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Sample records for flow cytometric kinetic

  1. Cell kinetics in a model of artificial skin. An immunohistochemical and flow cytometric analysis

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    A Casasco

    2009-12-01

    Full Text Available Bioengineered organs raised in vitro are candidate substitutes for natural organs in biological, pharmacological and clinical applications. We have studied cell kinetics in a human skin equivalent (HSE using a combined immunohistochemical and flow cytometric approach. Morphological analysis has shown that, relative to unstimulated natural skin, cell proliferation mainly occurs in the basal layer of the epidermal equivalent. Immunohistochemical and flow cytometric measurements of the growth fraction suggested a cell turnover comparable to that of natural skin. Immunohistochemical labelling indices matched well with flow cytometric data. These observations are consistent with morphological and histochemical data demonstrating normal cell differentiation and tissue architecture in HSE and suggest that such HSE may be a usefull substitute for human skin.

  2. Flow cytometric quantification of T cell proliferation and division kinetics in woodchuck model of hepatitis B.

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    Gujar, Shashi A; Michalak, Tomasz I

    2005-01-01

    Woodchucks infected with woodchuck hepatitis virus (WHV) represent the closest natural animal model to study the immunopathogenesis of liver injury caused by essentially noncytopathic, highly human specific hepatitis B virus (HBV). The importance of antiviral T cell response in induction of hepatitis and in control of HBV replication has been demonstrated. However, the understanding of how these responses contribute to the development of different immunomorphological forms of liver disease and their outcomes remain elusive. In this study, we established and standardized a flow cytometry assay using peripheral blood mononuclear cells labeled with carboxyfluorescein diacetate succinimidyl ester (CFSE) to assess WHV-specific and mitogen-driven T lymphocyte proliferative responses in woodchucks. The assay is of significantly greater sensitivity than the adenine incorporation assay currently used when applied to measure either WHV-specific T cell responses in acute (P measuring cell division rates. The study shows that woodchuck PBMC labeled with CFSE exhibit light scatter and fluorescence profiles compatible to those of human PBMC, allowing quantitation and deconvolution of the flow cytometric data by applying the existing analytical softwares. The availability of this novel assay should facilitate a more precise and comprehensive evaluation of hepadnavirus-specific and generalized T cell responses in experimental WHV hepatitis.

  3. Flow cytometric kinetic assay of calcium mobilization in whole blood platelets using Fluo-3 and CD41.

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    do Céu Monteiro, M; Sansonetty, F; Gonçalves, M J; O'Connor, J E

    1999-04-01

    Platelet activation plays a major role in the physiology and pathology of hemostasis. Flow cytometry is a promising approach for the structural and functional analysis of platelets. However, the choice of adequate biological parameters and most technical issues are still under discussion. A rise in cytosolic free Ca2+ is a key early event that follows platelet stimulation and precedes several activation responses, including shape change, aggregation, secretion, and expression of procoagulant activity. Our objective was to set up a fast and sensitive flow cytometric method to determine the kinetics of intracellular Ca2+ mobilization in platelets, which could be performed with the least artifactual perturbation of platelet function. Anticoagulated blood was diluted in Tyrode's buffer and incubated with Fluo-3-acetoxymethyl ester prior to staining with phycoerytrin-conjugated antiplatelet GPIIb/IIIa complex monoclonal antibody. Platelets were identified by a gate including only CD41+ events. After the determination of baseline Fluo-3 green fluorescence on a flow cytometer (EPICS XL-MCL, Coulter Electronics, Hialeah, FL), adequate agonists were added and time-dependent changes in Fluo-3 fluorescence were recorded on-line for up to 3 min. In these conditions, a very fast and transient increase of cytosolic-free Ca2+ was observed following the addition of thrombin, a strong platelet agonist. Stimulation with adenosine diphosphate (ADP), a weak agonist, also resulted in evident increase of Ca2+ levels. Our results show that this flow cytometric kinetic method provides a simple and sensitive tool to assess in vitro the time course and intensity of signal transduction responses to different platelet agonists under near physiological conditions. In this way, it may be useful to evaluate the degree of platelet reactivity and thus to monitor antiplatelet therapy.

