93.3% (95% CI: 77.9% to 99.2%) for the diagnosis of celiac disease in this setting. Flow cytometric analysis of IELs is useful for the diagnosis of celiac disease in difficult cases.

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

    Science.gov (United States)

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

    2015-01-01

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

  19. Flow cytometry apparatus

    Science.gov (United States)

    Pinkel, D.

    1987-11-30

    An obstruction across the flow chamber creates a one-dimensional convergence of a sheath fluid. A passageway in the obstruction directs flat cells near to the area of one-dimensional convergence in the sheath fluid to provide proper orientation of flat cells at fast rates. 6 figs.

  20. Immunophenotype Discovery, Hierarchical Organization, and Template-based Classification of Flow Cytometry Samples

    Directory of Open Access Journals (Sweden)

    Ariful Azad

    2016-08-01

    Full Text Available We describe algorithms for discovering immunophenotypes from large collections of flow cytometry (FC samples, and using them to organize the samples into a hierarchy based on phenotypic similarity. The hierarchical organization is helpful for effective and robust cytometry data mining, including the creation of collections of cell populations characteristic of different classes of samples, robust classification, and anomaly detection. We summarize a set of samples belonging to a biological class or category with a statistically derived template for the class. Whereas individual samples are represented in terms of their cell populations (clusters, a template consists of generic meta-populations (a group of homogeneous cell populations obtained from the samples in a class that describe key phenotypes shared among all those samples. We organize an FC data collection in a hierarchical data structure that supports the identification of immunophenotypes relevant to clinical diagnosis. A robust template-based classification scheme is also developed, but our primary focus is in the discovery of phenotypic signatures and inter-sample relationships in an FC data collection. This collective analysis approach is more efficient and robust since templates describe phenotypic signatures common to cell populations in several samples, while ignoring noise and small sample-specific variations.We have applied the template-base scheme to analyze several data setsincluding one representing a healthy immune system, and one of Acute Myeloid Leukemia (AMLsamples. The last task is challenging due to the phenotypic heterogeneity of the severalsubtypes of AML. However, we identified thirteen immunophenotypes corresponding to subtypes of AML, and were able to distinguish Acute Promyelocytic Leukemia from other subtypes of AML.

  1. A dual exterior point simplex type algorithm for the minimum cost network flow problem

    Directory of Open Access Journals (Sweden)

    Geranis George

    2009-01-01

    Full Text Available A new dual simplex type algorithm for the Minimum Cost Network Flow Problem (MCNFP is presented. The proposed algorithm belongs to a special 'exterior- point simplex type' category. Similarly to the classical network dual simplex algorithm (NDSA, this algorithm starts with a dual feasible tree-solution and reduces the primal infeasibility, iteration by iteration. However, contrary to the NDSA, the new algorithm does not always maintain a dual feasible solution. Instead, the new algorithm might reach a basic point (tree-solution outside the dual feasible area (exterior point - dual infeasible tree.

  2. Engine flow visualization using a copper vapor laser

    Science.gov (United States)

    Regan, Carolyn A.; Chun, Kue S.; Schock, Harold J., Jr.

    1987-01-01

    A flow visualization system has been developed to determine the air flow within the combustion chamber of a motored, axisymmetric engine. The engine has been equipped with a transparent quartz cylinder, allowing complete optical access to the chamber. A 40-Watt copper vapor laser is used as the light source. Its beam is focused down to a sheet approximately 1 mm thick. The light plane is passed through the combustion chamber, and illuminates oil particles which were entrained in the intake air. The light scattered off of the particles is recorded by a high speed rotating prism movie camera. A movie is then made showing the air flow within the combustion chamber for an entire four-stroke engine cycle. The system is synchronized so that a pulse generated by the camera triggers the laser's thyratron. The camera is run at 5,000 frames per second; the trigger drives one laser pulse per frame. This paper describes the optics used in the flow visualization system, the synchronization circuit, and presents results obtained from the movie. This is believed to be the first published study showing a planar observation of airflow in a four-stroke piston-cylinder assembly. These flow visualization results have been used to interpret flow velocity measurements previously obtained with a laser Doppler velocimetry system.

  3. DAFi: A directed recursive data filtering and clustering approach for improving and interpreting data clustering identification of cell populations from polychromatic flow cytometry data.

    Science.gov (United States)

    Lee, Alexandra J; Chang, Ivan; Burel, Julie G; Lindestam Arlehamn, Cecilia S; Mandava, Aishwarya; Weiskopf, Daniela; Peters, Bjoern; Sette, Alessandro; Scheuermann, Richard H; Qian, Yu

    2018-04-17

    Computational methods for identification of cell populations from polychromatic flow cytometry data are changing the paradigm of cytometry bioinformatics. Data clustering is the most common computational approach to unsupervised identification of cell populations from multidimensional cytometry data. However, interpretation of the identified data clusters is labor-intensive. Certain types of user-defined cell populations are also difficult to identify by fully automated data clustering analysis. Both are roadblocks before a cytometry lab can adopt the data clustering approach for cell population identification in routine use. We found that combining recursive data filtering and clustering with constraints converted from the user manual gating strategy can effectively address these two issues. We named this new approach DAFi: Directed Automated Filtering and Identification of cell populations. Design of DAFi preserves the data-driven characteristics of unsupervised clustering for identifying novel cell subsets, but also makes the results interpretable to experimental scientists through mapping and merging the multidimensional data clusters into the user-defined two-dimensional gating hierarchy. The recursive data filtering process in DAFi helped identify small data clusters which are otherwise difficult to resolve by a single run of the data clustering method due to the statistical interference of the irrelevant major clusters. Our experiment results showed that the proportions of the cell populations identified by DAFi, while being consistent with those by expert centralized manual gating, have smaller technical variances across samples than those from individual manual gating analysis and the nonrecursive data clustering analysis. Compared with manual gating segregation, DAFi-identified cell populations avoided the abrupt cut-offs on the boundaries. DAFi has been implemented to be used with multiple data clustering methods including K-means, FLOCK, FlowSOM, and

  4. A flow cytometry-based workflow for detection and quantification of anti-plasmodial antibodies in vaccinated and naturally exposed individuals

    DEFF Research Database (Denmark)

    Ajua, Anthony; Engleitner, Thomas; Esen, Meral

    2012-01-01

    information about natural exposure and vaccine immunogenicity. A novel, cytometry-based workflow for the quantitative detection of anti-plasmodial antibodies in human serum is presented. METHODS: Fixed red blood cells (RBCs), infected with late stages of P. falciparum were utilized to detect malaria...... vaccine trials in semiimmune adults and pre-school children residing in a malaria endemic area. RESULTS: Fixation, permeabilization, and staining of infected RBCs were adapted for best operation in flow cytometry. As asexual vaccine candidates are designed to induce antibody patterns similar to semi...... with those obtained by manual gating (r between 0.79 and 0.99) and outperformed other model-driven gating methods. Bland-Altman plots confirmed the agreement of manual gating and OSA derived results. A-1.33 fold increase (p=0.003) in the number of positive cells after vaccination in a subgroup of preschool...

  5. Prognostic value of ZAP-70 expression in chronic lymphocytic leukemia as assessed by quantitative polymerase chain reaction and flow cytometry.

    Science.gov (United States)

    Adams, Rebecca L C; Cheung, Catherine; Banh, Raymond; Saal, Russell; Cross, Donna; Gill, Devinder; Self, Marlene; Klein, Kerenaftali; Mollee, Peter

    2014-03-01

    Chronic lymphocytic leukemia (CLL) is a disorder in which the tempo of disease progression is highly variable, and prognostic markers that can be utilized at diagnosis are regarded as clinically important. Currently, there are several prognostic factors, such as immunoglobulin heavy chain (IgVH) mutational status, and ZAP-70 protein expression in neoplastic B-cells, that have demonstrated significant discriminative power in the prognostication of CLL. They are, however, largely unavailable in the routine diagnostic laboratory setting. In this study, we characterized the IgVH status and ZAP-70 expression by molecular techniques in a cohort of 108 patients with CLL, and correlated these results with three different methods of ZAP-70 expression by flow cytometry. We then assessed the results of these methods in terms of prognostic power as characterized by time to first treatment (TTFT). By comparing three different flow cytometry methods using receiver–operator curve (ROC) analysis, we identified that by utilizing a corrected mean fluorescence intensity (CorrMFI) algorithm for assessing ZAP-70 expression, there was good correlation with both IgVH mutational status, and ZAP-70 expression as assessed by qPCR. We were also able to show that ZAP-70 expression, as assessed by both qPCR and the CorrMFI method, was prognostic of TTFT. While confirmation in a larger patient cohort, with longer follow-up is required, we believe that the CorrMFI represents the most promising method currently available in a routine diagnostic setting for the assessment of ZAP-70 expression in CLL patients. © 2013 International Clinical Cytometry Society.

  6. Laser Treatment of Professional Tattoos With a 1064/532-nm Dual-Wavelength Picosecond Laser.

    Science.gov (United States)

    Kauvar, Arielle N B; Keaney, Terrence C; Alster, Tina

    2017-12-01

    Picosecond-domain laser pulses improve the photomechanical disruption of tattoos. This study evaluates the efficacy and safety of a novel, dual-wavelength, 1,064/532-nm, picosecond-domain laser for tattoo clearance. This was a prospective, self-controlled, clinical study of 34 subjects with 39 tattoos treated at 2 sites with an interval of 4.8 ± 1.6 weeks and up to 10 treatments (mean, 7.5). Blinded evaluation and investigator assessment of serial digital images was performed to evaluate treatment efficacy in the 36 tattoos that received at least 3 treatments. Investigators also assessed efficacy before each treatment visit up to 10 treatments. Safety and tolerability was evaluated for all 39 tattoos that underwent at least 1 treatment. Blinded evaluation demonstrated that lightening of tattoos was achieved in all subjects, with 86% (31 of 36 tattoos) showing at least a 50% clearance after 3 treatments. Adverse events were few and transient in nature. Patient satisfaction and treatment tolerability were high. Treatment of single-colored and multicolored tattoos with this novel 1,064/532-nm picosecond laser is highly safe and effective.

  7. Dual-wavelength mid-infrared CW and Q-switched laser in diode end-pumped Tm,Ho:GdYTaO4 crystal

    Science.gov (United States)

    Wang, Beibei; Gao, Congcong; Dou, Renqin; Nie, Hongkun; Sun, Guihua; Liu, Wenpeng; Yu, Haijuan; Wang, Guoju; Zhang, Qingli; Lin, Xuechun; He, Jingliang; Wang, Wenjun; Zhang, Bingyuan

    2018-02-01

    Dual-wavelength continuous-wave and Q-switched lasers are demonstrated in a Tm,Ho:GdYTaO4 crystal under 790 nm laser diode end pumping for the first time to the best of our knowledge. The laser operates with a dual wavelength at 1949.677 nm and 2070 nm for continuous-wave with a spacing of about 120 nm. The maximum output power is 0.332 W with a pump power of 3 W. By using graphene as the saturable absorber, a passively Q-switched operation is performed with a dual-wavelength at 1950.323 nm and 2068.064 nm with a wavelength interval of about 118 nm. The maximum average output power of the Q-switched laser goes up to 200 mW with a minimum pulse duration of 1.2 µs and a maximum repetition rate of 34.72 kHz.

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

    Science.gov (United States)

    Leif, Robert C.

    2007-02-01

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

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

    Science.gov (United States)

    Chen, Tiffany J; Kotecha, Nikesh

    2014-01-01

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

  10. Rapid Identification of Staphylococcus aureus and Methicillin Resistance by Flow Cytometry Using a Peptide Nucleic Acid Probe ▿

    Science.gov (United States)

    Shrestha, Nabin K.; Scalera, Nikole M.; Wilson, Deborah A.; Brehm-Stecher, Byron; Procop, Gary W.

    2011-01-01

    A total of 56 Staphylococcus aureus isolates incubated for 2 h in the presence or absence of oxacillin were analyzed by flow cytometry after labeling with an S. aureus-specific peptide nucleic acid (PNA) probe. Two defined ratios, the paired signal count ratio (PSCR) and the gate signal count ratio (GSCR), differentiated methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) with sensitivities of 100% each and specificities of 96% and 100%, respectively. PMID:21795508

  11. Differences in leukocyte differentiation molecule abundances on domestic sheep (Ovis aries) and bighorn sheep (Ovis canadensis) neutrophils identified by flow cytometry.

    Science.gov (United States)

    Highland, Margaret A; Schneider, David A; White, Stephen N; Madsen-Bouterse, Sally A; Knowles, Donald P; Davis, William C

    2016-06-01

    Although both domestic sheep (DS) and bighorn sheep (BHS) are affected by similar respiratory bacterial pathogens, experimental and field data indicate BHS are more susceptible to pneumonia. Cross-reactive monoclonal antibodies (mAbs) for use in flow cytometry (FC) are valuable reagents for interspecies comparative immune system analyses. This study describes cross-reactive mAbs that recognize leukocyte differentiation molecules (LDMs) and major histocompatibility complex antigens on DS and BHS leukocytes. Characterization of multichannel eosinophil autofluorescence in this study permitted cell-type specific gating of granulocytes for evaluating LDMs, specifically on neutrophils, by single-label FC. Evaluation of relative abundances of LDMs by flow cytometry revealed greater CD11a, CD11b, CD18 (β2 integrins) and CD 172a (SIRPα) on DS neutrophils and greater CD14 (lipopolysaccharide receptor) on BHS neutrophils. Greater CD25 (IL-2) was identified on BHS lymphocytes following Concavalin A stimulation. While DS and BHS have similar total peripheral blood leukocyte counts, BHS have proportionately more neutrophils. Published by Elsevier Ltd.

  12. Chromium-doped diamond-like carbon films deposited by dual-pulsed laser deposition

    Czech Academy of Sciences Publication Activity Database

    Písařík, Petr; Jelínek, Miroslav; Kocourek, Tomáš; Zezulová, M.; Remsa, Jan; Jurek, Karel

    2014-01-01

    Roč. 117, č. 1 (2014), s. 83-88 ISSN 0947-8396 R&D Projects: GA MŠk LD12069 Institutional support: RVO:68378271 Keywords : diamond like carbon * chromium * contact angle * surface free energy * dual laser deposition * zeta potential Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 1.704, year: 2014

  13. Accurate live and dead bacterial cell enumeration using flow cytometry (Conference Presentation)

    Science.gov (United States)

    Ou, Fang; McGoverin, Cushla; Swift, Simon; Vanholsbeeck, Frédérique

    2017-03-01

    Flow cytometry (FCM) is based on the detection of scattered light and fluorescence to identify cells with particular characteristics of interest. However most FCM cannot precisely control the flow through its interrogation point and hence the volume and concentration of the sample cannot be immediately obtained. The easiest, most reliable and inexpensive way of obtaining absolute counts with FCM is by using reference beads. We investigated a method of using FCM with reference beads to measure live and dead bacterial concentration over the range of 106 to 108 cells/mL and ratio varying from 0 to 100%. We believe we are the first to use this method for such a large cell concentration range while also establishing the effect of varying the live/dead bacteria ratios. Escherichia coli solutions with differing ratios of live:dead cells were stained with fluorescent dyes SYTO 9 and propidium iodide (PI), which label live and dead cells, respectively. Samples were measured using a LSR II Flow Cytometer (BD Biosciences); using 488 nm excitation with 20 mW power. Both SYTO 9 and PI fluorescence were collected and threshold was set to side scatter. Traditional culture-based plate count was done in parallel to the FCM analysis. The concentration of live bacteria from FCM was compared to that obtained by plate counts. Preliminary results show that the concentration of live bacteria obtained by FCM and plate counts correlate well with each other and indicates this may be extended to a wider concentration range or for studying other cell characteristics.

  14. High Power Q-Switched Dual-End-Pumped Ho:YAG Laser

    Energy Technology Data Exchange (ETDEWEB)

    Xiao-Ming, Duan; Ying-Jie, Shen; Tong-Yu, Dai; Bao-Quan, Yao; Wang Yue-Zhu, E-mail: xmduan@hit.edu.cn [National Key Laboratory of Tunable Laser Technology, Harbin Institute of Technology, Harbin 150001 (China)

    2012-09-15

    We report the high power acousto-optically Q-switched operation of a dual-end-pumped Ho:YAG laser at room temperature. For the Q-swithched mode, a maximum pulse energy of 2.4 mJ and a minimum pulse width of 23 ns at the repetition rate of 10 kHz are achieved, resulting in a peak power of 104.3 kW. The beam quality factor of M{sup 2} {approx} 1.5, which is demonstrated by a knife-edge method. In addition, the Ho:YAG laser is employed as a pumping source of ZGP optical parametric oscillator, and its total average output power is 13.2 W at 3.9 {mu}m and 4.4 {mu}m with a slope efficiency of 68.4%.

  15. Novel Strategy for Phenotypic Characterization of Human B Lymphocytes from Precursors to Effector Cells by Flow Cytometry.

    Directory of Open Access Journals (Sweden)

    Giovanna Clavarino

    Full Text Available A precise identification and phenotypic characterization of human B-cell subsets is of crucial importance in both basic research and medicine. In the literature, flow cytometry studies for the phenotypic characterization of B-lymphocytes are mainly focused on the description of a particular cell stage, or of specific cell stages observed in a single type of sample. In the present work, we propose a backbone of 6 antibodies (CD38, CD27, CD10, CD19, CD5 and CD45 and an efficient gating strategy to identify, in a single analysis tube, a large number of B-cell subsets covering the whole B-cell differentiation from precursors to memory and plasma cells. Furthermore, by adding two antibodies in an 8-color combination, our approach allows the analysis of the modulation of any cell surface marker of interest along B-cell differentiation. We thus developed a panel of seven 8-colour antibody combinations to phenotypically characterize B-cell subpopulations in bone marrow, peripheral blood, lymph node and cord blood samples. Beyond qualitative information provided by biparametric representations, we also quantified antigen expression on each of the identified B-cell subsets and we proposed a series of informative curves showing the modulation of seventeen cell surface markers along B-cell differentiation. Our approach by flow cytometry provides an efficient tool to obtain quantitative data on B-cell surface markers expression with a relative easy-to-handle technique that can be applied in routine explorations.

  16. Rapid dual-injection single-scan 13N-ammonia PET for quantification of rest and stress myocardial blood flows

    International Nuclear Information System (INIS)

    Rust, T C; DiBella, E V R; McGann, C J; Christian, P E; Hoffman, J M; Kadrmas, D J

    2006-01-01

    Quantification of myocardial blood flows at rest and stress using 13 N-ammonia PET is an established method; however, current techniques require a waiting period of about 1 h between scans. The objective of this study was to test a rapid dual-injection single-scan approach, where 13 N-ammonia injections are administered 10 min apart during rest and adenosine stress. Dynamic PET data were acquired in six human subjects using imaging protocols that provided separate single-injection scans as gold standards. Rest and stress data were combined to emulate rapid dual-injection data so that the underlying activity from each injection was known exactly. Regional blood flow estimates were computed from the dual-injection data using two methods: background subtraction and combined modelling. The rapid dual-injection approach provided blood flow estimates very similar to the conventional single-injection standards. Rest blood flow estimates were affected very little by the dual-injection approach, and stress estimates correlated strongly with separate single-injection values (r = 0.998, mean absolute difference = 0.06 ml min -1 g -1 ). An actual rapid dual-injection scan was successfully acquired in one subject and further demonstrates feasibility of the method. This study with a limited dataset demonstrates that blood flow quantification can be obtained in only 20 min by the rapid dual-injection approach with accuracy similar to that of conventional separate rest and stress scans. The rapid dual-injection approach merits further development and additional evaluation for potential clinical use

  17. Evaluating the effects of allelochemical ferulic acid on Microcystis aeruginosa by pulse-amplitude-modulated (PAM) fluorometry and flow cytometry.

    Science.gov (United States)

    Wang, Rui; Hua, Ming; Yu, Yang; Zhang, Min; Xian, Qi-Ming; Yin, Da-Qiang

    2016-03-01

    We investigated the effects of allelochemical ferulic acid (FA) on a series of physiological and biochemical processes of blue-green algae Microcystis aeruginosa, in order to find sensitive diagnostic variables for allelopathic effects. Algal cell density was significantly suppressed by FA (0.31-5.17 mM) only after 48 h exposure. Inhibitions of photosynthetic parameters (F(v)/F(m) and F(v)'/F(m)') occurred more rapidly than cell growth, and the stimulation of non-photochemical quenching was observed as a feed-back mechanisms induced by photosystem II blockage, determining by PAM fluorometry. Inhibitions on esterase activity, membrane potential and integrity, as well as disturbance on cell size, were all detected by flow cytometry with specific fluorescent markers, although exhibiting varied sensitivities. Membrane potential and esterase activity were identified as the most sensitive parameters (with relatively lower EC50 values), and responded more rapidly (significantly inhibited only after 8 h exposure) than photosynthetic parameters and cell growth, thus may be the primary responses of cyanobacteria to FA exposure. The use of PAM fluorometry and flow cytometry for rapid assessment of those sensitive variables may contribute to future mechanistic studies of allolepathic effects on phytoplankton. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. The auroral and ionospheric flow signatures of dual lobe reconnection

    Directory of Open Access Journals (Sweden)

    S. M. Imber

    2006-11-01

    Full Text Available We present the first substantial evidence for the occurrence of dual lobe reconnection from ionospheric flows and auroral signatures. The process of dual lobe reconnection refers to an interplanetary magnetic field line reconnecting with lobe field lines in both the northern and southern hemispheres. Two bursts of sunward plasma flow across the noon portion of the open/closed field line boundary (OCB, indicating magnetic flux closure at the dayside, were observed in SuperDARN radar data during a period of strongly northward IMF. The OCB is identified from spacecraft, radar backscatter, and auroral observations. In order for dual lobe reconnection to take place, we estimate that the interplanetary magnetic field clock angle must be within ±10° of zero (North. The total flux crossing the OCB during each burst is small (1.8% and 0.6% of the flux contained within the polar cap for the two flows. A brightening of the noon portion of the northern auroral oval was observed as the clock angle passed through zero, and is thought to be due to enhanced precipitating particle fluxes due to the occurrence of reconnection at two locations along the field line. The number of solar wind protons captured by the flux closure process was estimated to be ~2.5×1030 (4 tonnes by mass, sufficient to populate the cold, dense plasma sheet observed following this interval.

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

    Science.gov (United States)

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

    2014-09-01

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

  20. A Simplified Micromechanical Modeling Approach to Predict the Tensile Flow Curve Behavior of Dual-Phase Steels

    Science.gov (United States)

    Nanda, Tarun; Kumar, B. Ravi; Singh, Vishal

    2017-11-01

    Micromechanical modeling is used to predict material's tensile flow curve behavior based on microstructural characteristics. This research develops a simplified micromechanical modeling approach for predicting flow curve behavior of dual-phase steels. The existing literature reports on two broad approaches for determining tensile flow curve of these steels. The modeling approach developed in this work attempts to overcome specific limitations of the existing two approaches. This approach combines dislocation-based strain-hardening method with rule of mixtures. In the first step of modeling, `dislocation-based strain-hardening method' was employed to predict tensile behavior of individual phases of ferrite and martensite. In the second step, the individual flow curves were combined using `rule of mixtures,' to obtain the composite dual-phase flow behavior. To check accuracy of proposed model, four distinct dual-phase microstructures comprising of different ferrite grain size, martensite fraction, and carbon content in martensite were processed by annealing experiments. The true stress-strain curves for various microstructures were predicted with the newly developed micromechanical model. The results of micromechanical model matched closely with those of actual tensile tests. Thus, this micromechanical modeling approach can be used to predict and optimize the tensile flow behavior of dual-phase steels.

  1. Two new nuclear isolation buffers for plant DNA flow cytometry: A test with 37 species

    Czech Academy of Sciences Publication Activity Database

    Loureiro, J.; Rodriguez, E.; Doležel, Jaroslav; Santos, C.

    2007-01-01

    Roč. 100, č. 4 (2007), s. 875-888 ISSN 0305-7364 R&D Projects: GA ČR GA521/06/1723; GA ČR(CZ) GA521/05/0257; GA MŠk(CZ) LC06004 Grant - others:Mendelova zemědělská a lesnická univerzita v Brně / Agronomická fakulta(CZ) ME 844 Institutional research plan: CEZ:AV0Z50380511 Source of funding: V - iné verejné zdroje ; V - iné verejné zdroje ; V - iné verejné zdroje ; V - iné verejné zdroje Keywords : flow cytometry * general purpose buffer * genome size Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.939, year: 2007

  2. Hierarchical modeling for rare event detection and cell subset alignment across flow cytometry samples.

    Directory of Open Access Journals (Sweden)

    Andrew Cron

    Full Text Available Flow cytometry is the prototypical assay for multi-parameter single cell analysis, and is essential in vaccine and biomarker research for the enumeration of antigen-specific lymphocytes that are often found in extremely low frequencies (0.1% or less. Standard analysis of flow cytometry data relies on visual identification of cell subsets by experts, a process that is subjective and often difficult to reproduce. An alternative and more objective approach is the use of statistical models to identify cell subsets of interest in an automated fashion. Two specific challenges for automated analysis are to detect extremely low frequency event subsets without biasing the estimate by pre-processing enrichment, and the ability to align cell subsets across multiple data samples for comparative analysis. In this manuscript, we develop hierarchical modeling extensions to the Dirichlet Process Gaussian Mixture Model (DPGMM approach we have previously described for cell subset identification, and show that the hierarchical DPGMM (HDPGMM naturally generates an aligned data model that captures both commonalities and variations across multiple samples. HDPGMM also increases the sensitivity to extremely low frequency events by sharing information across multiple samples analyzed simultaneously. We validate the accuracy and reproducibility of HDPGMM estimates of antigen-specific T cells on clinically relevant reference peripheral blood mononuclear cell (PBMC samples with known frequencies of antigen-specific T cells. These cell samples take advantage of retrovirally TCR-transduced T cells spiked into autologous PBMC samples to give a defined number of antigen-specific T cells detectable by HLA-peptide multimer binding. We provide open source software that can take advantage of both multiple processors and GPU-acceleration to perform the numerically-demanding computations. We show that hierarchical modeling is a useful probabilistic approach that can provide a

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

    Science.gov (United States)

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

    2017-09-14

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

  4. Prospective study of removing solar lentigines in Asians using a novel dual-wavelength and dual-pulse width picosecond laser.

    Science.gov (United States)

    Negishi, Kei; Akita, Hirotaka; Matsunaga, Yukiko

    2018-04-02

    Quality-switched (QS) lasers are known to be an effective treatment for removing solar lentigines, however, high incidence of post-inflammatory hyperpigmentation (PIH) is a concern in darker skin types. The objective of this study was to evaluate the efficacy and safety of a dual-wavelength and dual-pulse width picosecond Nd:YAG laser for removing solar lentigines in Asians. This was a prospective, IRB-approved study. Twenty cases with solar lentigines on the face were enrolled for treatment and evaluated at 1- and 3-month after the final treatment. Results were assessed by blinded evaluators using a 5-grade percentage improvement scale and Melanin index (MI) measured by a reflectance spectrophotometer. A patient self-assessment questionnaire was also administered using a 5-grade improvement scale. Additional treatment was performed if the improvement was less than 75% or the lentigo partially remained after 4 weeks. Histological evaluation was performed to compare the differences between the current picosecond laser and a QS Nd:YAG laser 532-nm using light and electron microscopy. Forty-three lesions in 20 females, skin type III or IV, age 53.7 ± 9.75 were treated and evaluated. The laser setting was: 532-nm, 750 picoseconds, average fluence of 0.35 ± 0.06 J/cm [2] using a spot size of 3 or 4 mm. Forty lesions (93.02%) achieved over 75% clearance with a single treatment and the other three lesions (6.98%) needed two treatments. PIH occurred only in 4.65% of lesions. The average score of the blinded evaluators' assessment was 4.77 and 4.58 on a 5-grade percentage improvement scale. The patients' self-assessment rating was 4.76 and 4.67 on a 5-grade scale at 1- and 3-month follow-up, respectively. The improvement rate of relative MI (MI in the lesion minus that of the normal area) was 77.60 ± 36.27% and 76.93 ± 20.95% at 1-and 3-month follow-up. Histology showed vacuolar formation by both lasers in the epidermis that were different sizes

  5. Laser-based ultrasonics by dual-probe interferometer detection and narrow-band ultrasound generation

    Science.gov (United States)

    Huang, Jin

    1993-01-01

    Despite the advantages of laser-based ultrasonic (LBU) systems, the overall sensitivity of LBU systems needs to be improved for practical applications. Progress is reported to achieve better LBU detection accuracy and sensitivity for applications with surface waves and Lamb waves. A novel dual-probe laser interferometer has been developed to measure the same signal at two points. The dual-probe interferometer is a modification of a conventional single-probe interferometer in that the reference beam is guided to a second detecting point on the specimen surface to form a differential measurement mode, which measure the difference of the displacements at the two points. This dual-probe interferometer is particularly useful for accurate measurements of the speed and attenuation of surface waves and Lamb waves. The dual-probe interferometer has been applied to obtain accurate measurements of the surface wave speed and attenuation on surfaces of increasing surface roughness. It has also been demonstrated that with an appropriate signal processing method, namely, the power cepstrum method, the dual-probe interferometer is applicable to measure the local surface wave speed even when the probe separation is so small that the two waveforms in the interferometer output signal overlap in the time domain. Narrow-band signal generation and detection improve the sensitivity of LBU systems. It is proposed to use a diffraction grating to form an array of illuminating strips which form a source of narrowband surface and Lamb waves. The line-array of thermoelastic sources generates narrow-band signals whose frequency and bandwidth can be easily controlled. The optimum line-array parameters, such as width, spacing and the number of lines in the array have been derived theoretically and verified experimentally. Narrow-band signal generation with optimum parameters has been demonstrated. The enhanced LBU system with dual-probe detection and narrowband signal generation has been

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

    Science.gov (United States)

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

    1986-12-01

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

  7. Flowing Air-Water Cooled Slab Nd: Glass Laser

    Science.gov (United States)

    Lu, Baida; Cai, Bangwei; Liao, Y.; Xu, Shifa; Xin, Z.

    1989-03-01

    A zig-zag optical path slab geometry Nd: glass laser cooled through flowing air-water is developed by us. Theoretical studies on temperature distribution of slab and rod configurations in the unsteady state clarify the advantages of the slab geometry laser. The slab design and processing are also reported. In our experiments main laser output characteristics, e. g. laser efficiency, polarization, far-field divergence angle as well as resonator misalignment are investigated. The slab phosphate glass laser in combination with a crossed Porro-prism resonator demonstrates a good laser performance.

  8. In-situ characterization of symmetric dual-pass architecture of microfluidic co-laminar flow cells

    International Nuclear Information System (INIS)

    Ibrahim, Omar A.; Goulet, Marc-Antoni; Kjeang, Erik

    2016-01-01

    Highlights: • An analytical cell design is proposed for characterization of dual-pass flow cells • High power density up to 0.75 W cm −2 is demonstrated • The performance contributions of the inlet and outlet passes are of the same order • Downstream crossover is analyzed as a function of cell current and flow rate - Abstract: Microfluidic co-laminar flow cells with dual-pass architecture enable fuel recirculation and in-situ regeneration, and offer improvements in performance characteristics. In this work, a unique analytical cell design is proposed, with two split portions having flow-through porous electrodes. Each cell portion is first tested individually with vanadium redox species and the results are used to quantify the previously unknown crossover losses at the downstream portion of the cell, shown here to be a strong function of the flow rate. Moreover, the upstream cell portion demonstrates impressive room-temperature power density up to 0.75 W cm −2 at 1.0 A cm −2 , which is the highest performance reported to date for a microfluidic vanadium redox battery. Next, the two cell portions are connected in parallel to resemble a complete cell with dual-pass architecture, thereby enabling novel in-situ diagnostics of the inlet and outlet passes of the cell. For instance, the reactant utilization efficiency of the downstream cell portion is shown to be on the same order as that of the upstream portion at both low and high flow rates. Furthermore, in-situ regeneration is also demonstrated. Overall, the present results provide a deeper understanding of dual-pass reactant conversion and crossover which will be useful for future device optimization.

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

    NARCIS (Netherlands)

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

    2009-01-01

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

  10. Flow tube used to cool solar-pumped laser

    Science.gov (United States)

    1968-01-01

    A flow tube has been designed and constructed to provide two major functions in the application of a laser beam for transmission of both sound and video. It maintains the YAG laser at the proper operating temperature of 300 degrees K under solar pumping conditions, and it serves as a pump cavity for the laser crystal.

  11. A novel dual-wavelength laser stimulator to elicit transient and tonic nociceptive stimulation.

    Science.gov (United States)

    Dong, Xiaoxi; Liu, Tianjun; Wang, Han; Yang, Jichun; Chen, Zhuying; Hu, Yong; Li, Yingxin

    2017-07-01

    This study aimed to develop a new laser stimulator to elicit both transient and sustained heat stimulation with a dual-wavelength laser system as a tool for the investigation of both transient and tonic experimental models of pain. The laser stimulator used a 980-nm pulsed laser to generate transient heat stimulation and a 1940-nm continuous-wave (CW) laser to provide sustained heat stimulation. The laser with 980-nm wavelength can elicit transient pain with less thermal injury, while the 1940-nm CW laser can effectively stimulate both superficial and deep nociceptors to elicit tonic pain. A proportional integral-derivative (PID) temperature feedback control system was implemented to ensure constancy of temperature during heat stimulation. The performance of this stimulator was evaluated by in vitro and in vivo animal experiments. In vitro experiments on totally 120 specimens fresh pig skin included transient heat stimulation by 980-nm laser (1.5 J, 10 ms), sustained heat stimulation by 1940-nm laser (50-55 °C temperature control mode or 1.5 W, 5 min continuous power supply), and the combination of transient/sustained heat stimulation by dual lasers (1.5 J, 10 ms, 980-nm pulse laser, and 1940-nm laser with 50-55 °C temperature control mode). Hemoglobin brushing and wind-cooling methods were tested to find better stimulation model. A classic tail-flick latency (TFL) experiment with 20 Wistar rats was used to evaluate the in vivo efficacy of transient and tonic pain stimulation with 15 J, 100 ms 980-nm single laser pulse, and 1.5 W constant 1940-nm laser power. Ideal stimulation parameters to generate transient pain were found to be a 26.6 °C peak temperature rise and 0.67 s pain duration. In our model of tonic pain, 5 min of tonic stimulation produced a temperature change of 53.7 ± 1.3 °C with 1.6 ± 0.2% variation. When the transient and tonic stimulation protocols were combined, no significant difference was observed depending on the order

  12. Dual-wavelength DFB quantum cascade lasers: sources for multi-species trace gas spectroscopy

    Science.gov (United States)

    Kapsalidis, Filippos; Shahmohammadi, Mehran; Süess, Martin J.; Wolf, Johanna M.; Gini, Emilio; Beck, Mattias; Hundt, Morten; Tuzson, Béla; Emmenegger, Lukas; Faist, Jérôme

    2018-06-01

    We report on the design, fabrication, and performance of dual-wavelength distributed-feedback (DFB) quantum cascade lasers (QCLs) emitting at several wavelengths in the mid-infrared (mid-IR) spectrum. In this work, two new designs are presented: for the first one, called "Neighbour" DFB, two single-mode DFB QCLs are fabricated next to each other, with minimal lateral distance, to allow efficient beam-coupling into multi-pass gas cells. In addition, the minimal distance allows either laser to be used as an integrated heater for the other, allowing to extend the tuning range of its neighbour without any electrical cross-talk. For the second design, the Vernier effect was used to realize a switchable DFB laser, with two target wavelengths which are distant by about 300 cm^{-1}. These devices are promising laser sources for Tunable Diode Laser Absorption Spectroscopy applications targeting simultaneous detection of multiple gasses, with distant spectral features, in compact and mobile setups.

  13. Compound technology of manufacturing and multiple laser peening on microstructure and fatigue life of dual-phase spring steel

    Energy Technology Data Exchange (ETDEWEB)

    Prabhakaran, S., E-mail: spkaran.kmd@gmail.com; Kalainathan, S., E-mail: kalainathan@yahoo.com

    2016-09-30

    The present work proposes an advanced double quenching and tempering heat treatment based laser surface modification process of dual-phase spring steel. Multiple laser peening without coating process utilized the decarburized surface as the protective layer for the further cold working process. The electron backscattering diffraction analysis on crystallographic orientation of individual grains and phase map exhibits a perfect dual-phase steel. Also, the high resolution transmission electron microscopic study explains the high strain induced microstructural grain refinement features and plastic deformation behaviors. The laser peening technique taking an advantage that it induces a large and high magnitude compressive residual stress with good thermal stability. The micro and nano-hardness profile provides better surface and sub-surface mechanical properties. The controlled average surface roughness is achieved in this course of work. The stress-strain characteristics on tensile properties are analyzed through the pre-fatigued specimens. The fully reversed high cycle fatigue test indicates that the current laser peening has substantially improves the fatigue life of the specimens.

  14. Comparative analysis of minimal residual disease detection by multiparameter flow cytometry and enhanced ASO RQ-PCR in multiple myeloma

    International Nuclear Information System (INIS)

    Silvennoinen, R; Lundan, T; Kairisto, V; Pelliniemi, T-T; Putkonen, M; Anttila, P; Huotari, V; Mäntymaa, P; Siitonen, S; Uotila, L; Penttilä, T-L; Juvonen, V; Selander, T; Remes, K

    2014-01-01

    Multiparameter flow cytometry (MFC) and allele-specific oligonucleotide real-time quantitative PCR (ASO RQ-PCR) are the two most sensitive methods to detect minimal residual disease (MRD) in multiple myeloma (MM). We compared these methods in 129 paired post-therapy samples from 22 unselected, consecutive MM patients in complete/near complete remission. Appropriate immunophenotypic and ASO RQ-PCR-MRD targets could be detected and MRD analyses constructed for all patients. The high PCR coverage could be achieved by gradual widening of the primer sets used for clonality detection. In addition, for 13 (55%) of the patients, reverse orientation of the ASO primer and individual design of the TaqMan probe improved the sensitivity and specificity of ASO RQ-PCR analysis. A significant nonlinear correlation prevailed between MFC-MRD and PCR-MRD when both were positive. Discordance between the methods was found in 32 (35%) paired samples, which were negative by MFC-MRD, but positive by ASO RQ-PCR. The findings suggest that with the described technique, ASO RQ-PCR can be constructed for all patients with MM. ASO RQ-PCR is slightly more sensitive in MRD detection than 6−10-color flow cytometry. Owing to technical demands ASO RQ-PCR could be reserved for patients in immunophenotypic remission, especially in efficacy comparisons between different drugs and treatment modalities

  15. Assessment of a flow cytometry technique for studying signaling pathways in platelets: Monitoring of VASP phosphorylation in clinical samples

    Directory of Open Access Journals (Sweden)

    N. Mallouk

    2018-07-01

    Full Text Available A recently released kit (PerFix EXPOSE was reported to improve the measurement of the degree of phosphorylation of proteins in leukocytes by flow cytometry. We tested its adaptation for platelets to monitor vasodilator-stimulated phosphoprotein (VASP phosphorylation, which is the basis of a currently used test for the assessment of the pharmacological response to P2Y12 antagonists (PLT VASP/P2Y12. The PerFix EXPOSE kit was compared to the PLT VASP/P2Y12 kit by using blood samples drawn at 24 h post clopidogrel dose from 19 patients hospitalized for a non-cardio-embolic ischemic stroke and treated with clopidogrel monotherapy for at least five days in an observational study. The platelet PerFix method was based on adaptation of the volume of the sample, the centrifugation speed and the incubation temperature. Poor agreement between prevention by adenosine diphosphate (ADP of PGE1-induced cAMP-mediated VASP phosphorylation and ADP induced aggregation assessed by Light Transmittance Aggregometry was found. We found a significant correlation between the PLT VASP/P2Y12 kit and the PerFix EXPOSE kit. The PerFix EXPOSE kit may also be helpful to monitor adverse effects of second-generation tyrosine kinase inhibitors on platelets. Keywords: Platelet signaling, VASP, Flow cytometry, Clopidogrel

  16. Flow cytometry combined with viSNE for the analysis of microbial biofilms and detection of microplastics

    Science.gov (United States)

    Sgier, Linn; Freimann, Remo; Zupanic, Anze; Kroll, Alexandra

    2016-01-01

    Biofilms serve essential ecosystem functions and are used in different technical applications. Studies from stream ecology and waste-water treatment have shown that biofilm functionality depends to a great extent on community structure. Here we present a fast and easy-to-use method for individual cell-based analysis of stream biofilms, based on stain-free flow cytometry and visualization of the high-dimensional data by viSNE. The method allows the combined assessment of community structure, decay of phototrophic organisms and presence of abiotic particles. In laboratory experiments, it allows quantification of cellular decay and detection of survival of larger cells after temperature stress, while in the field it enables detection of community structure changes that correlate with known environmental drivers (flow conditions, dissolved organic carbon, calcium) and detection of microplastic contamination. The method can potentially be applied to other biofilm types, for example, for inferring community structure for environmental and industrial research and monitoring. PMID:27188265

  17. Continuous-wave dual-wavelength operation of a distributed feedback laser diode with an external cavity using a volume Bragg grating

    Science.gov (United States)

    Zheng, Yujin; Sekine, Takashi; Kurita, Takashi; Kato, Yoshinori; Kawashima, Toshiyuki

    2018-03-01

    We demonstrate continuous-wave dual-wavelength operation of a broad-area distributed feedback (DFB) laser diode with a single external-cavity configuration. This high-power DFB laser has a narrow bandwidth (current and temperature ranges.