  4. Recent advances in flow cytometric cell sorting.

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    Osborne, Geoffrey W

    2011-01-01

    The classification and separation of one cell type or particle from others is a fundamental task in many areas of science. Numerous techniques are available to perform this task; however, electrostatic cell sorting has gained eminence over others because, when combined with the analysis capabilities of flow cytometry it provides flexible separations based on multiple parameters. Unlike competing technologies, such as gradient or magnetic separations that offer much larger total throughput, flow cytometric cell sorting permits selections based on various levels of fluorescent reporters, rather the complete presence or absence of the reporter. As such, this technology has found application in a huge range of fields. This chapter aims to describe the utility of single-cell sorting with particular emphasis given to index sorting. This is followed by two recently developed novel techniques of sorting cells or particles. The first of these is positional sorting which is useful in cell-based studies where sorting can proceed and produce meaningful results without being inherently dependant on prior knowledge of where gates should be set. Secondly, reflective plate sorting is introduced which positionally links multiwell sample and collection plates in a convenient assay format so that cells in the collection plate "reflect" those in the sample plate.

  5. Howard University Flow Cytometric Sorter For Research and Education

    Science.gov (United States)

    2015-08-04

    SECURITY CLASSIFICATION OF: Howard University’s newly acquired Fluorescence Activated Cytometric Sorter (FACS) has been integrated into the new flow...Research Administrative Services Washington, DC 20059 -0001 11-Mar-2015 ABSTRACT Number of Papers published in peer-reviewed journals : Number of Papers...published in non peer-reviewed journals : Final Report: Howard University Flow Cytometric Sorter For Research and Education Report Title Howard

  6. Flow cytometric enumeration of marine viral populations at low abundances

    NARCIS (Netherlands)

    Mojica, K.D.A.; Evans, C.; Brussaard, C.P.D.

    2014-01-01

    Flow cytometric enumeration has advanced our ability to analyze aquatic virus samples and thereby our understanding of the ecological role viruses play in the oceans. However, low virus abundances are underestimated using the current flow cytometry (FCM) protocol. Our results revealed that low

  7. Flow cytometric immunoassay for sulfonamides in raw milk

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    Keizer, de W.; Bienenmann-Ploum, M.; Bergwerff, A.A.; Haasnoot, W.

    2008-01-01

    Sulfonamide antibiotics are applied in veterinary medicine for the treatment of microbial infections. For the detection of residues of sulfonamides in milk, a multi-sulfonamide flow cytometric immunoassay (FCI) was developed using the Luminex MultiAnalyte Profiling (xMAP) technology. In this automat

  8. Technical discussions II - Flow cytometric analysis

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    Cunningham, A; Cid, A; Buma, AGJ

    1996-01-01

    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 sp

  9. Technical discussions II - Flow cytometric analysis

    NARCIS (Netherlands)

    Cunningham, A; Cid, A; Buma, AGJ

    1996-01-01

    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 sp

  10. Flow cytometric detection of aberrant chromosomes

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    Gray, J.W.; Lucas, J.; Yu, L.C.; Langlois, R.

    1983-05-11

    This report describes the quantification of chromosomal aberrations by flow cytometry. Both homogeneously and heterogeneously occurring chromosome aberrations were studied. Homogeneously occurring aberrations were noted in chromosomes isolated from human colon carcinoma (LoVo) cells, stained with Hoechst 33258 and chromomycin A3 and analyzed using dual beam flow cytometry. The resulting bivariate flow karyotype showed a homogeneously occurring marker chromosome of intermediate size. Heterogeneously occurring aberrations were quantified by slit-scan flow cytometry in chromosomes isolated from control and irradiated Chinese hamster cells and stained with propidium iodide. Heterogeneously occurring dicentric chromosomes were detected by their shapes (two centrometers). The frequencies of such chromosomes estimated by slit-scan flow cytometry correlated well with the frequencies determined by visual microscopy.