  18. Stabilization of self-mode-locked quantum dash lasers by symmetric dual-loop optical feedback

    Science.gov (United States)

    Asghar, Haroon; Wei, Wei; Kumar, Pramod; Sooudi, Ehsan; McInerney, John. G.

    2018-02-01

    We report experimental studies of the influence of symmetric dual-loop optical feedback on the RF linewidth and timing jitter of self-mode-locked two-section quantum dash lasers emitting at 1550 nm. Various feedback schemes were investigated and optimum levels determined for narrowest RF linewidth and low timing jitter, for single-loop and symmetric dual-loop feedback. Two symmetric dual-loop configurations, with balanced and unbalanced feedback ratios, were studied. We demonstrate that unbalanced symmetric dual loop feedback, with the inner cavity resonant and fine delay tuning of the outer loop, gives narrowest RF linewidth and reduced timing jitter over a wide range of delay, unlike single and balanced symmetric dual-loop configurations. This configuration with feedback lengths 80 and 140 m narrows the RF linewidth by 4-67x and 10-100x, respectively, across the widest delay range, compared to free-running. For symmetric dual-loop feedback, the influence of different power split ratios through the feedback loops was determined. Our results show that symmetric dual-loop feedback is markedly more effective than single-loop feedback in reducing RF linewidth and timing jitter, and is much less sensitive to delay phase, making this technique ideal for applications where robustness and alignment tolerance are essential.

  19. Quantifying Information Flow between Two Chaotic Semiconductor Lasers Using Symbolic Transfer Entropy

    International Nuclear Information System (INIS)

    Li Nian-Qiang; Pan Wei; Yan Lian-Shan; Luo Bin; Xu Ming-Feng; Tang Yi-Long

    2012-01-01

    Symbolic transfer entropy (STE) is employed to quantify the dominant direction of information flow between two chaotic-semiconductor-laser time series. The information flow in unidirectionally and bidirectionally coupled systems was analyzed systematically. Numerical results show that the dependence relationship can be revealed if there exists any coupling between two chaotic semiconductor lasers. More importantly, in both unsynchronized and good synchronization regimes, the STE can be used to quantify the direction of information flow between the lasers, although the former case leads to a better identification. The results thus establish STE as an effective tool for quantifying the direction of information flow between chaotic-laser-based systems

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

    Science.gov (United States)

    Bhattacharyya, Kiran

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

  1. Coupling Bacterial Activity Measurements with Cell Sorting by Flow Cytometry.

    Science.gov (United States)

    Servais; Courties; Lebaron; Troussellier

    1999-08-01

    > Abstract A new procedure to investigate the relationship between bacterial cell size and activity at the cellular level has been developed; it is based on the coupling of radioactive labeling of bacterial cells and cell sorting by flow cytometry after SYTO 13 staining. Before sorting, bacterial cells were incubated in the presence of tritiated leucine using a procedure similar to that used for measuring bacterial production by leucine incorporation and then stained with SYTO 13. Subpopulations of bacterial cells were sorted according to their average right-angle light scatter (RALS) and fluorescence. Average RALS was shown to be significantly related to the average biovolume. Experiments were performed on samples collected at different times in a Mediterranean seawater mesocosm enriched with nitrogen and phosphorus. At four sampling times, bacteria were sorted in two subpopulations (cells smaller and larger than 0.25 µm(3)). The results indicate that, at each sampling time, the growth rate of larger cells was higher than that of smaller cells. In order to confirm this tendency, cell sorting was performed on six subpopulations differing in average biovolume during the mesocosm follow-up. A clear increase of the bacterial growth rates was observed with increasing cell size for the conditions met in this enriched mesocosm.http://link.springer-ny.com/link/service/journals/00248/bibs/38n2p180.html

  2. Isolation and Flow Cytometry Analysis of Innate Lymphoid Cells from the Intestinal Lamina Propria.

    Science.gov (United States)

    Gronke, Konrad; Kofoed-Nielsen, Michael; Diefenbach, Andreas

    2017-01-01

    The intestinal mucosa constitutes the biggest surface area of the body. It is constantly challenged by bacteria, commensal and pathogenic, protozoa, and food-derived irritants. In order to maintain homeostasis, a complex network of signaling circuits has evolved that includes contributions of immune cells. In recent years a subset of lymphocytes, which belong to the innate immune system, has caught particular attention. These so-called innate lymphoid cells (ILC) reside within the lamina propria of the small and large intestines and rapidly respond to environmental challenges. They provide immunity to various types of infections but may also contribute to organ homeostasis as they produce factors acting on epithelial cells thereby enhancing barrier integrity. Here, we describe how these cells can be isolated from their environment and provide an in-depth protocol how to visualize the various ILC subsets by flow cytometry.

  3. A study of light scattering of mononuclear blood cells with scanning flow cytometry

    International Nuclear Information System (INIS)

    Zharinov, Alexey; Tarasov, Peter; Shvalov, Alexander; Semyanov, Konstantin; Bockstaele, Dirk R. van; Maltsev, Valeri

    2006-01-01

    This study describes the measurement of light scattering of human mononuclear blood cells, the development of an appropriate optical model for those cells, and solution of the inverse light-scattering problem. The angular dependency of light-scattering intensity of mononuclear blood cells was experimentally measured by means of scanning flow cytometry. A sphere consisting of several concentric homogeneous layers with different refractive indices was tested as an optical model for mononuclear blood cells. A five-layer model has given the best agreement between experimental and theoretical light-scattering profiles. The inverse light-scattering problem was solved for a five-layer model with an optimization procedure that allows one to retrieve cell parameters: cell size relates to the outer diameter of the fifth layer; size of the nucleus relates to the outer diameter of the third layer. Mean values of cell size, nuclear size, refractive indices of nucleus and cellular cytoplasm were determined for blood monocytes and lymphocytes

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

    Directory of Open Access Journals (Sweden)

    Hanna L Sladitschek

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

  5. Two new polyploid species closely related to Alnus glutinosa in Europe and North Africa - An analysis based on morphometry, karyology, flow cytometry and microsatellites

    Czech Academy of Sciences Publication Activity Database

    Vít, Petr; Douda, Jan; Krak, Karol; Havrdová, Alena; Mandák, Bohumil

    2017-01-01

    Roč. 66, č. 3 (2017), s. 567-583 ISSN 0040-0262 R&D Projects: GA ČR(CZ) GAP504/11/0402 Institutional support: RVO:67985939 Keywords : Alnus glutinosa * flow cytometry * Balkan Peninsula Subject RIV: EF - Botanics OBOR OECD: Plant sciences, botany Impact factor: 2.447, year: 2016

  6. Performance evaluation of a dual-flow recharge filter for improving groundwater quality.

    Science.gov (United States)

    Samuel, Manoj P; Senthilvel, S; Mathew, Abraham C

    2014-07-01

    A dual-flow multimedia stormwater filter integrated with a groundwater recharge system was developed and tested for hydraulic efficiency and pollutant removal efficiency. The influent stormwater first flows horizontally through the circular layers of planted grass and biofibers. Subsequently, the flow direction changes to a vertical direction so that water moves through layers of pebbles and sand and finally gets recharged to the deep aquifers. The media in the sequence of vegetative medium:biofiber to pebble:sand were filled in nine proportions and tested for the best performing combination. Three grass species, viz., Typha (Typha angustifolia), Vetiver (Chrysopogon zizanioides), and St. Augustine grass (Stenotaphrum secundatum), were tested as the best performing vegetative medium. The adsorption behavior of Coconut (Cocos nucifera) fiber, which was filled in the middle layer, was determined by a series of column and batch studies.The dual-flow filter showed an increasing trend in hydraulic efficiency with an increase in flowrate. The chemical removal efficiency of the recharge dual-flow filter was found to be very high in case of K+ (81.6%) and Na+ (77.55%). The pH normalizing efficiency and electrical conductivity reduction efficiency were also recorded as high. The average removal percentage of Ca2+ was moderate, while that of Mg2+ was very low. The filter proportions of 1:1 to 1:2 (plant:fiber to pebble:sand) showed a superior performance compared to all other proportions. Based on the estimated annual costs and returns, all the financial viability criteria (internal rate of return, net present value, and benefit-cost ratio) were found to be favorable and affordable to farmers in terms of investing in the developed filtration system.

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

    Science.gov (United States)

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

    2017-02-01

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

  8. Flow cytometry for rapid detection of Salmonella spp. in seed sprouts

    Directory of Open Access Journals (Sweden)

    Bledar Bisha

    2014-12-01

    Full Text Available Seed sprouts (alfalfa, mung bean, radish, etc. have been implicated in several recent national and international outbreaks of salmonellosis. Conditions used for sprouting are also conducive to the growth of Salmonella. As a result, this pathogen can quickly grow to very high cell densities during sprouting without any detectable organoleptic impact. Seed sprouts typically also support heavy growth (~108 CFU g−1 of a heterogeneous microbiota consisting of various bacterial, yeast, and mold species, often dominated by non-pathogenic members of the family Enterobacteriaceae. This heavy background may present challenges to the detection of Salmonella, especially if this pathogen is present in relatively low numbers. We combined DNA-based fluorescence in situ hybridization (FISH with flow cytometry (FCM for the rapid molecular detection of Salmonella enterica ser. Typhimurium in artificially contaminated alfalfa and other seed sprouts. Components of the assay included a set of cooperatively binding probes, a chemical blocking treatment intended to reduce non-specific background, and sample concentration via tangential flow filtration (TFF. We were able to detect S. Typhimurium in sprout wash at levels as low as 103 CFU ml−1 sprout wash (104 CFU g−1 sprouts against high microbial backgrounds (~108 CFU g−1 sprouts. Hybridization times were typically 30 min, with additional washing, but we ultimately found that S. Typhimurium could be readily detected using hybridization times as short as 2 min, without a wash step. These results clearly demonstrate the potential of combined DNA-FISH and FCM for rapid detection of Salmonella in this challenging food matrix and provide industry with a useful tool for compliance with sprout production standards proposed in the Food Safety Modernization Act (FSMA.

  9. Nuclear DNA content of the pigeon orchid (Dendrobium crumenatum Sw. with the analysis of flow cytometry

    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.

  10. Acceleration of Gas Flow Simulations in Dual-Continuum Porous Media Based on the Mass-Conservation POD Method

    KAUST Repository

    Wang, Yi

    2017-09-12

    Reduced-order modeling approaches for gas flow in dual-porosity dual-permeability porous media are studied based on the proper orthogonal decomposition (POD) method combined with Galerkin projection. The typical modeling approach for non-porous-medium liquid flow problems is not appropriate for this compressible gas flow in a dual-continuum porous media. The reason is that non-zero mass transfer for the dual-continuum system can be generated artificially via the typical POD projection, violating the mass-conservation nature and causing the failure of the POD modeling. A new POD modeling approach is proposed considering the mass conservation of the whole matrix fracture system. Computation can be accelerated as much as 720 times with high precision (reconstruction errors as slow as 7.69 × 10−4%~3.87% for the matrix and 8.27 × 10−4%~2.84% for the fracture).

  11. Acceleration of Gas Flow Simulations in Dual-Continuum Porous Media Based on the Mass-Conservation POD Method

    KAUST Repository

    Wang, Yi; Sun, Shuyu; Yu, Bo

    2017-01-01

    Reduced-order modeling approaches for gas flow in dual-porosity dual-permeability porous media are studied based on the proper orthogonal decomposition (POD) method combined with Galerkin projection. The typical modeling approach for non-porous-medium liquid flow problems is not appropriate for this compressible gas flow in a dual-continuum porous media. The reason is that non-zero mass transfer for the dual-continuum system can be generated artificially via the typical POD projection, violating the mass-conservation nature and causing the failure of the POD modeling. A new POD modeling approach is proposed considering the mass conservation of the whole matrix fracture system. Computation can be accelerated as much as 720 times with high precision (reconstruction errors as slow as 7.69 × 10−4%~3.87% for the matrix and 8.27 × 10−4%~2.84% for the fracture).

  12. Ablative dual-phase Erbium:YAG laser treatment of atrophy-related vaginal symptoms in post-menopausal breast cancer survivors omitting hormonal treatment.

    Science.gov (United States)

    Mothes, A R; Runnebaum, M; Runnebaum, I B

    2018-05-01

    First evaluation of dual-phase vaginal Er:YAG laser to omit hormonal treatment for atrophy-related symptoms in post-menopausal breast cancer survivors following prolapse surgery. Patients with a history of breast cancer at the time of surgery for pelvic organ prolapse were offered non-hormonal vaginal Er:YAG laser treatment when complaining of atrophy-related genitourinary syndrome of menopause. A single 10-min course of dual-phase protocol of pulsed Er:YAG laser (2940 nm, fractional ablative and thermal mode, fluence according to tissue thickness). Follow-up included subjective satisfaction, vaginal pH, vaginal health index (VHI), and complications after 6 weeks. A total of 16 breast cancer survivors (age 71 years, SD 7) had been seeking treatment for pelvic floor symptoms related to vaginal atrophy at follow-up visits after prolapse surgery. All ablative vaginal Er:YAG laser outpatient procedures were successfully completed, all patients returned to daily activities without a need for analgetic medication. Evaluation was performed after 8.3 (SD 2.5) weeks. Pre-laser VHI scored 16 (SD 4.6) and post-laser VHI 20 (SD 3) with p = 0.01. Patients were satisfied in 94% (n = 15) regarding symptom relief. Breast cancer survivors with atrophy-related complaints after pelvic floor surgery may benefit from vaginal application of this innovative dual protocol of Er:YAG laser technology as a non-hormonal treatment approach.

  13. Tolerance Induction of Temperature and Starvation with Tricalcium Phosphate on Preservation and Sporulation in Bacillus amyloliquefaciens Detected by Flow Cytometry.

    Science.gov (United States)

    Shahrokh Esfahani, Samaneh; Emtiazi, Giti; Shafiei, Rasoul; Ghorbani, Najmeh; Zarkesh Esfahani, Seyed Hamid

    2016-09-01

    The Bacillus species have many applications in the preparation of various enzymes, probiotic, biofertilizer, and biomarkers for which the survival of resting cells and spore formation under different conditions are important. In this study, water and saline along with different mineral substances such as calcium carbonate, calcium phosphate, and silica were used for the detection of survival and preservation of Bacillus amyloliquefaciens. The results showed intensive death of resting cells at 8 °C, but significant survival at 28 °C after one month. However, preservation by minerals significantly decreased the rate of death and induced sporulation at both the temperatures. The resting cells were maintained at room temperature (about 60 % of the initial population survived after a month) in the presence of tricalcium phosphate. The results showed that temperature has more effect on sporulation compare with starvation. The sporulation in normal saline at 28 °C was 70 times more than that at 8 °C; meanwhile, addition of tricalcium phosphate increases sporulation by 90 times. Also, the FTIR data showed the interaction of tricalcium phosphate with spores and resting cells. The discrimination of sporulation from non-sporulation state was performed by nucleic acid staining with thiazole orange and detected by flow cytometry. The flow cytometric studies confirmed that the rates of sporulation in pure water were significantly more at 28 °C. This is the first report on the detection of bacterial spore with thiazole orange by flow cytometry and also on the interaction of tricalcium phosphate with spores by FTIR analyses.

  14. Picometer-resolution dual-comb spectroscopy with a free-running fibre laser

    OpenAIRE

    Zhao, Xin; Hu, Guoqing; Zhao, Bofeng; Li, Cui; Pan, Yingling; Liu, Ya; Yasui, Takeshi; Zheng, Zheng

    2016-01-01

    Dual-comb spectroscopy utilizes two sets of comb lines with slightly different comb-tooth-spacings, and optical spectral information is acquired by measuring the radio-frequency beat notes between the sets of comb lines. It holds the promise as a real-time, high-resolution analytical spectroscopy tool for a range of applications. However, the stringent requirement on the coherence between comb lines from two separate lasers and the sophisticated control system to achieve that have confined th...

  15. Development of a Flow Cytometry-Based Method for Rapid Detection of Escherichia coli and Shigella Spp. Using an Oligonucleotide Probe

    Science.gov (United States)

    Xue, Yong; Wilkes, Jon G.; Moskal, Ted J.; Williams, Anna J.; Cooper, Willie M.; Nayak, Rajesh; Rafii, Fatemeh; Buzatu, Dan A.

    2016-01-01

    Standard methods to detect Escherichia coli contamination in food use the polymerase chain reaction (PCR) and agar culture plates. These methods require multiple incubation steps and take a long time to results. An improved rapid flow-cytometry based detection method was developed, using a fluorescence-labeled oligonucleotide probe specifically binding a16S rRNA sequence. The method positively detected 51 E. coli isolates as well as 4 Shigella species. All 27 non-E. coli strains tested gave negative results. Comparison of the new genetic assay with a total plate count (TPC) assay and agar plate counting indicated similar sensitivity, agreement between cytometry cell and colony counts. This method can detect a small number of E.coli cells in the presence of large numbers of other bacteria. This method can be used for rapid, economical, and stable detection of E. coli and Shigella contamination in the food industry and other contexts. PMID:26913737

  16. Development of a Flow Cytometry-Based Method for Rapid Detection of Escherichia coli and Shigella Spp. Using an Oligonucleotide Probe.

    Directory of Open Access Journals (Sweden)

    Yong Xue

    Full Text Available Standard methods to detect Escherichia coli contamination in food use the polymerase chain reaction (PCR and agar culture plates. These methods require multiple incubation steps and take a long time to results. An improved rapid flow-cytometry based detection method was developed, using a fluorescence-labeled oligonucleotide probe specifically binding a16S rRNA sequence. The method positively detected 51 E. coli isolates as well as 4 Shigella species. All 27 non-E. coli strains tested gave negative results. Comparison of the new genetic assay with a total plate count (TPC assay and agar plate counting indicated similar sensitivity, agreement between cytometry cell and colony counts. This method can detect a small number of E.coli cells in the presence of large numbers of other bacteria. This method can be used for rapid, economical, and stable detection of E. coli and Shigella contamination in the food industry and other contexts.

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

    Science.gov (United States)

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

    2017-10-01

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

  18. Dual solutions in boundary layer flow of Maxwell fluid over a porous shrinking sheet

    International Nuclear Information System (INIS)

    Bhattacharyya Krishnendu; Hayat Tasawar; Alsaedi Ahmed

    2014-01-01

    An analysis is carried out for dual solutions of the boundary layer flow of Maxwell fluid over a permeable shrinking sheet. In the investigation, a constant wall mass transfer is considered. With the help of similarity transformations, the governing partial differential equations (PDEs) are converted into a nonlinear self-similar ordinary differential equation (ODE). For the numerical solution of transformed self-similar ODE, the shooting method is applied. The study reveals that the steady flow of Maxwell fluid is possible with a smaller amount of imposed mass suction compared with the viscous fluid flow. Dual solutions for the velocity distribution are obtained. Also, the increase of Deborah number reduces the boundary layer thickness for both solutions. (electromagnetism, optics, acoustics, heat transfer, classical mechanics, and fluid dynamics)

  19. Diode-pumped orthogonally polarized dual-wavelength Nd3+:LaBO2MoO4 laser

    Science.gov (United States)

    Chen, Y. J.; Gong, X. H.; Lin, Y. F.; Huang, J. H.; Luo, Z. D.; Huang, Y. D.

    2013-08-01

    Polarized spectroscopic properties related to 1.07 μm laser operation of a 1.8 at.% Nd3+:LaBO2MoO4 crystal grown by the Czochralski method were investigated at room temperature. Using a 2.2-mm-thick, Z-cut Nd3+:LaBO2MoO4 crystal as gain medium, orthogonally polarized dual-wavelength laser at 1,068 and 1,074 nm was first realized in a plano-concave resonator end-pumped by a quasi-continuous-wave 795 nm diode laser. A total output peak power of 1.2 W with slope efficiency of 26 % around 1.07 μm was obtained. The influences of resonator length and pump power on output laser wavelength were also investigated.

  20. Unilateral uveitis masquerade syndrome caused by diffuse large B-cell lymphoma diagnosed using multiparametric flow cytometry of the aqueous humor.

    Science.gov (United States)

    Monsalvo, Silvia; Serrano, Cristina; Prieto, Elena; Fernández-Sanz, Guillermo; Puente, Maria-Camino; Rodriguez-Pinilla, Maria; Garcia Raso, Aranzazu; Llamas, Pilar; Cordoba, Raul

    2017-07-01

    The uveitis masquerade syndromes (UMS) are a group of ocular diseases that may mimic chronic intraocular inflammation. Many malignant entities such as non-Hodgkin's lymphomas may masquerade as uveitis. We report a case of an HIV-positive patient with masquerade syndrome presenting unilateral uveitis. 45-year-old Caucasian man with a diagnosis of diffuse large B-cell lymphoma (DLBCL). The patient was diagnosed by a biopsy of an abdominal mass which showed fragments of gastric mucosa with diffuse growth of neoplastic cells. At diagnosis, the patient suffered from unilateral blurring of vision and a sudden decrease of left-eye visual acuity. A slit-lamp examination of the left eye revealed a diagnosis of anterior uveitis. The patient exhibited no signs of posterior uveitis. An anterior-chamber paracentesis was performed and analyzed by multiparameter flow cytometry (MFC), showing cells CD45, CD19, CD20, CD22, and CD38 positives, and moderate expression of CD10 with kappa light chain restriction, showing a monoclonal B-cell population. The patient received CHOP-R with intrathecal methotrexate followed by consolidation high dose methotrexate obtaining a complete response which is ongoing. Differential diagnosis between chronic uveitis and ocular lymphoma may be challenging. We advocate anterior-chamber paracentesis in cases of refractory uveitis in patients with hematologic malignancies. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

  1. Flow Cytometry Detection of Infectious Rotaviruses in Environmental and Clinical Samples

    Science.gov (United States)

    Abad, F. Xavier; Pintó, Rosa M.; Bosch, Albert

    1998-01-01

    A method for the detection of infectious human rotaviruses based on infection of CaCo-2 cells and detection of infected cells by indirect immunofluorescence and flow cytometry (IIF-FC) has been developed. The technique was validated by performing a seminested reverse transcription-PCR assay with sorted cell populations. The efficiency of the procedure has been compared with that of the standard method of infection of MA104 cells and ulterior detection by IIF and optical microscopy (IIF-OM) and with that of infection of MA104 cells and detection by IIF-FC. The limit of sensitivity for the detection of the cell-adapted strain Itor P13, expressed as the most probable number of cytopathogenic units, was established as 200 and 2 for MA104 and CaCo-2 cells, respectively, by the IIF-FC method. The ratio of infectious virus particles to total virus particles for a wild-type rotavirus was determined to be 1/2 × 106 and 1/2 × 104 for IIF-OM with MA104 cells and IIF-FC with CaCo-2 cells, respectively. The use of IIF-FC with CaCo-2 cells was tested with fecal and water samples and proved to be more effective than the standard procedure for rotavirus detection. PMID:9647805

  2. Melt flow characteristics in gas-assisted laser cutting

    Indian Academy of Sciences (India)

    R. Narasimhan (Krishtel eMaging) 1461 1996 Oct 15 13:05:22

    industrial applications. In this process the cut ... Under these assumptions the boundary layer theory flow over. Laser beam ... a flat plate can be applied to estimate the distance after which the flow separation takes place by calculating the ...

  3. Clinical cytometry and progress in HLA antibody detection.

    Science.gov (United States)

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

    2011-01-01

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

  4. Continuous-wave single-frequency laser with dual wavelength at 1064 and 532 nm.

    Science.gov (United States)

    Zhang, Chenwei; Lu, Huadong; Yin, Qiwei; Su, Jing

    2014-10-01

    A continuous-wave high-power single-frequency laser with dual-wavelength output at 1064 and 532 nm is presented. The dependencies of the output power on the transmission of the output coupler and the phase-matching temperature of the LiB(3)O(5) (LBO) crystal are studied. An output coupler with transmission of 19% is used, and the temperature of LBO is controlled to the optimal phase-matching temperature of 422 K; measured maximal output powers of 33.7 W at 1064 nm and of 1.13 W at 532 nm are obtained with optical-optical conversion efficiency of 45.6%. The laser can be single-frequency operated stably and mode-hop-free, and the measured frequency drift is less than 15 MHz in 1 min. The measured Mx2 and My2 for the 1064 nm laser are 1.06 and 1.09, respectively. The measured Mx2 and My2 for the 532 nm laser are 1.12 and 1.11, respectively.

  5. Compact 84 GHz passive mode-locked fiber laser using dual-fiber coupled fused-quartz microresonator

    Science.gov (United States)

    Liu, Tze-An; Hsu, Yung; Chow, Chi-Wai; Chuang, Yi-Chen; Ting, Wei-Jo; Wang, Bo-Chun; Peng, Jin-Long; Chen, Guan-Hong; Chang, Yuan-Chia

    2017-10-01

    We propose and demonstrate a compact and portable-size 84-GHz passive mode-locked fiber laser, in which a dual-fiber coupled fused-quartz microresonator is employed as the intracavity optical comb filter as well as the optical nonlinear material for optical frequency comb generation. About eight coherent optical tones can be generated in the proposed fiber laser. The 20-dB bandwidth is larger than 588 GHz. The full-width half-maximum pulse-width of the proposed laser is 2.5 ps. We also demonstrate the feasibility of using the proposed passive mode-locked fiber laser to carry a 5-Gbit/s on-off-keying signal and transmit over 20-km standard single mode fiber. A 7% forward error correction requirement can be achieved, showing the proposed fiber laser can be a potential candidate for fiber-wireless applications.

  6. Measurements of gas velocity in supersonic flow using a laser beam

    International Nuclear Information System (INIS)

    Airoldi, V.J.T.; Santos, R. dos

    1982-01-01

    A study of measurements of supersonic velocities in a wind tunnel using a laser beam was performed. Techniques using lasers are most suitable because they do not disturb the gas flow. This work presents the technique entitled as fringe technique. It works using interference patterns due to two perpendicular laser beams crossing the sample (i.e. the gas flow). Experimental results are compared with other usual techniques. (R.S.)

  7. Panel development for multicolor flow-cytometry testing of proliferation and immunophenotype in hMSCs.

    Science.gov (United States)

    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.

  8. Relationship between conventional culture and flow cytometry for the diagnosis of urinary tract infection.

    Science.gov (United States)

    García-Coca, Marta; Gadea, Ignacio; Esteban, Jaime

    2017-06-01

    Urine culture is the gold standard for the diagnosis of urinary tract infections (UTI). The use of flow cytometry analyzers (FCA) prior to culture allows for the quantification and recognition of cell components in urine to be automated and makes it possible to relate these data to the urine pathogens subsequently identified in cultures. Urine samples were assessed with the Sysmex UF-1000i analyzer. Those that met the criteria for culture (> 25 leukocytes/μL or > 385 bacteria/μL) were subjected to quantitative urine culture on chromogenic agar. Counts of red blood cells (RBC), white blood cells (WBC), epithelial cells (EC), and the kind of microorganisms identified in cultures were evaluated. A total of 17,483 samples were processed by FCA. Of these, 9057 met the criteria for culture. Urine cultures were reduced by 48.2%. The most common urine pathogen was Escherichia coli (60.3%). Negative urine cultures were significantly (p flow cytometer for screening urine samples allows for a reduction in the number of urine cultures. WBC values correlate well with the main urine pathogens related to UTI. The results observed for Enterococcus spp. suggest a low impact of these pathogens as a cause of UTI.

  9. Synthetic Aperture Flow Imaging Using a Dual Beamformer Approach

    DEFF Research Database (Denmark)

    Li, Ye

    Color flow mapping systems have become widely used in clinical applications. It provides an opportunity to visualize the velocity profile over a large region in the vessel, which makes it possible to diagnose, e.g., occlusion of veins, heart valve deficiencies, and other hemodynamic problems....... However, while the conventional ultrasound imaging of making color flow mapping provides useful information in many circumstances, the spatial velocity resolution and frame rate are limited. The entire velocity distribution consists of image lines from different directions, and each image line...... on the current commercial ultrasound scanner. The motivation for this project is to develop a method lowering the amount of calculations and still maintaining beamforming quality sufficient for flow estimation. Synthetic aperture using a dual beamformer approach is investigated using Field II simulations...

  10. A highly stable and switchable dual-wavelength laser using coupled microfiber Mach-Zehnder interferometer as an optical filter

    Science.gov (United States)

    Jasim, A. A.; Ahmad, H.

    2017-12-01

    The generation and switching of dual-wavelength laser based on compact coupled microfiber Mach-Zehnder interferometer (CM-MZI) is reported. The CM-MZI is constructed by overlapping two portions of a single tapered optical fiber which has a diameter of 9 μm as to create multi-mode interference and also to produce spatial mode beating as to suppress mode competition in the homogeneous gain medium. The system is able to generate a dual-wavelength laser output that can be switched with the aid of the polarization rotation technique. Four dual-wavelength oscillation pairs are obtained from the interference fringe peaks of the CM-MZI comb filter with a switched channel spacing of 1.5 nm, 3.0 nm, and 6.0 nm. The wavelength spacing is stable at different pump powers. The lasing wavelength has a 3-dB linewidth of about 30 pm and peak-to-floor ration of about 55 dB at a pump power of 38 mW.

  11. Microfluidics and photonics for Bio-System-on-a-Chip: a review of advancements in technology towards a microfluidic flow cytometry chip.

    Science.gov (United States)

    Godin, Jessica; Chen, Chun-Hao; Cho, Sung Hwan; Qiao, Wen; Tsai, Frank; Lo, Yu-Hwa

    2008-10-01

    Microfluidics and photonics come together to form a field commonly referred to as 'optofluidics'. Flow cytometry provides the field with a technology base from which both microfluidic and photonic components be developed and integrated into a useful device. This article reviews some of the more recent developments to familiarize a reader with the current state of the technologies and also highlights the requirements of the device and how researchers are working to meet these needs.

  12. Combining Cell Type-Restricted Adenoviral Targeting with Immunostaining and Flow Cytometry to Identify Cells-of-Origin of Lung Cancer.

    Science.gov (United States)

    Best, Sarah A; Kersbergen, Ariena; Asselin-Labat, Marie-Liesse; Sutherland, Kate D

    2018-01-01

    Lung cancers display considerable intertumoral heterogeneity, leading to the classification of distinct tumor subtypes. Our understanding of the genetic aberrations that underlie tumor subtypes has been greatly enhanced by recent genomic sequencing studies and state-of-the-art gene targeting technologies, highlighting evidence that distinct lung cancer subtypes may be derived from different "cells-of-origin". Here, we describe the intra-tracheal delivery of cell type-restricted Ad5-Cre viruses into the lungs of adult mice, combined with immunohistochemical and flow cytometry strategies for the detection of lung cancer-initiating cells in vivo.

  13. Tunable and switchable dual-wavelength passively mode-locked Bi-doped all-fiber ring laser based on nonlinear polarization rotation

    International Nuclear Information System (INIS)

    Luo, A-P; Luo, Z-C; Xu, W-C; Dvoyrin, V V; Mashinsky, V M; Dianov, E M

    2011-01-01

    We demonstrate a tunable and switchable dual-wavelength passively mode-locked Bi-doped all-fiber ring laser by using nonlinear polarization rotation (NPR) technique. Exploiting the spectral filtering effect caused by the combination of the polarizer and intracavity birefringence, the wavelength separation of dual-wavelength mode-locked pulses can be flexibly tuned between 2.38 and 20.45 nm. Taking the advantage of NPR-induced intensity-dependent loss to suppress the mode competition, the stable dual-wavelength pulses output is obtained at room temperature. Moreover, the dual-wavelength switchable operation is achieved by simply rotating the polarization controllers (PCs)

  14. Hydrostatic Pressure Enhances Vital Staining with Carboxyfluorescein or Carboxydichlorofluorescein in Saccharomyces cerevisiae: Efficient Detection of Labeled Yeasts by Flow Cytometry

    Science.gov (United States)

    Abe, Fumiyoshi

    1998-01-01

    The extent of intracellular accumulation of the fluorescent dye carboxyfluorescein or carboxydichlorofluorescein (CDCF) in Saccharomyces cerevisiae was found to be increased 5- to 10-fold under a nonlethal hydrostatic pressure of 30 to 50 MPa. This observation was confirmed by analysis of individual labeled cells by flow cytometry. The pressure-induced enhancement of staining with CDCF required d-glucose and was markedly inhibited by 2-deoxy-d-glucose, suggesting that glucose metabolism has a role in the process. PMID:9501452

  15. La citometría de flujo en el estudio de tumores mediastinales ricos en elementos linfoides Flow cytometry for the study of mediastinal tumors with abundant lymphoid elements

    Directory of Open Access Journals (Sweden)

    Gerardo Vides Almonacid

    2008-02-01

    Full Text Available El mediastino anterior es un sitio frecuente de localización de tumores ricos en elementos linfoides. La identificación correcta de cada entidad es de importancia en el tratamiento de los pacientes. En ocasiones puede plantearse el diagnóstico diferencial entre timoma y linfoma linfoblástico con fenotipo de precursor T (LLB-T. La Citometría de Flujo (CF es una técnica complementaria útil para estos tumores de la cual se obtiene información cualitativa y cuantitativa. Revisamos 38 tumores mediastinales que tenían estudio de CF. Además comparamos los resultados de CF de timomas y tejido tímico normal con 42 casos de LLB-T de otras localizaciones anatómicas. De los 38 tumores mediastinales 6 eran lesiones benignas, 9 linfomas difusos de células grandes con fenotipo B (LDCG-B, 10 linfomas de Hodgkin (LH, 11 timomas y 2 LLB-T. En 24 casos la CF aportó información positiva, definiendo el inmunofenotipo de las células linfoides neoplásicas, o los linfocitos característicos que acompañan a los timomas. La CF en los 10 casos de LH y en 4 lesiones benignas permitió descartar otros tipos de linfoma (LDCG-B, LLB-T, etc.. Las marcaciones para CD3, CD4 y CD8 fueron las más útiles para el diagnóstico diferencial entre timomas y LLB-T. En conclusión, la CF es una técnica complementaria de utilidad que aporta información en lesiones mediastinales de manera rápida, requiriendo cantidades pequeñas de material, tanto para el diagnóstico inicial como para el monitoreo de estas enfermedades.The anterior mediastinum is a common site of tumors with abundant lymphoid elements. Flow cytometry is a useful complementary technique to analyze this type of tumors, which provides qualitative and quantitative information. A differential diagnosis can be sometimes made between thymoma and precursor T-lymphoblastic lymphoma (T-LBL. Correct identification is of utmost importance for patient treatment. A total of 38 mediastinal tumors were analyzed, and

  16. Mapping cell populations in flow cytometry data for cross‐sample comparison using the Friedman–Rafsky test statistic as a distance measure

    Science.gov (United States)

    Hsiao, Chiaowen; Liu, Mengya; Stanton, Rick; McGee, Monnie; Qian, Yu

    2015-01-01

    Abstract Flow cytometry (FCM) is a fluorescence‐based single‐cell experimental technology that is routinely applied in biomedical research for identifying cellular biomarkers of normal physiological responses and abnormal disease states. While many computational methods have been developed that focus on identifying cell populations in individual FCM samples, very few have addressed how the identified cell populations can be matched across samples for comparative analysis. This article presents FlowMap‐FR, a novel method for cell population mapping across FCM samples. FlowMap‐FR is based on the Friedman–Rafsky nonparametric test statistic (FR statistic), which quantifies the equivalence of multivariate distributions. As applied to FCM data by FlowMap‐FR, the FR statistic objectively quantifies the similarity between cell populations based on the shapes, sizes, and positions of fluorescence data distributions in the multidimensional feature space. To test and evaluate the performance of FlowMap‐FR, we simulated the kinds of biological and technical sample variations that are commonly observed in FCM data. The results show that FlowMap‐FR is able to effectively identify equivalent cell populations between samples under scenarios of proportion differences and modest position shifts. As a statistical test, FlowMap‐FR can be used to determine whether the expression of a cellular marker is statistically different between two cell populations, suggesting candidates for new cellular phenotypes by providing an objective statistical measure. In addition, FlowMap‐FR can indicate situations in which inappropriate splitting or merging of cell populations has occurred during gating procedures. We compared the FR statistic with the symmetric version of Kullback–Leibler divergence measure used in a previous population matching method with both simulated and real data. The FR statistic outperforms the symmetric version of KL‐distance in distinguishing

  17. Optofluidic fluorescent imaging cytometry on a cell phone.

    Science.gov (United States)

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

    2011-09-01

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

  18. Interlaboratory assessment of mitotic index by flow cytometry confirms superior reproducibility relative to microscopic scoring.

    Science.gov (United States)

    Roberts, D J; Spellman, R A; Sanok, K; Chen, H; Chan, M; Yurt, P; Thakur, A K; DeVito, G L; Murli, H; Stankowski, L F

    2012-05-01

    A flow cytometric procedure for determining mitotic index (MI) as part of the metaphase chromosome aberrations assay, developed and utilized routinely at Pfizer as part of their standard assay design, has been adopted successfully by Covance laboratories. This method, using antibodies against phosphorylated histone tails (H3PS10) and nucleic acid stain, has been evaluated by the two independent test sites and compared to manual scoring. Primary human lymphocytes were treated with cyclophosphamide, mitomycin C, benzo(a)pyrene, and etoposide at concentrations inducing dose-dependent cytotoxicity. Deming regression analysis indicates that the results generated via flow cytometry (FCM) were more consistent between sites than those generated via microscopy. Further analysis using the Bland-Altman modification of the Tukey mean difference method supports this finding, as the standard deviations (SDs) of differences in MI generated by FCM were less than half of those generated manually. Decreases in scoring variability owing to the objective nature of FCM, and the greater number of cells analyzed, make FCM a superior method for MI determination. In addition, the FCM method has proven to be transferable and easily integrated into standard genetic toxicology laboratory operations. Copyright © 2012 Wiley Periodicals, Inc.

  19. FLOCK cluster analysis of mast cell event clustering by high-sensitivity flow cytometry predicts systemic mastocytosis.

    Science.gov (United States)

    Dorfman, David M; LaPlante, Charlotte D; Pozdnyakova, Olga; Li, Betty

    2015-11-01

    In our high-sensitivity flow cytometric approach for systemic mastocytosis (SM), we identified mast cell event clustering as a new diagnostic criterion for the disease. To objectively characterize mast cell gated event distributions, we performed cluster analysis using FLOCK, a computational approach to identify cell subsets in multidimensional flow cytometry data in an unbiased, automated fashion. FLOCK identified discrete mast cell populations in most cases of SM (56/75 [75%]) but only a minority of non-SM cases (17/124 [14%]). FLOCK-identified mast cell populations accounted for 2.46% of total cells on average in SM cases and 0.09% of total cells on average in non-SM cases (P < .0001) and were predictive of SM, with a sensitivity of 75%, a specificity of 86%, a positive predictive value of 76%, and a negative predictive value of 85%. FLOCK analysis provides useful diagnostic information for evaluating patients with suspected SM, and may be useful for the analysis of other hematopoietic neoplasms. Copyright© by the American Society for Clinical Pathology.

  20. Development of electrical excited CO2-laser with transversal gas flow as well an axial flowed CO2-laser for material treatment, in particular for cutting

    International Nuclear Information System (INIS)

    Wollermann-Windgasse, R.; Ackermann, F.

    1987-04-01

    The project describes the development of a new generation of CO 2 -lasers using high frequency discharge (13.56 MHz) for laser excitation by capacitive dielectrical input. HF-excitation has a lot of advantages compared with direct current technology, these are higher electrical input power into the plasma, better homogeneity and stability of discharges. In addition to this, HF-excitation shows excellent possibilities for pulsing and modulation. As a result of this, there are compact powerful laser systems with the possibility of scaling up to the multi-kW-range. The examination included fast transversal flowed as well as fast axial flowed systems. In the end of this project development prototypes with laser output power of 1000 W, 1500 W and 3000 W were available. Detailed attempts of application show that these lasers on grounds of excellent laser output quality and controlability of laser power specifically to each process make possible new ways for material treatment by laser. (orig./HP) [de

  1. Plasma measurement by feedback-stabilized dual-beam laser interferometer

    International Nuclear Information System (INIS)

    Yasuda, Akio; Kawahata, Kazuo; Kanai, Yasubumi.