  11. Flow cytometric studies of human osteosarcoma.

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    Mankin, H J; Gebhardt, M C; Springfield, D S; Litwak, G J; Kusazaki, K; Rosenberg, A E

    1991-09-01

    A number of recent studies have emphasized the potential value of flow cytometry as a "marker" to assess the malignity and therefore to help predict the biologic behavior of neoplasms, including bone tumors. Using propidium iodide and a home-built flow cytometer, the authors have studied the DNA distribution in 95 patients with osteosarcoma and determined the percentage of cells in diploidy, S-phase, tetraploidy, and aneuploidy. Using these values and a derived one, mean DNA concentration, it was possible to demonstrate the extent of the abnormalities observed in this group of neoplasms and show their severity as compared with the normal pattern. When the data are compared against disease-free survival and total survival, correlations were noted that, although weak, suggested that some patterns were predictive of increased risk of metastasis and death. The effect of treatment could also be assessed by evaluating the pattern before and after chemotherapy and correlating these with survival. It seems likely that with some improvement in technology, flow cytometry will be of value in the future in assessing the prognosis for osteosarcoma and predicting whether treatment has been effective.

  12. Flow cytometric detection method for DNA samples

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    Nasarabadi,Shanavaz (Livermore, CA); Langlois, Richard G. (Livermore, CA); Venkateswaran, Kodumudi S. (Round Rock, TX)

    2011-07-05

    Disclosed herein are two methods for rapid multiplex analysis to determine the presence and identity of target DNA sequences within a DNA sample. Both methods use reporting DNA sequences, e.g., modified conventional Taqman.RTM. probes, to combine multiplex PCR amplification with microsphere-based hybridization using flow cytometry means of detection. Real-time PCR detection can also be incorporated. The first method uses a cyanine dye, such as, Cy3.TM., as the reporter linked to the 5' end of a reporting DNA sequence. The second method positions a reporter dye, e.g., FAM.TM. on the 3' end of the reporting DNA sequence and a quencher dye, e.g., TAMRA.TM., on the 5' end.

  13. Epstein-Barr Virus, Human Papillomavirus, and Flow Cytometric Cell Cycle Kinetics in Nasopharyngeal Carcinoma and Inverted Papilloma among Egyptian Patients

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    S. K. Kassim

    1998-01-01

    Full Text Available It is widely accepted that the Epstein-Barr virus is etiologically associated with the development of nasopharyngeal carcinoma. The human papillomavirus is also associated with inverted papilloma. We used the polymerase chain reaction technique to detect both viruses in both types of tumors. Flow cytometry was also used to study the DNA pattern and proliferative behavior of the tumors in relation to the viruses. EBV was detected in 13/20 (65% of NPC specimens, and in none of IP (n = 10 or control specimens (n = 10. This indicates the contribution of EBV as an etiologic factor in NPC. Five cases of NPC (25% were positive for HPV 16, two of them were EBV positive. Four HPV 16 positive cases were found among cases with inverted papilloma, but none among the control cases. Flow cytometry revealed that all NPC, IP, and control samples were diploid except one aneuploid NPC sample. Proliferative capacity (PC of primary tumors was predictive of tumor recurrence in NPC. Using 13.6% as a cut-off point for PC, we were able to discriminate between high risk and low risk groups with 100% sensitivity and 86% specificity. PC can be used as a baseline prognostic parameter in NPC, making it possible to modify courses of treatment in an attempt to inhibit tumor recurrence.

  14. Flow-cytometric Analysis of Bacillus anthracis Spores

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    D. V. Kamboj

    2006-11-01

    Full Text Available Flow-cytometric technique has been established as a powerful tool for detection andidentification of microbiological agents. Unambiguous and rapid detection of Bacillus anthracisspores has been reported using immunoglobulin G-fluorescein isothiocyanate conjugate againstlive spores. In addition to the high sensitivity, the present technique could differentiate betweenspores of closely related species, eg, Bacillus cereus and Bacillus subtilis using fluorescenceintensity. The technique can be used for detection of live as well as inactivated spores makingit more congenial for screening of suspected samples of bioterrorism.