    1982-03-01

    The plasma density in a dynamic magneto arcjet is measured by a stabilized dual-beam laser interferometer proposed by the authors. The fringe shift for a 0.63 μm beam of He-Ne laser is used to stabilize the interferometer against the effect of mechanical vibration by means of a feedback controlled speaker coil, while the other beam of 3.39 μm, for which the effect of mechanical vibrations is excluded, is used to measure plasma density. Stability of --1/500 of one fringe for 0.63 μm is obtained during a long period for frequencies lower than a few Hertz. Stability for higher frequencies, which determines the accuracy of the present measurement, is limited to --1/30 of one fringe for 0.63 μm, which corresponds to --1/200 of one fringe and a line electron density of --1.5 x 10 14 cm - 2 for 3.39 μm, by acoustic noise picked up by the speaker coil. The advantage of this technique over the single-laser technique is that the frequency response of the interferometer extends down to zero frequency. Since the effect of the neutral gas background is practically reduced to zero, the present interferometer is to be applied advantageously to the diagnostics of the plasma produced in high pressure gases. (author)

  2. Emission Characteristics of Laser-Induced Plasma Using Collinear Long and Short Dual-Pulse Laser-Induced Breakdown Spectroscopy (LIBS).

    Science.gov (United States)

    Wang, Zhenzhen; Deguchi, Yoshihiro; Liu, Renwei; Ikutomo, Akihiro; Zhang, Zhenzhen; Chong, Daotong; Yan, Junjie; Liu, Jiping; Shiou, Fang-Jung

    2017-09-01

    Collinear long and short dual-pulse laser-induced breakdown spectroscopy (DP-LIBS) was employed to clarify the emission characteristics from laser-induced plasma. The plasma was sustained and became stable by the long pulse-width laser with the pulse width of 60 μs under free running (FR) conditions as an external energy source. Comparing the measurement results of stainless steel in air using single-pulse LIBS (SP-LIBS) and DP-LIBS, the emission intensity was markedly enhanced using DP-LIBS. The temperature of plasma induced by DP-LIBS was maintained at a higher temperature under different gate delay time and short pulse-width laser power conditions compared with those measured using short SP-LIBS. Moreover, the variation rates of plasma temperatures measured using DP-LIBS were also lower. The superior detection ability was verified by the measurement of aluminum sample in water. The spectra were clearly detected using DP-LIBS, whereas it cannot be identified using SP-LIBS of short and long pulse widths. The effects of gate delay time and short pulse-width laser power were also discussed. These results demonstrate the feasibility and enhanced detection ability of the proposed collinear long and short DP-LIBS method.

  3. Flow cytometry in the diagnosis of myelodysplastic syndromes (MDS) and the value of myeloid nuclear differentiation antigen (MNDA).

    Science.gov (United States)

    Bellos, Frauke; Kern, Wolfgang

    2014-09-25

    Background: Confirming diagnosis of myelodysplastic syndromes (MDS) is often challenging. Standard diagnostic methods are cytomorphology (CM) and cytogenetics (CG). Multiparameter flow cytometry (MFC) is upcoming in MDS diagnostic work up, comparability and investigator experience are critical. Myeloid nuclear differentiation antigen (MNDA) in myelomonocytic cells might be expressed more weakly in patients with MDS. The analysis of MNDA may thus improve diagnostic capabilities of MFC in MDS. Methods: Staining methods and antibody combinations for MFC in MDS are outlined, giving details for interpretation of results in regard to dyspoiesis. MFC results are correlated with CM and CG and with survival data. Use of myeloid nuclear differentiation antigen (MNDA) in MDS diagnostics was evaluated in 239 patients with MDS, AML, other cytopenic conditions and in 30 negative controls. Results: Strong correlation between findings in CM and MFC was found; MFC results correlated well with those of CG. Patients with higher grades of dysplasia in MFC had shorter overall survival. Percentages of granulocytes and monocytes with diminished MNDA expression (%dimG, %dimM) were higher in patients with MDS and AML. Mean fluorescence intensity (MFI) of MNDA in monocytes was lower in MDS and AML. Cut-off values for %dimG (12%) and %dimM (22%) as well as for MFI in monocytes (72) were defined discriminating between MDS and non-MDS. Conclusion: MFC adds significant information on dyspoiesis in the diagnostic work up for MDS and provides prognostic information. MNDA expression can be assessed by MFC and may facilitate evaluation of dyspoiesis when added to MDS MFC panels. © 2014 Clinical Cytometry Society. Copyright © 2014 Clinical Cytometry Society.

  4. Flexible temperature and flow sensor from laser-induced graphene

    KAUST Repository

    Marengo, Marco

    2017-12-25

    Herein we present a flexible temperature sensor and a flow speed sensor based on laser-induced graphene. The main benefits arise from peculiar electrical, thermal and mechanical performances of the material thus obtained, along with a cheap and simple fabrication process. The temperature sensor is a negative temperature coefficient thermistor with non-linear response typical of semi-metals. The thermistor shows a 4% decrease of the resistance in a temperature range of 20–60 °C. The flow sensor exploits the piezoresistive properties of laser-induced graphene and can be used both in gaseous and liquid media thanks to a protective polydimethylsiloxane coating. Main characteristics are ultra-fast response and versatility in design offered by the laser technology.

  5. Tunable and switchable dual-wavelength single polarization narrow linewidth SLM erbium-doped fiber laser based on a PM-CMFBG filter.

    Science.gov (United States)

    Yin, Bin; Feng, Suchun; Liu, Zhibo; Bai, Yunlong; Jian, Shuisheng

    2014-09-22

    A tunable and switchable dual-wavelength single polarization narrow linewidth single-longitudinal-mode (SLM) erbium-doped fiber (EDF) ring laser based on polarization-maintaining chirped moiré fiber Bragg grating (PM-CMFBG) filter is proposed and demonstrated. For the first time as we know, the CMFBG inscribed on the PM fiber is applied for the wavelength-tunable and-switchable dual-wavelength laser. The PM-CMFBG filter with ultra-narrow transmission band (0.1 pm) and a uniform polarization-maintaining fiber Bragg grating (PM-FBG) are used to select the laser longitudinal mode. The stable single polarization SLM operation is guaranteed by the PM-CMFBG filter and polarization controller. A tuning range of about 0.25 nm with about 0.075 nm step is achieved by stretching the uniform PM-FBG. Meanwhile, the linewidth of the fiber laser for each wavelength is approximate 6.5 and 7.1 kHz with a 20 dB linewidth, which indicates the laser linewidth is approximate 325 Hz and 355 Hz FWHM.

  6. Dual fluorescence and laser emissions from fluorescein-Na and eosin-B

    International Nuclear Information System (INIS)

    Math, N.N.; Naik, L.R.; Suresh, H.M.; Inamdar, S.R.

    2006-01-01

    Dual laser emissions were observed from fluorescein-Na and eosin-B in ethanolic solutions individually in the concentration range from 10 -2 to 10 -3 mol dm -3 under N 2 laser excitation. The first compound was found to lase at two distinct regions with wavelength maxima around 540, 550 nm, while the second one around 558, 574 nm. Steady-state absorption, fluorescence excitation, fluorescence polarization, fluorescence emission and decays of the dyes in various solvents under varying conditions of excitation and detection systems were carried out to identify the nature of the emitting species responsible for laser emissions in two distinct regions. Both the dyes exhibited concentration and excitation wavelength dependence of fluorescence and the effects were found to be more pronounced in binary solution. The fluorescence decays of dyes were monoexponential in ethanol, while in some other solvents used, the decays showed biexponential behavior. The absorption and excitation studies using thin layers of solutions revealed the formation of dimers with the dye concentration around 1x10 -3 mol dm -3 . Fluorescence polarization and decay studies confirmed the presence of dimers. The two laser bands observed in the shorter and longer wavelengths were respectively ascribed to monomeric and dimeric species

  7. Dual fluorescence and laser emissions from fluorescein-Na and eosin-B

    Energy Technology Data Exchange (ETDEWEB)

    Math, N.N. [Laser Spectroscopy (DRDO/KU) Programme, Department of Physics, Karnatak University, Dharwad 580 003 (India)]. E-mail: nnm31@rediffmail.com; Naik, L.R. [Laser Spectroscopy (DRDO/KU) Programme, Department of Physics, Karnatak University, Dharwad 580 003 (India); Suresh, H.M. [Laser Spectroscopy (DRDO/KU) Programme, Department of Physics, Karnatak University, Dharwad 580 003 (India); Inamdar, S.R. [Laser Spectroscopy (DRDO/KU) Programme, Department of Physics, Karnatak University, Dharwad 580 003 (India)

    2006-12-15

    Dual laser emissions were observed from fluorescein-Na and eosin-B in ethanolic solutions individually in the concentration range from 10{sup -2} to 10{sup -3} mol dm{sup -3} under N{sub 2} laser excitation. The first compound was found to lase at two distinct regions with wavelength maxima around 540, 550 nm, while the second one around 558, 574 nm. Steady-state absorption, fluorescence excitation, fluorescence polarization, fluorescence emission and decays of the dyes in various solvents under varying conditions of excitation and detection systems were carried out to identify the nature of the emitting species responsible for laser emissions in two distinct regions. Both the dyes exhibited concentration and excitation wavelength dependence of fluorescence and the effects were found to be more pronounced in binary solution. The fluorescence decays of dyes were monoexponential in ethanol, while in some other solvents used, the decays showed biexponential behavior. The absorption and excitation studies using thin layers of solutions revealed the formation of dimers with the dye concentration around 1x10{sup -3} mol dm{sup -3}. Fluorescence polarization and decay studies confirmed the presence of dimers. The two laser bands observed in the shorter and longer wavelengths were respectively ascribed to monomeric and dimeric species.

  8. Effect of pump-beam conditions on dual polarization oscillations in a microchip Nd:GdVO{sub 4} laser

    Energy Technology Data Exchange (ETDEWEB)

    Lin, C-C; Jiang, I-M [Department of Physics, National Sun Yat-Sen University, Kaohsiung 804, Taiwan (China); Ko, J-Y; Tsai, K-T; Cheng, Y-T; Ho, M-C, E-mail: jyko@nknucc.nknu.edu.t [Department of Physics, National Kaohsiung Normal University, Kaohsiung 824, Taiwan (China)

    2009-08-28

    This study investigated the input-output characteristics of a laser-diode-end-pumped microchip Nd:GdVO{sub 4} laser under different pump-beam focusing conditions by varying the magnifications of the microscope objective lenses and pump-beam positions on a chip. Dual-polarization oscillations were generated in the entire pump region using pumping conditions associated with different temperature gradients.

  9. CW and AO Q-switched operation of a dual-crystal Tm, Ho:GdVO4 laser pumped by two diodes

    International Nuclear Information System (INIS)

    Li, L J; Bai, Y F; Liu, Y W; He, Z L; Wang, J; Yao, B Q; Zhou, S; Xing, M N

    2013-01-01

    Continuous wave (CW) mode and acousto-optic (AO) Q-switched mode operation of a dual-crystal Tm, Ho:GdVO 4 laser is reported. The dual-crystal Tm, Ho:GdVO 4 laser with output wavelength of 2.05 μm was pumped by two laser diodes (LDs). The Tm, Ho:GdVO 4 crystals were cooled by liquid nitrogen and pumped by two fiber-coupled LDs with a center output wavelength of 801.0 nm. A 20.5 W output power was obtained at a 255 mm physical cavity length in CW mode operation, and a 19.6 W average power was obtained at a pulse repetition frequency (PRF) of 10 kHz with a 19 ns pulse duration. Also, the efficiency loss of the laser is not more than 4.4% from CW mode to Q-switch mode, and the M 2 factor, which is measured by the traveling knife-edge method, does not exceed 1.2. (paper)

  10. Application of a flow generated by IR laser and AC electric field in micropumping and micromixing

    International Nuclear Information System (INIS)

    Nakano, M; Mizuno, A

    2008-01-01

    In this paper, it is described that measurement of fluid flow generated by simultaneous operation of an infrared (IR) laser and AC electric field in a microfabricated channel. When an IR laser (1026 nm) was focused under an intense AC electric field, a circulating flow was generated around the laser focus. The IR laser and the electric field generate two flow patterns of the electrohydrodynamicss. When the laser focus is placed at the center of the gap between electrodes, the flow pattern is parallel to the AC electric field toward electrodes from the centre. On the other hand, when the laser focus is placed close to one of the electrodes, one directional flow is generated. First flow pattern can be used as a micromixer and the second one as a micropump. Flow velocity profiles of the two flow patterns were measured as a function of the laser power, intensity of the AC electric field and AC frequency.

  11. Avoiding false positive antigen detection by flow cytometry on blood cell derived microparticles: the importance of an appropriate negative control.

    Directory of Open Access Journals (Sweden)

    Emerence Crompot

    Full Text Available Microparticles (MPs, also called microvesicles (MVs are plasma membrane-derived fragments with sizes ranging from 0.1 to 1μm. Characterization of these MPs is often performed by flow cytometry but there is no consensus on the appropriate negative control to use that can lead to false positive results.We analyzed MPs from platelets, B-cells, T-cells, NK-cells, monocytes, and chronic lymphocytic leukemia (CLL B-cells. Cells were purified by positive magnetic-separation and cultured for 48h. Cells and MPs were characterized using the following monoclonal antibodies (CD19,20 for B-cells, CD3,8,5,27 for T-cells, CD16,56 for NK-cells, CD14,11c for monocytes, CD41,61 for platelets. Isolated MPs were stained with annexin-V-FITC and gated between 300nm and 900nm. The latex bead technique was then performed for easy detection of MPs. Samples were analyzed by Transmission (TEM and Scanning Electron microscopy (SEM.Annexin-V positive events within a gate of 300-900nm were detected and defined as MPs. Our results confirmed that the characteristic antigens CD41/CD61 were found on platelet-derived-MPs validating our technique. However, for MPs derived from other cell types, we were unable to detect any antigen, although they were clearly expressed on the MP-producing cells in the contrary of several data published in the literature. Using the latex bead technique, we confirmed detection of CD41,61. However, the apparent expression of other antigens (already deemed positive in several studies was determined to be false positive, indicated by negative controls (same labeling was used on MPs from different origins.We observed that mother cell antigens were not always detected on corresponding MPs by direct flow cytometry or latex bead cytometry. Our data highlighted that false positive results could be generated due to antibody aspecificity and that phenotypic characterization of MPs is a difficult field requiring the use of several negative controls.

  12. Flow cytometric characterization of cerebrospinal fluid cells.

    Science.gov (United States)

    de Graaf, Marieke T; de Jongste, Arjen H C; Kraan, Jaco; Boonstra, Joke G; Sillevis Smitt, Peter A E; Gratama, Jan W

    2011-09-01

    Flow cytometry facilitates the detection of a large spectrum of cellular characteristics on a per cell basis, determination of absolute cell numbers and detection of rare events with high sensitivity and specificity. White blood cell (WBC) counts in cerebrospinal fluid (CSF) are important for the diagnosis of many neurological disorders. WBC counting and differential can be performed by microscopy, hematology analyzers, or flow cytometry. Flow cytometry of CSF is increasingly being considered as the method of choice in patients suspected of leptomeningeal localization of hematological malignancies. Additionally, in several neuroinflammatory diseases such as multiple sclerosis and paraneoplastic neurological syndromes, flow cytometry is commonly performed to obtain insight into the immunopathogenesis of these diseases. Technically, the low cellularity of CSF samples, combined with the rapidly declining WBC viability, makes CSF flow cytometry challenging. Comparison of flow cytometry with microscopic and molecular techniques shows that each technique has its own advantages and is ideally combined. We expect that increasing the number of flow cytometric parameters that can be simultaneously studied within one sample, will further refine the information on CSF cell subsets in low-cellular CSF samples and enable to define cell populations more accurately. Copyright © 2011 International Clinical Cytometry Society.

  13. Multiphase Venturi Dual Energy Gamma Ray combination performance in NUEX flow loop; Desempenho no flowloop do NUEX da medicao multifasica Venturi Dual Energy Gamma Ray

    Energy Technology Data Exchange (ETDEWEB)

    Barreiros, Claudio; Taranto, Cleber; Costa, Alcemir [PETROBRAS S.A., Rio de Janeiro, RJ (Brazil); Pinguet, Bruno; Heluey, Vitor; Bessa, Fabiano; Loicq, Olivier [Schlumberger Servicos de Petroleo Ltda., Rio de Janeiro, RJ (Brazil)

    2008-07-01

    The Multiphase Venturi Dual Energy Gamma Ray Combination, Vx* technology, arrived in Brazil in 2000. PETROBRAS, Brazilian Oil Company, has been putting big efforts in its production business and also has demonstrated a large interest in having a multiphase meter approved by ANP for back allocation purposes. The oil industry was looking for ways to improve the back allocation process using an approved on line multiphase flow measurement device, thus replacing punctual test done today by a permanent monitoring device. Considering this scenario, a partnership project between PETROBRAS and Schlumberger was created in Brazil. The main objective of this project, which was held in NUEX flow loop, was to demonstrate to INMETRO (Brazilian Metrology Institute) that the Multiphase Venturi Dual Energy Gamma Ray Combination meter is able to be used for back allocation purpose. PETROBRAS and Schlumberger elaborated a complete methodology in the NUEX flow loop to demonstrate the results and benefits of the Multiphase Venturi Dual Energy Gamma Ray Combination meter. The test was witnessed by INMETRO and had a very good performance at the end. The results were within what was expected by Schlumberger, PETROBRAS and INMETRO. These results has been very useful to PETROBRAS in order to start using the Venturi Dual Energy Gamma Ray technology for well allocation purposes. (author)

  14. Experimental study of gas-liquid flow local characteristics in rectangular microchannel

    Directory of Open Access Journals (Sweden)

    Bartkus German

    2017-01-01

    Full Text Available Using high-speed video recording and the method of dual laser scanning the gas-liquid flow was investigated in a rectangular microchannel with an aspect ratio of 0.74 (cross section 269×362 μm. The T-mixer was used at the channel’s inlet for the two-phase flow formation. The peculiarity of this work is using a number of liquids (ethanol, distilled water, 40% aqueous ethanol with different physical properties, including surface tension, viscosity, and density, with nitrogen. Experiments were carried out for the vertically upward and horizontal flow. Using laser scanning method the maps of flow patterns were obtained for all mixtures.

  15. Nuclear DNA content of the hybrid plant pathogen Phytophthora andina determined by flow cytometry.

    Science.gov (United States)

    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.

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

    OpenAIRE

    Chen , Xiaoyi

    2015-01-01

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

  17. Laser Sensing of Vegetation Based on Dual Spectrum Measurements of Reflection Coefficients

    Directory of Open Access Journals (Sweden)

    M. L. Belov

    2017-01-01

    Full Text Available Currently, a promising trend in remote sensing of environment is to monitor the vegetative cover: evaluate the productivity of agricultural crops; evaluate the moisture content of soils and the state of ecosystems; provide mapping the sites of bogging, desertification, drought, etc.; control the phases of vegetation of crops, etc.Development of monitoring systems for remote detection of vegetation sites being under unfavorable conditions (low or high temperature, excess or lack of water, soil salinity, disease, etc. is of relevance. Optical methods are the most effective for this task. These methods are based on the physical features of reflection spectra in the visible and near infrared spectral range for vegetation under unfavorable conditions and vegetation under normal conditions.One of the options of optoelectronic equipment for monitoring vegetation condition is laser equipment that allows remote sensing of vegetation from the aircraft and mapping of vegetation sites with abnormal (inactive periods of vegetation reflection spectra with a high degree of spatial resolution.The paper deals with development of a promising dual-spectrum method for laser remote sensing of vegetation. Using the experimentally measured reflection spectra of different vegetation types, mathematical modeling of probability for appropriate detection and false alarms to solve a problem of detecting the vegetation under unfavorable conditions (with abnormal reflection spectra is performed based on the results of dual-spectrum measurements of the reflection coefficient.In mathematical modeling, the lidar system was supposed to provide sensing at wavelengths of 0.532 μm and 0.85 μm. The noise of the measurement was supposed to be normal with zero mean value and mean-square value of 1% -10%.It is shown that the method of laser sensing of vegetation condition based on the results of dual-spectrum measurement of the reflection coefficient at wavelengths of 0.532 μm and 0

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

    Science.gov (United States)

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

    2016-09-01

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

  19. Experimental and modelling study of reverse flow catalytic converters for natural gas/diesel dual fuel engine pollution control

    Energy Technology Data Exchange (ETDEWEB)

    Liu, B.