  15. Automated High-Dimensional Flow Cytometric Data Analysis

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    Pyne, Saumyadipta; Hu, Xinli; Wang, Kui; Rossin, Elizabeth; Lin, Tsung-I.; Maier, Lisa; Baecher-Allan, Clare; McLachlan, Geoffrey; Tamayo, Pablo; Hafler, David; de Jager, Philip; Mesirov, Jill

    Flow cytometry is widely used for single cell interrogation of surface and intracellular protein expression by measuring fluorescence intensity of fluorophore-conjugated reagents. We focus on the recently developed procedure of Pyne et al. (2009, Proceedings of the National Academy of Sciences USA 106, 8519-8524) for automated high- dimensional flow cytometric analysis called FLAME (FLow analysis with Automated Multivariate Estimation). It introduced novel finite mixture models of heavy-tailed and asymmetric distributions to identify and model cell populations in a flow cytometric sample. This approach robustly addresses the complexities of flow data without the need for transformation or projection to lower dimensions. It also addresses the critical task of matching cell populations across samples that enables downstream analysis. It thus facilitates application of flow cytometry to new biological and clinical problems. To facilitate pipelining with standard bioinformatic applications such as high-dimensional visualization, subject classification or outcome prediction, FLAME has been incorporated with the GenePattern package of the Broad Institute. Thereby analysis of flow data can be approached similarly as other genomic platforms. We also consider some new work that proposes a rigorous and robust solution to the registration problem by a multi-level approach that allows us to model and register cell populations simultaneously across a cohort of high-dimensional flow samples. This new approach is called JCM (Joint Clustering and Matching). It enables direct and rigorous comparisons across different time points or phenotypes in a complex biological study as well as for classification of new patient samples in a more clinical setting.

  16. Uncovering Aberrant Mutant PKA Function with Flow Cytometric FRET

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    Shin-Rong Lee

    2016-03-01

    Full Text Available Biology has been revolutionized by tools that allow the detection and characterization of protein-protein interactions (PPIs. Förster resonance energy transfer (FRET-based methods have become particularly attractive as they allow quantitative studies of PPIs within the convenient and relevant context of living cells. We describe here an approach that allows the rapid construction of live-cell FRET-based binding curves using a commercially available flow cytometer. We illustrate a simple method for absolutely calibrating the cytometer, validating our binding assay against the gold standard isothermal calorimetry (ITC, and using flow cytometric FRET to uncover the structural and functional effects of the Cushing-syndrome-causing mutation (L206R on PKA’s catalytic subunit. We discover that this mutation not only differentially affects PKAcat’s binding to its multiple partners but also impacts its rate of catalysis. These findings improve our mechanistic understanding of this disease-causing mutation, while illustrating the simplicity, general applicability, and power of flow cytometric FRET.

  17. An automated flow cytometric micronucleus assay for human lymphocytes.

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    Schreiber, G A; Beisker, W; Braselmann, H; Bauchinger, M; Bögl, K W; Nüsse, M

    1992-12-01

    A new flow cytometric method is presented for scoring micronuclei (MN) in human lymphocytes after in vitro gamma-irradiation. Fifty to fifty-five hours after PHA-stimulation, the frequency of micronuclei per nucleus and the fraction of cells in the second cell cycle were measured using flow cytometry. All data were automatically analysed using our DAS-software package. Eight individual linear-quadratic dose response curves derived from five donors revealed inter- and intra-individual variabilities of all curve parameters. Since also an age dependence was found for spontaneous MN-frequencies and for the linear curve parameter, a combined linear-quadratic age-dose-effect model was used to fit the data. The 90% prediction intervals show that a reliable individual dose estimation for donors aged between 23 and 54 years cannot be achieved for exposures below 1 Gy.

  18. Comparative flow cytometric analysis of immunofunctionalized nanowire and nanoparticle signatures.

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    Prina-Mello, Adriele; Whelan, Aine M; Atzberger, Ann; McCarthy, Joseph E; Byrne, Fiona; Davies, Gemma-Louise; Coey, J M D; Volkov, Yuri; Gun'ko, Yurii K

    2010-01-01

    Flow cytometry is one of the gold-standard techniques used in clinical medicine for quantitative immunoassaying. The continuous development of its probes, commonly fluorescent nanoparticles, is important. Lately, the introduction of quantitative multiplexed immunoassay has challenged the use of nanoparticles as probes. Functionalized fluorescent silica-based magnetic nanowires are investigated under flow cytometry as a novel probe category. The preparation and full characterization of these multimodal nanowires is reported and compared to those of silica-based magnetic nanoparticles by flow cytometry. Full characterization includes transmission electron microscopy and fluorescence microscopy imaging, flow cytometric assaying, superconducting quantum interference device (SQUID) magnetization, and Mössbauer spectroscopy measurements. This work shows that loaded silica nanowires have intrinsic geometrical advantages when compared to similar spherical particles due to their unique "flow cytometry fingerprint" when utilized as magnetic carriers for immunodetection applications. These advantages account for a 17% yield in detecting the functional binding between THP-1 and ICAM-1, by utilizing a much lower concentration than that required for the nanoparticles.