    2000-07-01

    There is renewed interest in the development of natural gas vehicles in response to the challenge to reduce urban air pollution and consumption of petroleum. The natural gas/diesel dual fuel engine is one way to apply natural gas to the conventional diesel engine. Dual fuel engines operating on natural gas and diesel emit less nitrogen oxides, and less carbon soot to the air compared to conventional diesel engines. The problem is that at light loads, fuel efficiency is reduced and emissions of hydrocarbons and carbon monoxide are increased. This thesis focused on control methods for emissions of hydrocarbons and carbon monoxide in the dual fuel engine at light loads. This was done by developing a reverse flow catalytic converter to complement dual fuel engine exhaust characteristics. Experimental measurements and numerical simulations of reverse flow catalytic converters were conducted. Reverse flow creates a high reactor temperature even when the engine is run at low exhaust temperature levels at light loads. The increase in reactor temperature from reverse flow could be 2 or 3 times higher than the adiabatic temperature increase, which is based on the reactor inlet temperature and concentration. This temperature makes it possible for greater than 90 per cent of the hydrocarbon and carbon monoxide to be converted with a palladium based catalyst. Reverse flow appears to be better than conventional unidirectional flow to deal with natural gas/diesel dual fuel engine pollution at light loads. Reverse flow could also maintain reactor temperature at over 800 K and hydrocarbon conversion at about 80 per cent during testing. The newly presented model simulates reactor performance with reasonable accuracy. Both carbon monoxide and methane oxidation over the palladium catalyst in excess oxygen and water were described using first order kinetics.

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

    Science.gov (United States)

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

    2013-09-01

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

  1. Flow cytometric evaluation of antibiotic effects on viability and mitochondrial function of refrigerated spermatozoa of Nile tilapia

    Science.gov (United States)

    Segovia, M.; Jenkins, J.A.; Paniagua-Chavez, C.; Tiersch, T.R.

    2000-01-01

    Improved techniques for storage and evaluation of fish sperm would enhance breeding programs around the world. The goal of this study was to test the effect of antibiotics on refrigerated sperm from Nile tilapia (Oreochromis niloticus) by use of flow cytometry with 2 dual-staining protocols for objective assessment of sperm quality. Concentrations of 1 x 109 sperm/mL were suspended in Ringer's buffer at 318 mOsmol/kg (pH 8.0). The fluorescent stains Sybr 14 (10 ??M), propidium iodide (2.4 mM), and rhodamine 123 (0.13 ??M) were used to assess cell viability and mitochondrial function. Three concentrations of ampicillin, gentamicin, and an antibiotic/antimycotic solution were added to fresh spermatozoa. Motility estimates and flow cytometry measurements were made daily during 7 d of refrigerated storage (4 ??C). The highest concentrations of gentamicin and antibiotic/antimycotic and all 3 concentrations of ampicillin significantly reduced sperm viability. The highest of each of the 3 antibiotic concentrations significantly reduced mitochondrial function. This study demonstrates that objective sperm quality assessments can be made using flow cytometry and that addition of antibiotics at appropriate concentrations can lengthen refrigerated storage time for tilapia spermatozoa. With minor modifications, these protocols can be adapted for use with sperm from other species and with other tissue types.

  2. Corrected Lymphocyte Percentages Reduce the Differences in Absolute CD4+ T Lymphocyte Counts between Dual-Platform and Single-Platform Flow Cytometric Approaches.

    Science.gov (United States)

    Noulsri, Egarit; Abudaya, Dinar; Lerdwana, Surada; Pattanapanyasat, Kovit

    2018-03-13

    To determine whether a corrected lymphocyte percentage could reduce bias in the absolute cluster of differentiation (CD)4+ T lymphocyte counts obtained via dual-platform (DP) vs standard single-platform (SP) flow cytometry. The correction factor (CF) for the lymphocyte percentages was calculated at 6 laboratories. The absolute CD4+ T lymphocyte counts in 300 blood specimens infected with human immunodeficiency virus (HIV) were determined using the DP and SP methods. Applying the CFs revealed that 4 sites showed a decrease in the mean bias of absolute CD4+ T lymphocyte counts determined via DP vs standard SP (-109 vs -84 cells/μL, -80 vs -58 cells/μL, -52 vs -45 cells/μL, and -32 vs 1 cells/μL). However, 2 participating laboratories revealed an increase in the difference of the mean bias (-42 vs -49 cells/μL and -20 vs -69 cells/μL). Use of the corrected lymphocyte percentage shows potential for decreasing the difference in CD4 counts between DP and the standard SP method.

  3. An Integrated Workflow To Assess Technical and Biological Variability of Cell Population Frequencies in Human Peripheral Blood by Flow Cytometry.

    Science.gov (United States)

    Burel, Julie G; Qian, Yu; Lindestam Arlehamn, Cecilia; Weiskopf, Daniela; Zapardiel-Gonzalo, Jose; Taplitz, Randy; Gilman, Robert H; Saito, Mayuko; de Silva, Aruna D; Vijayanand, Pandurangan; Scheuermann, Richard H; Sette, Alessandro; Peters, Bjoern

    2017-02-15

    In the context of large-scale human system immunology studies, controlling for technical and biological variability is crucial to ensure that experimental data support research conclusions. In this study, we report on a universal workflow to evaluate both technical and biological variation in multiparameter flow cytometry, applied to the development of a 10-color panel to identify all major cell populations and T cell subsets in cryopreserved PBMC. Replicate runs from a control donation and comparison of different gating strategies assessed the technical variability associated with each cell population and permitted the calculation of a quality control score. Applying our panel to a large collection of PBMC samples, we found that most cell populations showed low intraindividual variability over time. In contrast, certain subpopulations such as CD56 T cells and Temra CD4 T cells were associated with high interindividual variability. Age but not gender had a significant effect on the frequency of several populations, with a drastic decrease in naive T cells observed in older donors. Ethnicity also influenced a significant proportion of immune cell population frequencies, emphasizing the need to account for these covariates in immune profiling studies. We also exemplify the usefulness of our workflow by identifying a novel cell-subset signature of latent tuberculosis infection. Thus, our study provides a universal workflow to establish and evaluate any flow cytometry panel in systems immunology studies. Copyright © 2017 by The American Association of Immunologists, Inc.

  4. Breakdown plasma and vortex flow control for laser ignition using a combination of nano- and femto-second lasers.

    Science.gov (United States)

    Kojima, Hirokazu; Takahashi, Eiichi; Furutani, Hirohide

    2014-01-13

    The breakdown plasma and successive flow leading to combustion are controlled by the combination of a nano-second Nd:YAG laser and a femto-second Ti:Sapphire (TiS) laser. The behaviors are captured by an intensified charged coupled device (ICCD) camera and a high-speed schlieren optical system. The TiS laser determines the initial position of the breakdown by supplying the initial electrons in the optical axis of focusing YAG laser pulses. We show that the initial position of the breakdown can be controlled by the incident position of the TiS laser. In addition, the ignition lean limit of the flammable mixture changes depending on the TiS laser incident position, which is influenced by hot gas distribution and the flow in the flame kernel.

  5. Investigation of Dual-Vortical-Flow Hybrid Rocket Engine without Flame Holding Mechanism

    Directory of Open Access Journals (Sweden)

    A. Lai

    2018-01-01

    Full Text Available A 250 kgf thrust hybrid rocket engine was designed, tested, and verified in this work. Due to the injection and flow pattern of this engine, this engine was named dual-vortical-flow engine. This propulsion system uses N2O as oxidizer and HDPE as fuel. This engine was numerically investigated using a CFD tool that can handle reacting flow with finite-rate chemistry and coupled with the real-fluid model. The engine was further verified via a hot-fire test for 12 s. The ground Isp of the engine was 232 s and 221 s for numerical and hot-fire tests, respectively. An oscillation frequency with an order of 100 Hz was observed in both numerical and hot-fire tests with less than 5% of pressure oscillation. Swirling pattern on the fuel surface was also observed in both numerical and hot-fire test, which proves that this swirling dual-vortical-flow engine works exactly as designed. The averaged regression rate of the fuel surface was found to be 0.6~0.8 mm/s at the surface of disk walls and 1.5~1.7 mm/s at the surface of central core of the fuel grain.

  6. Flow with vibrational energy exchange, application to CO2 electric laser

    International Nuclear Information System (INIS)

    Dahan, Claude.

    1974-01-01

    The performances of a continuous wave (CO 2 , N 2 , He) laser ionized by an electron beam are calculated. Several types of phenomena are considered: energy exchange processes between molecules of laser medium, electron molecular excitation processes, aerodynamic phenomena: the energy exchanges accompanying the laser effect generate important quantities of heat, which have to be evacuated by the flow. After a survey of the fundamental assumptions on molecular phenomena, a computer code was developed for following, along the flow, the evolution of the thermodynamic parameters (pressure, temperature), of the laser gain, and of the electrical properties (electron density and temperature). To provide a finer description of the last ones, a model giving the energy distribution of the electrons in the laser medium was established [fr

  7. Flow cytometry of human primary epidermal and follicular keratinocytes.

    Science.gov (United States)

    Gragnani, Alfredo; Ipolito, Michelle Zampieri; Sobral, Christiane S; Brunialti, Milena Karina Coló; Salomão, Reinaldo; Ferreira, Lydia Masako

    2008-02-19

    The aim of this study was to characterize using flow cytometry cultured human primary keratinocytes isolated from the epidermis and hair follicles by different methods. Human keratinocytes derived from discarded fragments of total skin and scalp hair follicles from patients who underwent plastic surgery in the Plastic Surgery Division at UNIFESP were used. The epidermal keratinocytes were isolated by using 3 different methods: the standard method, upon exposure to trypsin for 30 minutes; the second, by treatment with dispase for 18 hours and with trypsin for 10 minutes; and the third, by treatment with dispase for 18 hours and with trypsin for 30 minutes. Follicular keratinocytes were isolated using the standard method. On comparing the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with dispase for 18 hours and with trypsin for 30 minutes, it was observed that the first group presented the largest number of viable cells, the smallest number of cells in late apoptosis and necrosis with statistical significance, and no difference in apoptosis. When we compared the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with trypsin, the first group presented the largest number of viable cells, the smallest number of cells in apoptosis with statistical significance, and no difference in late apoptosis and necrosis. When we compared the results of the group treated with dispase for 18 hours and with trypsin for 10 minutes with the results for follical isolation, there was a statistical difference in apoptosis and viable cells. The isolation method of treatment with dispase for 18 hours and with trypsin for 10 minutes produced the largest number of viable cells and the smallest number of cells in apoptosis/necrosis.

  8. Transverse-flow quasi-cw HF chemical laser: design and preliminary performance

    International Nuclear Information System (INIS)

    Gagne, J.M.; Mah, S.Q.; Conturie, Y.

    1974-01-01

    A small transverse-flow HF chemical laser has been constructed using a large volume microwave plasma generator for the production of F atoms. The F atoms react with hydrogen to form the lasing HF molecules. The active medium is about 5 cm long, and the maximum average laser power was found to be 560 mW for all lines. Three laser lines with wavelengths 2.61 μm, 2.64 μm, and 2.73 μm were observed. The time-varying laser transition profile closely resembles the density profile of the excited fluorine atoms in the plasma. Both profiles are greatly affected by changes in flow conditions

  9. Evaluation of Escherichia coli viability by flow cytometry: A method for determining bacterial responses to antibiotic exposure.

    Science.gov (United States)

    Boi, Paola; Manti, Anita; Pianetti, Anna; Sabatini, Luigia; Sisti, Davide; Rocchi, Marco Bruno; Bruscolini, Francesca; Galluzzi, Luca; Papa, Stefano

    2015-01-01

    In this study, we check for the presence of specific resistance genes by polymerase chain reaction (PCR) and then we used flow cytometry (FCM) to evaluate antibiotic-induced effects in different strains of Escherichia coli (E. coli). The presence of resistance genes was investigated by PCR in 10 strains of E. coli isolated from Foglia River. Bacterial responses to different antibiotics were also tested with FCM techniques by evaluating both the degree of decrease in viability and the light scatter changes in all of the strains. PCR revealed that only one strain exhibits the presence of one resistance gene. Despite this, analyses of strains using FCM evidenced the presence of viable subpopulations after antibiotic treatment. Furthermore, analyses of scatter signals revealed profound changes in the Forward Scatter and Side Scatter of the bacterial populations as a consequence of antibiotic exposure, confirming the viability and membrane potential data. The riverine strains were in general less sensitive to antibiotics than the reference strain (ATCC 25922). Antibiotic resistance is a widespread phenomena. The multiparametric approach based on FCM used in this study, providing results about different aspects (cell viability, membrane potential, light scatter changes), may overcome the limitation of PCR and could represent an adequate method for the evaluation of bacteria responses to antibiotic exposure. © 2014 International Clinical Cytometry Society.

  10. Cytometry of mammalian sperm

    Energy Technology Data Exchange (ETDEWEB)

    Gledhill, B.L.

    1983-10-11

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

  11. Cytometry of mammalian sperm

    International Nuclear Information System (INIS)

    Gledhill, B.L.

    1983-01-01

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

  12. Unsteady supercritical/critical dual flowpath inlet flow and its control methods

    Directory of Open Access Journals (Sweden)

    Jun LIU

    2017-12-01

    Full Text Available The characteristics of unsteady flow in a dual-flowpath inlet, which was designed for a Turbine Based Combined Cycle (TBCC propulsion system, and the control methods of unsteady flow were investigated experimentally and numerically. It was characterized by large-amplitude pressure oscillations and traveling shock waves. As the inlet operated in supercritical condition, namely the terminal shock located in the throat, the shock oscillated, and the period of oscillation was about 50 ms, while the amplitude was 6 mm. The shock oscillation was caused by separation in the diffuser. This shock oscillation can be controlled by extending the length of diffuser which reduces pressure gradient along the flowpath. As the inlet operated in critical condition, namely the terminal shock located at the shoulder of the third compression ramp, the shock oscillated, and the period of oscillation was about 7.5 ms, while the amplitude was 12 mm. At this condition, the shock oscillation was caused by an incompatible backpressure in the bleed region. It can be controlled by increasing the backpressure of the bleed region. Keywords: Airbreathing hypersonic vehicle, Dual flowpath inlet, Terminal shock oscillation, Turbine based combined cycle, Unsteady flow

  13. Monitoring Plasmodium falciparum growth and development by UV flow cytometry using an optimized Hoechst-thiazole orange staining strategy.

    Science.gov (United States)

    Grimberg, Brian T; Erickson, John J; Sramkoski, R Michael; Jacobberger, James W; Zimmerman, Peter A

    2008-06-01

    The complex life cycle of Plasmodium falciparum (Pf) makes it difficult to limit infections and reduce the risk of severe malaria. Improved understanding of Pf blood-stage growth and development would provide new opportunities to evaluate and interfere with successful completion of the parasite's life cycle. Cultured blood stage Pf was incubated with Hoechst 33342 (HO) and thiazole orange (TO) to stain DNA and total nucleic acids, respectively. Correlated HO and TO fluorescence emissions were then measured by flow cytometry. Complex bivariate data patterns were analyzed by manual cluster gating to quantify parasite life cycle stages. The permutations of viable staining with both reagents were tested for optimal detection of parasitized RBC (pRBC). Pf cultures were exposed to HO and TO simultaneously to achieve optimal staining of pRBC and consistent quantification of early and late stages of the replicative cycle (rings through schizonts). Staining of Pf nucleic acids allows for analysis of parasite development in the absence of fixatives, lysis, or radioactivity to enable examination of erythrocytes from parasite invasion through schizont rupture using sensitive and rapid assay procedures. Investigation of the mechanisms by which anti-malarial drugs and antibodies act against different Pf lifecycle stages will be aided by this cytometric strategy. (c) 2008 International Society for Advancement of Cytometry.

  14. A quasi-three-level dual-wavelength thin-disk laser at 1024 and 1030 nm based on a diode-pumped Yb:YAG crystal

    International Nuclear Information System (INIS)

    Sun, G C; Li, Y D; Zhao, M; Chen, X Y; Wang, J B; Chen, G B

    2013-01-01

    A diode-end-pumped Yb:YAG dual-wavelength continuous-wave (cw) laser that generates simultaneous laser action at wavelengths of 1024 and 1030 nm is demonstrated for the first time. A total output power of 897 mW for the dual-wavelength was achieved at an incident pump power of 17.8 W. Furthermore, intracavity sum-frequency mixing at 1024 and 1030 nm was then realized in an LBO crystal to reach the green range. We obtained a total cw output power of 85 mW at 513.5 nm. (paper)

  15. Dual-plane ultrasound flow measurements in liquid metals

    International Nuclear Information System (INIS)

    Büttner, Lars; Nauber, Richard; Burger, Markus; Czarske, Jürgen; Räbiger, Dirk; Franke, Sven; Eckert, Sven

    2013-01-01

    An ultrasound measurement system for dual-plane, two-component flow velocity measurements especially in opaque liquids is presented. Present-day techniques for measuring local flow structures in opaque liquids disclose considerable drawbacks concerning line-wise measurement of single ultrasound probes. For studying time-varying flow patterns, conventional ultrasound techniques are either limited by time-consuming mechanical traversing or by the sequential operation of single probes. The measurement system presented within this paper employs four transducer arrays with a total of 100 single elements which allows for flow mapping without mechanical traversing. A high frame rate of several 10 Hz has been achieved due to an efficient parallelization scheme using time-division multiplexing realized by a microcontroller-based electronic switching matrix. The functionality and capability of the measurement system are demonstrated on a liquid metal flow at room temperature inside a cube driven by a rotating magnetic field (RMF). For the first time, the primary and the secondary flow have been studied in detail and simultaneously using a configuration with two crossed measurement planes. The experimental data confirm predictions made by numeric simulation. After a sudden switching on of the RMF, inertial oscillations of the secondary flow were observed by means of a time-resolved measurement with a frame rate of 3.4 Hz. The experiments demonstrate that the presented measurement system is able to investigate complex and transient flow structures in opaque liquids. Due to its ability to study the temporal evolution of local flow structures, the measurement system could provide considerable progress for fluid dynamics research, in particular for applications in the food industry or liquid metal technologies. (paper)

  16. Dual-plane ultrasound flow measurements in liquid metals

    Science.gov (United States)

    Büttner, Lars; Nauber, Richard; Burger, Markus; Räbiger, Dirk; Franke, Sven; Eckert, Sven; Czarske, Jürgen

    2013-05-01

    An ultrasound measurement system for dual-plane, two-component flow velocity measurements especially in opaque liquids is presented. Present-day techniques for measuring local flow structures in opaque liquids disclose considerable drawbacks concerning line-wise measurement of single ultrasound probes. For studying time-varying flow patterns, conventional ultrasound techniques are either limited by time-consuming mechanical traversing or by the sequential operation of single probes. The measurement system presented within this paper employs four transducer arrays with a total of 100 single elements which allows for flow mapping without mechanical traversing. A high frame rate of several 10 Hz has been achieved due to an efficient parallelization scheme using time-division multiplexing realized by a microcontroller-based electronic switching matrix. The functionality and capability of the measurement system are demonstrated on a liquid metal flow at room temperature inside a cube driven by a rotating magnetic field (RMF). For the first time, the primary and the secondary flow have been studied in detail and simultaneously using a configuration with two crossed measurement planes. The experimental data confirm predictions made by numeric simulation. After a sudden switching on of the RMF, inertial oscillations of the secondary flow were observed by means of a time-resolved measurement with a frame rate of 3.4 Hz. The experiments demonstrate that the presented measurement system is able to investigate complex and transient flow structures in opaque liquids. Due to its ability to study the temporal evolution of local flow structures, the measurement system could provide considerable progress for fluid dynamics research, in particular for applications in the food industry or liquid metal technologies.