  19. Flow cytometric detection of anti-gliadin antibodies.

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    Presani, G; Perticarari, S; Mangiarotti, M A

    1989-05-12

    A very sensitive solid-phase fluorescent immunoassay to detect anti-alpha-gliadin IgA class antibodies is described. The solid phase consisted of polystyrene carboxylated microspheres, of 5 microns diameter, coated with alpha-gliadin. Serum-specific antibodies bound to the alpha-gliadin were measured by flow cytometry using fluorescein-conjugated anti-human IgA. 41 samples were tested and the results compared with those obtained by a standard method: an enzyme-linked immunosorbent assay (ELISA). A good correlation was found between the two techniques (r = 0.96). The sera of untreated coeliac children showed significantly higher antibody values than the sera of children on a gluten-free diet or healthy control groups. The flow cytometric method was more sensitive when the Kolgomorov/Smirnov test was used to analyse the histograms. This method provides an alternative screening test for coeliac disease and may also be used to confirm borderline results obtained in the ELISA test.

  20. Flow cytometric detection of immunoglobulin light chain in hematolymphoid immunophenotyping

    Institute of Scientific and Technical Information of China (English)

    Xu Dongsheng

    2011-01-01

    During B cell development and maturation, the antigen receptor,which is encoded by the immunoglobulin heavy-(IgH)and light chain genes,rearrange to associate one of a number of variable, diverse, and joining gene segments. A single mature Bcell expresses an IgH chain and either a kappa or lambda light chain, which is known as allelic or isotypic exclusion. In normal or reactive conditions, lymphoid cells comprise the mixtures of lymphocytes with either kappa or lambda expression. The norreal proportion of kappa to lambda (κ/λ ratio) is within the range of 0. 5 - 3. 0 in peripheral blood or bone marrow and 1.2 2.7 in lymph nodes[1].The most useful feature for diagnosing mature B cell neoplasm is light chain restriction or monotypic staining with κ or λ light chain. Currently, it is a common assumption that demonstration of light chain restriction in a B lymphocyte population is generally considered proof of monoclonality and indicates malignancy although monotypic B cell populations have been infrequently demonstrated in patients with no definitive evidence ofB cell malignancy[2-4]. The most common flow cytometric analysis for determining B cell monotype is the percent κ and λimmunoglobulin light chains.Because of the importance of light chain restriction in the diagnosis of B cell neoplasm, anti immunoglobulin antibodies (e. g. anti-κ and anti-λ) are vital tools in the detection of monotypic B cell populations. Accurate determination of surface light chain expression depends on many factors, such as proper washing procedure, lysing solution, type of antibody used, specimen type or lymphoma type[5]. This article will discuss some common problems encountered in flow cytometric(FCM)deterruination of surface immunoglobulin light chain expression inhematolymphoid immunophenotyping.%@@ During B-cell development and maturation,the antigen receptor,which is encoded by the immunoglobulin heavy-(IgH) and light-chain genes,rearrange to associate one of a number of

  1. Flow cytometric and laser scanning microscopic approaches in epigenetics research.

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    Szekvolgyi, Lorant; Imre, Laszlo; Minh, Doan Xuan Quang; Hegedus, Eva; Bacso, Zsolt; Szabo, Gabor

    2009-01-01

    Our understanding of epigenetics has been transformed in recent years by the advance of technological possibilities based primarily on a powerful tool, chromatin immunoprecipitation (ChIP). However, in many cases, the detection of epigenetic changes requires methods providing a high-throughput (HTP) platform. Cytometry has opened a novel approach for the quantitative measurement of molecules, including PCR products, anchored to appropriately addressed microbeads (Pataki et al. 2005. Cytometry 68, 45-52). Here we show selected examples for the utility of two different cytometry-based platforms of epigenetic analysis: ChIP-on-beads, a flow-cytometric test of local histone modifications (Szekvolgyi et al. 2006. Cytometry 69, 1086-1091), and the laser scanning cytometry-based measurement of global epigenetic modifications that might help predict clinical behavior in different pathological conditions. We anticipate that such alternative tools may shortly become indispensable in clinical practice, translating the systematic screening of epigenetic tags from basic research into routine diagnostics of HTP demand.

  2. A Flow Cytometric and Computational Approaches to Carbapenems Affinity to the Different Types of Carbapenemases

    Science.gov (United States)

    Pina-Vaz, Cidália; Silva, Ana P.; Faria-Ramos, Isabel; Teixeira-Santos, Rita; Moura, Daniel; Vieira, Tatiana F.; Sousa, Sérgio F.; Costa-de-Oliveira, Sofia; Cantón, Rafael; Rodrigues, Acácio G.

    2016-01-01

    The synergy of carbapenem combinations regarding Enterobacteriaceae producing different types of carbapenemases was study through different approaches: flow cytometry and computational analysis. Ten well characterized Enterobacteriaceae (KPC, verona integron-encoded metallo-β-lactamases –VIM and OXA-48-like enzymes) were selected for the study. The cells were incubated with a combination of ertapenem with imipenem, meropenem, or doripenem and killing kinetic curves performed with and without reinforcements of the drugs. A cephalosporin was also used in combination with ertapenem. A flow cytometric assay with DiBAC4-(3), a membrane potential dye, was developed in order to evaluate the cellular lesion after 2 h incubation. A chemical computational study was performed to understand the affinity of the different drugs to the different types of enzymes. Flow cytometric analysis and time-kill assays showed a synergic effect against KPC and OXA-48 producing-bacteria with all combinations; only ertapenem with imipenem was synergic against VIM producing-bacteria. A bactericidal effect was observed in OXA-48-like enzymes. Ceftazidime plus ertapenem was synergic against ESBL-negative KPC producing-bacteria. Ertapenem had the highest affinity for those enzymes according to chemical computational study. The synergic effect between ertapenem and others carbapenems against different carbapenemase-producing bacteria, representing a therapeutic choice, was described for the first time. Easier and faster laboratorial methods for carbapenemase characterization are urgently needed. The design of an ertapenem derivative with similar affinity to carbapenemases but exhibiting more stable bonds was demonstrated as highly desirable. PMID:27555844

  3. A flow cytometric and computational approaches to carbapenems affinity to the different types of carbapenemases

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    Cidália Pina-Vaz

    2016-08-01

    Full Text Available The synergy of carbapenem combinations regarding Enterobacteriaceae producing different types of carbapenemases was study through different approaches: flow cytometry and computa-tional analysis. Ten well characterized Enterobacteriaceae (KPC, verona integron-encoded metallo-β-lactamases –VIM and OXA-48-like enzymes were selected for the study. The cells were incubated with a combination of ertapenem with imipenem, meropenem or doripenem and killing kinetic curves performed with and without reinforments of the drugs. A cephalosporin was also used in combination with ertapenem. A flow cytometric assay with DiBAC4-(3, a membrane potential dye, was developed in order to evaluate the cellular lesion after 2 h incuba-tion. A chemical computational study was performed to understand the affinity of the different drugs to the different types of enzymes. Flow cytometric analysis and time-kill assays showed a synergic effect against KPC and OXA-48 producing-bacteria with all combinations; only ertapenem with imipenem was synergic against VIM producing-bacteria. A bactericidal effect was observed in OXA-48-like enzymes. Ceftazidime plus ertapenem was synergic against ESBL-negative KPC producing-bacteria. Ertapenem had the highest affinity for those enzymes according to chemical computational study. The synergic effect between ertapenem and others carbapenems against different carbapenemase-producing bacteria, representing a therapeutic choice, was described for the first time. Easier and faster laboratorial methods for car-bapenemase characterization are urgently needed. The design of an ertapenem derivative with similar affinity to carbapenemases but exhibiting more stable bonds was demonstrated as highly desirable.