  17. Flow cytometry evidence of human granulocytes interaction with polyhedral oligomeric silsesquioxanes: effect of nanoparticle charge

    International Nuclear Information System (INIS)

    Renò, Filippo; Rizzi, Manuela; Pittarella, Pamela; Carniato, Fabio; Olivero, Francesco; Marchese, Leonardo

    2013-01-01

    Nanoparticles (NPs) entering the human body are immediately confronted with the innate part of human immune system. In particular, monocyte and neutrophil granulocytes readily clear particles by phagocytosis, even if in the case of NPs the uptake mechanism may be classified as macropinocytosis. Among engineered nanoparticles, in the last years, siliceous materials have emerged as promising materials for several applications ranging from catalysis to biomedical. The polyhedral oligomeric silsesquioxanes (POSS) are nanodimensional, easily synthesizable molecular compounds and POSS-based systems are promising carriers for biological molecules. In this work, the ability of human granulocytes to uptake positively and negatively charged POSS was measured using a simple flow cytometry analysis based on cell size modifications. The data obtained showed that after a 30 min exposure only positive NPs were uptaken by human granulocyte using both macropinocytosis and clathrin-mediated mechanisms as demonstrated by uptake inhibition mediated by amiloride and chlorpromazine. (paper)

  18. Application of the flow cytometry for determination of the amount of DNA in Yersinia pestis cells under the influence of serotonin (5-hydroxytryptamine)

    Science.gov (United States)

    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.

  19. A novel quantitative kinase assay using bacterial surface display and flow cytometry.

    Directory of Open Access Journals (Sweden)

    Sónia Troeira Henriques

    Full Text Available The inhibition of tyrosine kinases is a successful approach for the treatment of cancers and the discovery of kinase inhibitor drugs is the focus of numerous academic and pharmaceutical laboratories. With this goal in mind, several strategies have been developed to measure kinase activity and to screen novel tyrosine kinase inhibitors. Nevertheless, a general non-radioactive and inexpensive approach, easy to implement and adapt to a range of applications, is still missing. Herein, using Bcr-Abl tyrosine kinase, an oncogenic target and a model protein for cancer studies, we describe a novel cost-effective high-throughput screening kinase assay. In this approach, named the BacKin assay, substrates displayed on a Bacterial cell surface are incubated with Kinase and their phosphorylation is examined and quantified by flow cytometry. This approach has several advantages over existing approaches, as using bacteria (i.e. Escherichia coli to display peptide substrates provides a self renewing solid support that does not require laborious chemical strategies. Here we show that the BacKin approach can be used for kinetic and mechanistic studies, as well as a platform to characterize and identify small-molecule or peptide-based kinase inhibitors with potential applications in drug development.

  20. A rapid method for infectivity titration of Andes hantavirus using flow cytometry.

    Science.gov (United States)

    Barriga, Gonzalo P; Martínez-Valdebenito, Constanza; Galeno, Héctor; Ferrés, Marcela; Lozach, Pierre-Yves; Tischler, Nicole D

    2013-11-01

    The focus assay is currently the most commonly used technique for hantavirus titer determination. This method requires an incubation time of between 5 and 11 days to allow the appearance of foci after several rounds of viral infection. The following work presents a rapid Andes virus (ANDV) titration assay, based on viral nucleocapsid protein (N) detection in infected cells by flow cytometry. To this end, an anti-N monoclonal antibody was used that was developed and characterized previously. ANDV N could be detected as early as 6 h post-infection, while viral release was not observed until 24-48 h post-infection. Given that ANDV detection was performed during its first round of infection, a time reduction for titer determination was possible and provided results in only two days. The viral titer was calculated from the percentage of N positive cells and agreed with focus assay titers. Furthermore, the assay was applied to quantify the inhibition of ANDV cell entry by patient sera and by preventing endosome acidification. This novel hantavirus titration assay is a highly quantitative and sensitive tool that facilitates infectivity titration of virus stocks, rapid screening for antiviral drugs, and may be further used to detect and quantify infectious virus in human samples. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. Evaporation-induced gas-phase flows at selective laser melting

    Science.gov (United States)

    Zhirnov, I.; Kotoban, D. V.; Gusarov, A. V.

    2018-02-01

    Selective laser melting is the method for 3D printing from metals. A solid part is built from powder layer-by-layer. A continuum-wave laser beam scans every powder layer to fuse powder. The process is studied with a high-speed CCD camera at the frame rate of 104 fps and the resolution up to 5 µm per pixel. Heat transfer and evaporation in the laser-interaction zone are numerically modeled. Droplets are ejected from the melt pool in the direction around the normal to the melt surface and the powder particles move in the horizontal plane toward the melt pool. A vapor jet is observed in the direction of the normal to the melt surface. The velocities of the droplets, the powder particles, and the jet flow and the mass loss due to evaporation are measured. The gas flow around the vapor jet is calculated by Landau's model of submerged jet. The measured velocities of vapor, droplets, and powder particles correlate with the calculated flow field. The obtained results show the importance of evaporation and the flow of the vapor and the ambient gas. These gas-dynamic phenomena can explain the formation of the denudated zones and the instability at high-energy input.

  2. Measurement of Vertical Oil-in-water Two-phase Flow Using Dual-modality ERT-EMF System

    OpenAIRE

    Faraj, Yousef; Wang, Mi; Jia, Jiabin; Wang, Qiang; Xie, Cheng-gang; Oddie, Gary; Primrose , Ken; Qiu, Changhua

    2015-01-01

    Oil-in-water two-phase flows are often encountered in the upstream petroleum industry. The measurement of phase flow rates is of particular importance for managing oil production and water disposal and/or water reinjection. The complexity of oil-in-water flow structures creates a challenge to flow measurement. This paper proposes a new method of two-phase flow metering, which is based on the use of dual-modality system and multidimensional data fusion. The Electrical Resistance Tomography sys...

  3. Adaptive real-time dual-comb spectroscopy

    Science.gov (United States)

    Ideguchi, Takuro; Poisson, Antonin; Guelachvili, Guy; Picqué, Nathalie; Hänsch, Theodor W.

    2014-01-01

    The spectrum of a laser frequency comb consists of several hundred thousand equally spaced lines over a broad spectral bandwidth. Such frequency combs have revolutionized optical frequency metrology and they now hold much promise for significant advances in a growing number of applications including molecular spectroscopy. Despite an intriguing potential for the measurement of molecular spectra spanning tens of nanometres within tens of microseconds at Doppler-limited resolution, the development of dual-comb spectroscopy is hindered by the demanding stability requirements of the laser combs. Here we overcome this difficulty and experimentally demonstrate a concept of real-time dual-comb spectroscopy, which compensates for laser instabilities by electronic signal processing. It only uses free-running mode-locked lasers without any phase-lock electronics. We record spectra spanning the full bandwidth of near-infrared fibre lasers with Doppler-limited line profiles highly suitable for measurements of concentrations or line intensities. Our new technique of adaptive dual-comb spectroscopy offers a powerful transdisciplinary instrument for analytical sciences. PMID:24572636

  4. Adaptive real-time dual-comb spectroscopy

    Science.gov (United States)

    Ideguchi, Takuro; Poisson, Antonin; Guelachvili, Guy; Picqué, Nathalie; Hänsch, Theodor W.

    2014-02-01

    The spectrum of a laser frequency comb consists of several hundred thousand equally spaced lines over a broad spectral bandwidth. Such frequency combs have revolutionized optical frequency metrology and they now hold much promise for significant advances in a growing number of applications including molecular spectroscopy. Despite an intriguing potential for the measurement of molecular spectra spanning tens of nanometres within tens of microseconds at Doppler-limited resolution, the development of dual-comb spectroscopy is hindered by the demanding stability requirements of the laser combs. Here we overcome this difficulty and experimentally demonstrate a concept of real-time dual-comb spectroscopy, which compensates for laser instabilities by electronic signal processing. It only uses free-running mode-locked lasers without any phase-lock electronics. We record spectra spanning the full bandwidth of near-infrared fibre lasers with Doppler-limited line profiles highly suitable for measurements of concentrations or line intensities. Our new technique of adaptive dual-comb spectroscopy offers a powerful transdisciplinary instrument for analytical sciences.

  5. A novel multiparametric flow cytometry-based cytotoxicity assay simultaneously immunophenotypes effector cells: Comparisons to a 4 h 51Cr-release assay

    OpenAIRE

    Kim, GG; Donnenberg, VS; Donnenberg, AD; Gooding, W; Whiteside, TL

    2007-01-01

    Natural killer (NK) cell- or T cell-mediated cytotoxicity traditionally is measured in 4-16h 51Cr-release assays (CRA). A new four-color flow cytometry-based cytotoxicity assay (FCC) was developed to simultaneously measure NK cell cytotoxicity and NK cell phenotype (CD3−CD16+CD56+). Target cells, K562 or Daudi, were labeled with Cell Tracker Orange (CTO) prior to the addition of effector cells. Following co-incubation, 7 amino-actinomycin D (7-AAD) was added to measure death of target cells. ...

  6. Shot-Noise-Limited Dual-Beam Detector for Atmospheric Trace-Gas Monitoring with Near-Infrared Diode Lasers

    Science.gov (United States)

    Durry, Georges; Pouchet, Ivan; Amarouche, Nadir; Danguy, Théodore; Megie, Gerard

    2000-10-01

    A dual-beam detector is used to measure atmospheric trace species by differential absorption spectroscopy with commercial near-infrared InGaAs laser diodes. It is implemented on the Spectrom tre Diodes Laser Accordables, a balloonborne tunable diode laser spectrometer devoted to the in situ monitoring of CH 4 and H 2 O. The dual-beam detector is made of simple analogical subtractor circuits combined with InGaAs photodiodes. The detection strategy consists in taking the balanced analogical difference between the reference and the sample signals detected at the input and the output of an open optical multipass cell to apply the full dynamic range of the measurements (16 digits) to the weak molecular absorption information. The obtained sensitivity approaches the shot-noise limit. With a 56-m optical cell, the detection limit obtained when the spectra is recorded within 8 ms is 10 4 (expressed in absorbance units). The design and performances of both a simple substractor and an upgraded feedback substractor circuit are discussed with regard to atmospheric in situ CH 4 absorption spectra measured in the 1.653- m region. Mixing ratios are obtained from the absorption spectra by application of a nonlinear least-squares fit to the full molecular line shape in conjunction with in situ P and T measurements.

  7. Analysis of multiphase flows using dual-energy gamma densitometry and neural networks

    International Nuclear Information System (INIS)

    Bishop, C.M.; James, G.D.

    1993-01-01

    Dual-energy gamma densitometry offers a powerful technique for the non-intrusive analysis of multiphase flows. By employing multiple beam lines, information on the phase configuration can be obtained. Once the configuration is known, it then becomes possible in principle to determine the phase fractions. In practice, however, the extraction of the phase fractions from the densitometer data is complicated by the wide variety of phase configurations which can arise, and by the considerable difficulties of modelling multiphase flows. In this paper we show that neural network techniques provide a powerful approach to the analysis of data from dual-energy gamma densitometers, allowing both the phase configuration and the phase fractions to be determined with high accuracy, whilst avoiding the uncertainties associated with modelling. The technique is well suited to the determination of oil, water and gas fractions in multiphase oil pipelines. Results from linear and non-linear network models are compared, and a new technique for validating the network output is described. (orig.)

  8. A high-performance dual-scale porous electrode for vanadium redox flow batteries

    Science.gov (United States)

    Zhou, X. L.; Zeng, Y. K.; Zhu, X. B.; Wei, L.; Zhao, T. S.

    2016-09-01

    In this work, we present a simple and cost-effective method to form a dual-scale porous electrode by KOH activation of the fibers of carbon papers. The large pores (∼10 μm), formed between carbon fibers, serve as the macroscopic pathways for high electrolyte flow rates, while the small pores (∼5 nm), formed on carbon fiber surfaces, act as active sites for rapid electrochemical reactions. It is shown that the Brunauer-Emmett-Teller specific surface area of the carbon paper is increased by a factor of 16 while maintaining the same hydraulic permeability as that of the original carbon paper electrode. We then apply the dual-scale electrode to a vanadium redox flow battery (VRFB) and demonstrate an energy efficiency ranging from 82% to 88% at current densities of 200-400 mA cm-2, which is record breaking as the highest performance of VRFB in the open literature.

  9. Flow-Induced Vibration Measurement of an Inner Cladding Tube in a Simulated Dual-Cooled Fuel Rod

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Kang Hee; Kim, Hyung Kyu; Yoon, Kyung Ho; Lee, Young Ho; Kim, Jae Yong [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2009-05-15

    To create an internal coolant flow passage in a dual cooled fuel rod, an inner cladding tube cannot have intermediate supports enough to relieve its vibration. Thus it can be suffered from a flow-induced vibration (FIV) more severely than an outer cladding tube which will be supported by series of spacer grids. It may cause a fatigue failure at welding joints on the cladding's end plug or fluid elastic instability of long, slender inner cladding due to decrease of a critical flow velocity. This is one of the challenging technical issues when a dual cooled fuel assembly is to be realized into a conventional reactor core To study an actual vibration phenomenon of a dual cooled fuel rod, FIV tests using a small-scale test bundle are being carried out. Measurement results of inner cladding tube of two typically simulated rods are presented. Causes of the differences in the vibration amplitude and response spectrum of the inner cladding tube in terms of intermediate support condition and pellet stacking are discussed.

  10. Optimization of the C11-BODIPY(581/591) dye for the determination of lipid oxidation in Chlamydomonas reinhardtii by flow cytometry.

    Science.gov (United States)

    Cheloni, Giulia; Slaveykova, Vera I

    2013-10-01

    Lipid oxidation is a recognized end point for the study of oxidative stress and is an important parameter to describe the mode of micropollutant action on aquatic microorganisms. Therefore, the development of quick and reliable methodologies probing the oxidative stress and damage in living cells is highly sought. In the present proof-of-concept work, we examined the potential of the fluorescent dye C11-BODIPY(591/581) to probe lipid oxidation in the green microalga Chlamydomonas reinhardtii. C11-BODIPY(591/581) staining was combined with flow cytometry measurements to obtain multiparameter information on cellular features and oxidative stress damage within single cells. First, staining conditions were optimized by exploring the capability of the dye to stain algal cells under increasing cell and dye concentrations and different staining procedures. Then lipid oxidation in algae induced by short- and long-term exposures to the three metallic micropollutants, copper, mercury, and nanoparticulate copper oxide, and the two organic contaminants, diethyldithiocarbamate (DDC) and diuron was determined. In this work we pointed out C11-BODIPY(591/581) applicability in a wide range of exposure conditions, including studies of oxidation as a function of time and that it is suitable for in vivo measurements of lipid oxidation due to its high permeation and stability in cells and its low interference with algal autofluorescence. © 2013 International Society for Advancement of Cytometry. Copyright © 2013 International Society for Advancement of Cytometry.

  11. Nature of convection-stabilized dc arcs in dual-flow nozzle geometry

    International Nuclear Information System (INIS)

    Serbetci, I.; Nagamatsu, H.T.

    1990-01-01

    In this paper, an experimental investigation of the steady-state low-current air arcs in a dual-flow nozzle system is presented. First, the cold flow with no arc as determined for various nozzle geometries, i.e., two- and three-dimensional and orifice nozzles, and nozzle pressure ratios. Supersonic flow separation and oblique and detached shock waves were observed in the flow field. Using a finite-element computer program, the Mach number contours were determined in the flow field for various nozzle-gap spacings and pressure ratios. In addition, the dc arc voltage and current measurements were made for an electrode gap spacing of ∼ 5.5 cm and current levels of I ∼ 25, 50, and 100 A for the three nozzle geometries. The arc voltage and arc power increased rapidly as the flow speed increased from zero to sonic velocity at the nozzle throat. The shock waves in the converging-diverging nozzles resulted in a decrease in the overall resistance by about 15 percent

  12. Use of flow cytometry for the adhesion analysis of Streptococcus pyogenes mutant strains to epithelial cells: investigation of the possible role of surface pullulanase and cysteine protease, and the transcriptional regulator Rgg

    Directory of Open Access Journals (Sweden)

    Finne Jukka

    2006-02-01

    Full Text Available Abstract Background Flow cytometry based adherence assay is a potentially powerful but little used method in the study of bacterial binding to host structures. We have previously characterized a glycoprotein-binding activity in Streptococcus pyogenes called 'strepadhesin' binding to thyroglobulin, submaxillar mucin, fetuin and asialofetuin. We have identified surface-associated pullulanase (PulA and cysteine protease (SpeB as carriers of strepadhesin activity. In the present paper, we investigated the use of flow cytometry as a method to study the binding of Rgg, SpeB and PulA knock-out strains to cultured human epithelial cells. Results Streptococcal mutants were readily labelled with CFDA-SE and their binding to epithelial cells could be effectively studied by flow cytometry. A strain deficient in Rgg expression showed increased binding to the analyzed epithelial cell lines of various origin. Inactivation of SpeB had no effect on the adhesion, while PulA knock-out strains displayed decreased binding to the cell lines. Conclusion These results suggest that the flow cytometric assay is a valuable tool in the analysis of S. pyogenes adherence to host cells. It appears to be an efficient and sensitive tool for the characterization of interactions between the bacteria and the host at the molecular level. The results also suggest a role for Rgg regulated surface molecules, like PulA, in the adhesion of S. pyogenes to host cells.

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

    Directory of Open Access Journals (Sweden)

    Monika Lukomska-Szymanska

    2017-04-01

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

  14. Generation of dual-wavelength, synchronized, tunable, high energy, femtosecond laser pulses with nearly perfect gaussian spatial profile

    Science.gov (United States)

    Wang, J.-K.; Siegal, Y.; Lü, C.; Mazur, E.

    1992-07-01

    We use self-phase modulation in a single-mode fiber to produce broadband femtosecond laser pulses. Subsequent amplification through two Bethune cells yields high-energy, tunable, pulses synchronized with the output of an amplified colliding-pulse-modelocked (CPM) laser. We routinely obtain tunable 200 μJ pulses of 42 fs (fwhm) duration with a nearly perfect gaussian spatial profile. Although self-phase modulation in a single-mode fiber is widely used in femtosecond laser systems, amplification of a fiber-generated supercontinuum in a Bethune cell amplifier is a new feature which maintains the high-quality spatial profile while providing high gain. This laser system is particularly well suited for high energy dual-wavelength pump=probe experiments and time-resolved four-wave mixing spectroscopy.

  15. Launch and capture of a single particle in a pulse-laser-assisted dual-beam fiber-optic trap

    Science.gov (United States)

    Fu, Zhenhai; She, Xuan; Li, Nan; Hu, Huizhu

    2018-06-01

    The rapid loading and manipulation of microspheres in optical trap is important for its applications in optomechanics and precision force sensing. We investigate the microsphere behavior under coaction of a dual-beam fiber-optic trap and a pulse laser beam, which reveals a launched microsphere can be effectively captured in a spatial region. A suitable order of pulse duration for launch is derived according to the calculated detachment energy threshold of pulse laser. Furthermore, we illustrate the effect of structural parameters on the launching process, including the spot size of pulse laser, the vertical displacement of beam waist and the initial position of microsphere. Our result will be instructive in the optimal design of the pulse-laser-assisted optical tweezers for controllable loading mechanism of optical trap.

  16. Convection flows driven by laser heating of a liquid layer

    OpenAIRE

    Rivière , David; Selva , Bertrand; Chraibi , Hamza; Delabre , Ulysse; Delville , Jean-Pierre

    2016-01-01

    International audience; When a fluid is heated by the absorption of a continuous laser wave, the fluid density decreases in the heated area. This induces a pressure gradient that generates internal motion of the fluid. Due to mass conservation, convection eddies emerge in the sample. To investigate these laser-driven bulk flows at the microscopic scale, we built a setup to perform temperature measurements with a fluorescent-sensitive dye on the one hand, and measured the flow pattern at diffe...

  17. Directional synthetic aperture flow imaging using a dual stage beamformer approach

    DEFF Research Database (Denmark)

    Li, Ye; Jensen, Jørgen Arendt

    2011-01-01

    . The new method has been studied using the Field II simulations and experimental flow rig measurements. A linear array transducer with 7 MHz center frequency is used, and 64 elements are active to transmit and receive signals. The data is processed in two stages. The first stage has a fixed focus point......A new method for directional synthetic aperture flow imaging using a dual stage beamformer approach is presented. The velocity estimation is angle independent and the amount of calculations is reduced compared to full synthetic aperture, but still maintains all the advantages at the same time....... In the second stage, focal points are considered as virtual sources and data is beamformed along the flow direction. Then the velocities are estimated by finding the spatial shift between two signals. In the experimental measurements the angle between the transmit beam and flow vessel was 70 and a laminar flow...