  4. Flow cytometric immunophenotypic characteristics of plasma cell leukemia

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    Barbara Kruk

    2011-04-01

    Full Text Available The aim of this prospective study was to define the flow cytometric characteristics of simultaneously investigated bone marrow and peripheral blood plasma cells antigens expression in 36 plasma cell leukemia (PCL patients. The immunophenotypic profile of plasma cells was determined with a panel of monoclonal antibodies. The antigen expression intensity was calculated as relative fluorescence intensity (RFI. Bone marrow plasma cells showed expression of particular antigens in the following proportion of cases: CD49d 100%, CD29 94%, CD54 93%, CD44 83%, CD56 60%, CD18 26%, CD11b 29%, CD11a 19%, CD117 27%, CD71 30%, CD126 100% and CD19 0%, while the expression of those antigens on peripheral blood plasma cells was present in the following percentage of patients: CD49d 100%, CD29 96%, CD54 93%, CD44 95%, CD56 56%, CD18 50%, CD11b 53%, CD11a 29%, CD117 26%, CD71 28%, CD126 100% and CD19 0%. The expression of CD54 was significantly higher than that of adhesion molecules belonging to the integrin b2 family: CD11a, CD18 and CD11b, on both bone marrow and peripheral blood cells (p < 0.01. Expression of CD18, CD11a and CD11b was differential between two cell compartments: lower on bone marrow and higher on peripheral blood cells. We found that plasma cells in the bone marrow of patients with plasma cell leukaemia showed significantly greater granularity and size than those in the peripheral blood (p = 0.0001 and p = 0.04, respectively. However, no differences in cell size or granularity were revealed between bone marrow plasma cells from patients with PCL and multiple myeloma. In conclusion, impaired expression of adhesion molecules such as CD11a/CD18 (LFA-1 or CD56 may explain hematogenic dissemination characterizing PCL. The following pattern of adhesion molecule expression according to the proportion of plasma cells expressing a given antigen in peripheral blood and bone marrow and arranged in diminishing order may be established: CD49d > CD44 > CD54

  5. Flow cytometric DNA analysis of ducks accumulating 137Cs on a reactor reservoir.

    Science.gov (United States)

    George, L S; Dallas, C E; Brisbin, I L; Evans, D L

    1991-06-01

    The objective of this study was to detect red blood cell (rbc) DNA abnormalities in male, game-farm mallard ducks as they ranged freely and accumulated 137Cs (radiocesium) from an abandoned nuclear reactor cooling reservoir. Prior to release, the ducks were tamed to enable recapture at will. Flow cytometric measurements conducted at intervals during the first year of exposure yielded cell cycle percentages of DNA (G0/G1, S, G2 + M phases) of rbc, as well as coefficients of variation (CV) in the G0/G1 phase. DNA histograms of exposed ducks were compared with two sets of controls which were maintained 30 and 150 miles from the study site. 137Cs live wholebody burdens were also measured in these animals in a parallel kinetics study, and an approximate steady-state equilibrium was attained after about 8 months. DNA histograms from 2 of the 14 contaminated ducks revealed DNA aneuploid-like patterns after 9 months exposure. These two ducks were removed from the experiment at this time, and when sampled again 1 month later, one continued to exhibit DNA aneuploidy. None of the control DNA histograms demonstrated DNA aneuploid-like patterns. There were no significant differences in cell cycle percentages at any time point between control and exposed animals. A significant increase in CV was observed at 9 months exposure, but after removal of the two ducks with DNA aneuploidy, no significant difference was detected in the group monitored after 12 months exposure. An increased variation in the DNA and DNA aneuploidy could, therefore, be detected in duck rbc using flow cytometric analysis, with the onset of these effects being related to the attainment of maximal levels of 137Cs body burdens in the exposed animals.