  18. Expression of the chondroitin sulphate proteoglycan molecular complex in six human melanoma xenograft lines studied by flow cytometry and immunohistochemistry.

    Science.gov (United States)

    Nagelhus, T A; Rofstad, E K

    1993-06-01

    The expression of the chondroitin sulphate proteoglycan (CSP) molecular complex in six human melanoma xenograft lines (BEX-t, COX-t, HUX-t, ROX-t, SAX-t, WIX-t) was studied by flow cytometry and immunohistochemistry using the monoclonal antibodies 9.2.27, ME31.3, G7A5, and NKI.M6. The two methods and the four antibodies gave consistent results. The six melanoma lines could be divided into three distinct groups of two lines each; expression was high in the HUX-t and ROX-t lines and intermediate in the BEX-t and SAX-t lines, whereas the COX-t and WIX-t lines were negative. The mean number of epitopes per cell for 9.2.27 was approximately twice as high as for ME31.3, G7A5, and NKI.M6 and was estimated to range from 0.8 +/- 0.1 x 10(5) to 1.9 +/- 0.2 x 10(5) in the positive xenograft lines. The expression of the CSP complex was heterogeneous. The immunofluorescence histograms measured by flow cytometry were therefore broad for all tumour lines. A significant fraction of the HUX-t cells was negative or weakly stained. These cells appeared as clear negative patches in the immunohistochemical preparations. Moreover, most morphologically intact tumour cells adjacent to necrotic areas did not show significant expression of the CSP complex, irrespective of tumour line. These cells were probably hypoxic and thus resistant to radiation therapy. The expression of the CSP complex in the xenograft lines was similar to that reported for melanoma in man.

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

    Science.gov (United States)

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

    2017-09-05

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

  20. Dual wavelength Mode-Locking of InAs/InP quantum dot laser diodes at 1.5µm

    NARCIS (Netherlands)

    Tahvili, M.S.; Heck, M.J.R.; Nötzel, R.; Smit, M.K.; Bente, E.A.J.M.

    2011-01-01

    We report on stable dual-wavelength mode-locking of 3.1GHz and 10GHz two-section InAs/InP(100) quantum dot laser diodes. Evaluation of relative time delay between different spectral components indicates opposite sign of chirp over the two spectral lobes

  1. Ploidy levels among species in the 'Oxalis tuberosa alliance' as inferred by flow cytometry.

    Science.gov (United States)

    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.

  2. Dual plane problems for creeping flow of power-law incompressible medium

    Directory of Open Access Journals (Sweden)

    Dmitriy S. Petukhov

    2016-09-01

    Full Text Available In this paper, we consider the class of solutions for a creeping plane flow of incompressible medium with power-law rheology, which are written in the form of the product of arbitrary power of the radial coordinate by arbitrary function of the angular coordinate of the polar coordinate system covering the plane. This class of solutions represents the asymptotics of fields in the vicinity of singular points in the domain occupied by the examined medium. We have ascertained the duality of two problems for a plane with wedge-shaped notch, at which boundaries in one of the problems the vector components of the surface force vanish, while in the other—the vanishing components are the vector components of velocity, We have investigated the asymptotics and eigensolutions of the dual nonlinear eigenvalue problems in relation to the rheological exponent and opening angle of the notch for the branch associated with the eigenvalue of the Hutchinson–Rice–Rosengren problem learned from the problem of stress distribution over a notched plane for a power law medium. In the context of the dual problem we have determined the velocity distribution in the flow of power-law medium at the vertex of a rigid wedge, We have also found another two eigenvalues, one of which was determined by V. V. Sokolovsky for the problem of power-law fluid flow in a convergent channel.

  3. 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)

  4. Vitreous flow rates through dual pneumatic cutters: effects of duty cycle and cut rate

    Directory of Open Access Journals (Sweden)

    Abulon DJK

    2015-02-01

    Full Text Available Dina Joy K Abulon Medical Affairs, Alcon Research, Ltd, Lake Forest, CA, USA Purpose: We aimed to investigate effects of instrument settings on porcine vitreous flow rates through dual pneumatic high-speed vitrectomy probes. Methods: The CONSTELLATION® Vision System was tested with 250, 450, and 650 mmHg of vacuum using six ULTRAVIT® vitrectomy probes of each diameter (25+®, 25, 23, and 20 gauge operated from 500 cuts per minute (cpm up to 5,000 cpm. Duty cycle modes tested included biased open, 50/50, and biased closed. Flow rates were calculated by assessing the change in weight of porcine eyes during vitreous aspiration. Volumetric flow rate was measured with a computer-connected electronic scale. Results: At lower cut rates, the biased open mode produced higher flow than did the 50/50 mode, which produced higher flow than did the biased closed mode. In the biased closed and 50/50 modes, vitreous flow rates tended to increase with increasing cut rate. Vitreous flow rates in the biased open duty cycle mode remained relatively constant across cut rates. Conclusion: Vitreous flow rates through dual pneumatic vitrectomy probes could be manipulated by changing the duty cycle modes on the vitrectomy system. Differences in duty cycle behavior suggest that high-speed cut rates of 5,000 cpm may optimize vitreous aspiration. Keywords: enhanced 25-gauge vitrectomy, 25-gauge vitrectomy, 20-gauge vitrectomy, 23-gauge vitrectomy, aspiration, Constellation Vision System

  5. The Diagnosis of Gastric Mucosa-associated Lymphoid Tissue Lymphoma by Flow Cytometry and Fluorescence in situ Hybridization of Biopsy Specimens.

    Science.gov (United States)

    Matsueda, Katsunori; Omote, Sizuma; Sakata, Masahiro; Fujita, Isao; Horii, Jouichiro; Toyokawa, Tatsuya

    2018-04-15

    Mucosa-associated lymphoid tissue (MALT) lymphoma and reactive inflammatory lymphoid changes are frequently difficult to distinguish based on a routine histological differential diagnosis. We were unable to diagnose gastric MALT lymphoma histologically using specimens obtained by endoscopy, although a flow cytometry (FCM) analysis demonstrated clonality of neoplastic cells by separating cells by CD45 gating. Furthermore, a fluorescence in situ hybridization (FISH) analysis showed trisomy 18. We therefore diagnosed gastric MALT lymphoma with trisomy 18. We recommend that FCM and FISH analyses of biopsy specimens be considered for diagnosing gastric MALT lymphoma if this diagnosis is suspected based on endoscopic findings.

  6. A radio-high-performance liquid chromatography dual-flow cell gamma-detection system for on-line radiochemical purity and labeling efficiency determination

    DEFF Research Database (Denmark)

    Lindegren, S; Jensen, H; Jacobsson, L

    2014-01-01

    In this study, a method of determining radiochemical yield and radiochemical purity using radio-HPLC detection employing a dual-flow-cell system is evaluated. The dual-flow cell, consisting of a reference cell and an analytical cell, was constructed from two PEEK capillary coils to fit into the w...

  7. Dual-wavelength passive and hybrid mode-locking of 3, 4.5 and 10 GHz InAs/InP(100) quantum dot lasers.

    Science.gov (United States)

    Tahvili, M S; Du, L; Heck, M J R; Nötzel, R; Smit, M K; Bente, E A J M

    2012-03-26

    We present an investigation of passive and hybrid mode-locking in Fabry-Pérot type two-section InAs/InP(100) quantum dot lasers that show dual wavelength operation. Over the whole current and voltage range for mode-locking of these lasers, the optical output spectra show two distinct lobes. The two lobes provide a coherent bandwidth and are verified to lead to two synchronized optical pulses. The generated optical pulses are elongated in time due to a chirp which shows opposite signs over the two spectral lobes. Self-induced mode-locking in the single-section laser shows that the dual-wavelength spectra correspond to emission from ground state. In the hybrid mode-locking regime, a map of locking range is presented by measuring the values of timing jitter for several values of power and frequency of the external electrical modulating signal. An overview of the systematic behavior of InAs/InP(100) quantum dot mode-locked lasers is presented as conclusion.

  8. Local regulation of blood flow evaluated simultaneously by 133-xenon washout and laser Doppler flowmetry

    International Nuclear Information System (INIS)

    Engelhart, M.; Petersen, L.J.; Kristensen, J.K.

    1988-01-01

    The laser Doppler flowmeter and the 133-Xenon washout techniques of measuring cutaneous blood flow were compared for measuring the vasoconstrictor response of the hand during orthostatic maneuvres. Important discrepancies were detected for the two methods. When the hand was lowered by 40 cm a 40% decrease in blood flow was detected by the 133-Xenon method, while a 60% decrease was seen by the laser Doppler technique. Lowering the hand by 50 cm resulted in no further blood flow decrease when using the 133-Xenon method, but an 80% blood flow decrease was recorded with the laser Doppler method. A marked decrease in blood flow was recorded by the laser Doppler technique in hands that were sympathectomized or a hand that was subjected to a nerve blockade, strategies which should eliminate the orthostatic vasoconstrictor response of superficial cutaneous vessels. The 133-Xenon technique did not detect any blood flow changes in hands without sympathetic tone. We found the laser Doppler flowmetry technique unsatisfactory for measurement of blood flow changes that occur in nutritional vessels as this method measures total skin blood flow including non-capillary vessels

  9. Magnetic field sensor based on the Ampere's force using dual-polarization DBR fiber laser

    Science.gov (United States)

    Yao, Shuang; Zhang, Yang; Guan, Baiou

    2015-08-01

    A novel magnetic field sensor using distributed Bragg reflector (DBR) fiber laser by Ampere's force effect is proposed and experimentally demonstrated. The key sensing element, that is the dual-polarization DBR fiber laser, is fixed on the middle part of two copper plates which carry the current. Ampere's force is applied onto the coppers due to an external magnetic field generated by a DC solenoid. Thus, the lateral force from the coppers is converted to a corresponding beat frequency signal shift produced by the DBR laser. The electric current sensing is also realized by the same configuration and same principle simultaneously in an intuitive manner. Good agreement between the theory calculation and the experimental results is obtained, which shows a good linearity. This sensor's sensitivity to the magnetic field and to the electric current finally reaches ~258.92 kHz/mT and ~1.08727 MHz/A, respectively.

  10. Effect of Matrix-Wellbore Flow and Porosity on Pressure Transient Response in Shale Formation Modeling by Dual Porosity and Dual Permeability System

    Directory of Open Access Journals (Sweden)

    Daolun Li

    2015-01-01

    Full Text Available A mathematical dual porosity and dual permeability numerical model based on perpendicular bisection (PEBI grid is developed to describe gas flow behaviors in shale-gas reservoirs by incorporating slippage corrected permeability and adsorbed gas effect. Parametric studies are conducted for a horizontal well with multiple infinite conductivity hydraulic fractures in shale-gas reservoir to investigate effect of matrix-wellbore flow, natural fracture porosity, and matrix porosity. We find that the ratio of fracture permeability to matrix permeability approximately decides the bottom hole pressure (BHP error caused by omitting the flow between matrix and wellbore and that the effect of matrix porosity on BHP is related to adsorption gas content. When adsorbed gas accounts for large proportion of the total gas storage in shale formation, matrix porosity only has a very small effect on BHP. Otherwise, it has obvious influence. This paper can help us understand the complex pressure transient response due to existence of the adsorbed gas and help petroleum engineers to interpret the field data better.

  11. Asymmetric dual-loop feedback to suppress spurious tones and reduce timing jitter in self-mode-locked quantum-dash lasers emitting at 155 μm

    Science.gov (United States)

    Asghar, Haroon; McInerney, John G.

    2017-09-01

    We demonstrate an asymmetric dual-loop feedback scheme to suppress external cavity side-modes induced in self-mode-locked quantum-dash lasers with conventional single and dual-loop feedback. In this letter, we achieved optimal suppression of spurious tones by optimizing the length of second delay time. We observed that asymmetric dual-loop feedback, with large (~8x) disparity in cavity lengths, eliminates all external-cavity side-modes and produces flat RF spectra close to the main peak with low timing jitter compared to single-loop feedback. Significant reduction in RF linewidth and reduced timing jitter was also observed as a function of increased second feedback delay time. The experimental results based on this feedback configuration validate predictions of recently published numerical simulations. This interesting asymmetric dual-loop feedback scheme provides simplest, efficient and cost effective stabilization of side-band free optoelectronic oscillators based on mode-locked lasers.

  12. Measuring cell cycle progression kinetics with metabolic labeling and flow cytometry.

    Science.gov (United States)

    Fleisig, Helen; Wong, Judy

    2012-05-22

    Precise control of the initiation and subsequent progression through the various phases of the cell cycle are of paramount importance in proliferating cells. Cell cycle division is an integral part of growth and reproduction and deregulation of key cell cycle components have been implicated in the precipitating events of carcinogenesis. Molecular agents in anti-cancer therapies frequently target biological pathways responsible for the regulation and coordination of cell cycle division. Although cell cycle kinetics tend to vary according to cell type, the distribution of cells amongst the four stages of the cell cycle is rather consistent within a particular cell line due to the consistent pattern of mitogen and growth factor expression. Genotoxic events and other cellular stressors can result in a temporary block of cell cycle progression, resulting in arrest or a temporary pause in a particular cell cycle phase to allow for instigation of the appropriate response mechanism. The ability to experimentally observe the behavior of a cell population with reference to their cell cycle progression stage is an important advance in cell biology. Common procedures such as mitotic shake off, differential centrifugation or flow cytometry-based sorting are used to isolate cells at specific stages of the cell cycle. These fractionated, cell cycle phase-enriched populations are then subjected to experimental treatments. Yield, purity and viability of the separated fractions can often be compromised using these physical separation methods. As well, the time lapse between separation of the cell populations and the start of experimental treatment, whereby the fractionated cells can progress from the selected cell cycle stage, can pose significant challenges in the successful implementation and interpretation of these experiments. Other approaches to study cell cycle stages include the use of chemicals to synchronize cells. Treatment of cells with chemical inhibitors of key

  13. A tunable and switchable single-longitudinal-mode dual-wavelength fiber laser with a simple linear cavity.

    Science.gov (United States)

    He, Xiaoying; Fang, Xia; Liao, Changrui; Wang, D N; Sun, Junqiang

    2009-11-23

    A simple linear cavity erbium-doped fiber laser based on a Fabry-Perot filter which consists of a pair of fiber Bragg gratings is proposed for tunable and switchable single-longitudinal-mode dual-wavelength operation. The single-longitudinal-mode is obtained by the saturable absorption of an unpumed erbium-doped fiber together with a narrow-band fiber Bragg grating. Under the high pump power (>166 mW) condition, the stable dual-wavelength oscillation with uniform amplitude can be realized by carefully adjusting the polarization controller in the cavity. Wavelength selection and switching are achieved by tuning the narrow-band fiber Bragg grating in the system. The spacing of the dual-wavelength can be selected at 0.20 nm (approximately 25.62 GHz), 0.22 nm (approximately 28.19 GHz) and 0.54 nm (approximately 69.19 GHz).

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

    Science.gov (United States)

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

    2013-01-01

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

  15. Technical discussions II - Flow cytometric analysis

    NARCIS (Netherlands)

    Cunningham, A; Cid, A; Buma, AGJ

    In this paper the potencial of flow cytometry as applied to the aquatic life sciences is discussed. The use of flow cytometry for studying the ecotoxicology of phytoplankton was introduced. On the other hand, the new flow cytometer EUROPA was presented. This is a multilaser machine which has been

  16. Diode-pumped dual-frequency microchip Nd : YAG laser with tunable frequency difference

    Energy Technology Data Exchange (ETDEWEB)

    Ren Cheng; Zhang Shulian, E-mail: ren-c06@mails.tsinghua.edu.c [State Key Laboratory of Precision Measurement Technology and Instruments, Department of Precision Instruments and Mechanology, Tsinghua University, Beijing 100084 (China)

    2009-08-07

    The diode-pumped dual-frequency microchip Nd : YAG laser with tunable frequency difference is presented. The gain medium used is a microchip 2 mm in thickness for miniaturized and integrated design. Two quarter-wave plates are placed into the laser cavity and the intra-cavity birefringence produces two orthogonally linearly polarized modes. The rotation of one of the two quarter-wave plates introduces a controlled and variable cavity birefringence which causes a variable frequency difference between the two orthogonally polarized modes. The frequency difference can be tuned through the whole cavity free spectral range. The obtained frequency difference ranges from 14 MHz to 1.5 GHz. The variation of the beat frequency over a period of 10 min is less than 10 kHz. The lock-in between modes is not found. Experimental results are presented, which match well with the theoretical analysis based on Jones matrices.

  17. Determination of freeze damage on HPV vaccines by use of flow cytometry.

    Science.gov (United States)

    Østergaard, Erik; Frandsen, Peer Lyng; Sandberg, Eva

    2015-07-01

    The human papillomavirus (HPV) vaccines Gardasil, Silgard and Cervarix were labeled with antibodies against HPV strain 6 or 16/FITC conjugated secondary antibodies and analyzed by flow cytometry. The vaccines showed distinct peaks of fluorescent particles, and a shift towards decreased fluorescent particles was observed after incubation of the vaccines over night at -20 °C. Since parallel distributed vaccines could have longer route of transportation there is an increased risk of freeze damage for these types of vaccine. Shift in fluorescence of labeled vaccine particles was used to indicate whether parallel distributed Silgard, which is a vaccine type identical to Gardasil, was exposed to freeze damage during transportation, but no shift was observed. Additional experiments showed that the HPV vaccines could be degraded to smaller particles by citric acid/phosphate buffer treatment. The majority of particles detected in degraded Gardasil were very small indicating that the particles are HPV virus like particle (VLPs) labeled with antibodies, but Cervarix could only be degraded partially due to the presence of another type adjuvant in this vaccine. The described method may be useful in characterization of adjuvanted vaccines with respect to freeze damage, and to characterize vaccines containing particles corresponding to VLPs in size. Copyright © 2015 The International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

  18. Assessment of the Utility of Cytology and Flow Cytometry of Cerebrospinal Fluid Samples in Clinical Practice.

    Science.gov (United States)

    Nam, Anna S; Giorgadze, Tamara; Tam, Wayne; Chadburn, Amy

    2018-01-01

    We sought to assess the utility and limitations of both flow cytometry (FC) and cytology for the analysis of cerebrospinal fluid (CSF) in a practical clinical setting. A total of 393 consecutive CSF samples from 171 patients submitted for both cytomorphologic and FC assessments were analyzed. Both FC and cytology findings were negative for malignancy in 315/393 samples (80%), and either positive (POS) or suspicious/atypical (SUSP/AT) in 7% of samples. This resulted in high agreement between FC and cytology (87%). Minor discrepancies were present in 4% of the cases. In 28 samples, an abnormal population was detected by FC but not by cytology. FC and cytology are important complementary methods for analyzing CSF samples. In cases where cytology is SUSP/AT and FC is inconclusive or negative, additional specimens should be submitted for immunostaining, cytogenetics, and/or molecular studies. © 2018 S. Karger AG, Basel.

  19. Enhancement of conversion efficiency of extreme ultraviolet radiation from a liquid aqueous solution microjet target by use of dual laser pulses

    Science.gov (United States)

    Higashiguchi, Takeshi; Dojyo, Naoto; Hamada, Masaya; Kawasaki, Keita; Sasaki, Wataru; Kubodera, Shoichi

    2006-03-01

    We demonstrated a debris-free, efficient laser-produced plasma extreme ultraviolet (EUV) source by use of a regenerative liquid microjet target containing tin-dioxide (SnO II) nano-particles. By using a low SnO II concentration (6%) solution and dual laser pulses for the plasma control, we observed the EUV conversion efficiency of 1.2% with undetectable debris.

  20. Internal flow measurement in transonic compressor by PIV technique

    Science.gov (United States)

    Wang, Tongqing; Wu, Huaiyu; Liu, Yin

    2001-11-01

    The paper presents some research works conducted in National Key Laboratory of Aircraft Engine of China on the shock containing supersonic flow measurement as well as the internal flow measurement of transoijc compressor by PIC technique. A kind of oil particles in diameter about 0.3 micrometers containing in the flow was discovered to be a very good seed for the PIV measurement of supersonic jet flow. The PIV measurement in over-expanded supersonic free jet and in the flow over wages show a very clear shock wave structure. In the PIV internal flow measurement of transonic compressor a kind of liquid particle of glycol was successful to be used as the seed. An illumination periscope with sheet forming optics was designed and manufactured, it leaded the laser shot generated from an integrate dual- cavity Nd:YAG laser of TSI PIV results of internal flow of an advanced low aspect ratio transonic compressor were shown and discussed briefly.