  6. Cluster Analysis of Flow Cytometric List Mode Data on a Personal Computer

    NARCIS (Netherlands)

    Bakker Schut, Tom C.; Bakker schut, T.C.; de Grooth, B.G.; Greve, Jan

    1993-01-01

    A cluster analysis algorithm, dedicated to analysis of flow cytometric data is described. The algorithm is written in Pascal and implemented on an MS-DOS personal computer. It uses k-means, initialized with a large number of seed points, followed by a modified nearest neighbor technique to reduce

  7. Long-term storage of samples for flow cytometric DNA analysis

    DEFF Research Database (Denmark)

    Vindeløv, L L; Christensen, I J; Keiding, N

    1983-01-01

    A simple procedure for long-term storage of cells for flow cytometric DNA analysis was developed and tested. The cells were stored as single cells or fine-needle aspirates suspended in a citrate buffer with dimethylsulfoxide (DMSO), or as small blocks of tissue from solid tumors. The cells were...

  8. Flow cytometric sorting of paraffin-embedded tumor tissues considerably improves molecular genetic analysis

    NARCIS (Netherlands)

    Jordanova, ES; Corver, WE; Vonk, MJ; Leers, MPG; Riemersma, SA; Schuuring, E; Kluin, PM

    2003-01-01

    The characterization of genetic aberrations in paraffin-embedded tumor material is impaired by contaminating normal cells. In the present study on the genetic causes of loss of HLA expression in diffuse large B-cell lymphoma (DLBCL), we compared the efficacy of microdissection with flow cytometric s

  9. Flow cytometric determination of circulating immune complexes with the indirect granulocyte phagocytosis test

    NARCIS (Netherlands)

    Terstappen, L.W.M.M.; Grooth, de B.G.; Nolten, G.M.J.; Napel, ten C.H.H.; Berkel, van W.; Greve, J.

    1985-01-01

    A method for the determination of circulating immune complexes (CIC) was adapted for flow cytometric analysis. Human granulocytes were used to phagocytose IgG-bearing CIC of serum from systemic lupus erythematosus (SLE) patients. A method for labeling the phagocytosed CIC with FITC-conjugated anti-h

  10. Discrimination of bromodeoxyuridine labelled and unlabelled mitotic cells in flow cytometric bromodeoxyuridine/DNA analysis

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J K; Christensen, I J

    1994-01-01

    Bromodeoxyuridine (BrdUrd) labelled and unlabelled mitotic cells, respectively, can be discriminated from interphase cells using a new method, based on immunocytochemical staining of BrdUrd and flow cytometric four-parameter analysis of DNA content, BrdUrd incorporation, and forward and orthogona...

  11. A flow cytometric technique for quantification and differentiation of bacteria in bulk tank milk

    DEFF Research Database (Denmark)

    Holm, C.; Mathiasen, T.; Jespersen, Lene

    2004-01-01

    AIMS: The present study describes a flow cytometric technique for quantification and differentiation of bacteria in bulk tank milk according to the main cause of elevated counts. METHODS AND RESULTS: A total of 75 Danish bulk tank milk samples exceeding the grading level of 3.0 x 10(4) CFU ml(-1)...

  12. A flow-cytometric method to evaluate eosinophil-mediated uptake of probiotic Lactobacillus reuteri.

    Science.gov (United States)

    Kraemer, Laura S; Brenner, Todd A; Krumholz, Julia O; Rosenberg, Helene F

    2017-03-27

    Eosinophils are resident leukocytes of gut mucosa. Here we present a combined flow cytometric-antibiotic protection assay to identify mouse eosinophils capable of bacterial uptake, specifically, Gram-positive Lactobacillus reuteri, in studies performed ex vivo. The assay may be adapted for use in vivo.

  13. Simultaneous preparation of RNA and nuclei for Northern blot and flow cytometric analysis

    Energy Technology Data Exchange (ETDEWEB)

    Tesfaigzi, J.; Jaramillo, R. [Inhalation Toxicology Research Inst., Albuquerque, NM (United States)

    1996-11-01

    Several methods have been developed to quantify RNA synthesis during the progression of the cell cycle. The rate of RNA synthesis can be detected during different stages of the cell cycle by staining cells with agents that intercalate with nucleic acids. For example, following staining of mammalian cells with acridin orange, the green and red fluorescence that correlates with DNA and RNA content, respectively, can be analyzed by flow cytometry. Increase in RNA content during the progression of cells through the cell cycle can be measured after staining with acridin orange. RNA synthesis resulting from the stimulation of quiescent cells with various growth factors has also been demonstrated by labeling cells with bromo-uridine and using the anti-bromo-deoxyuridine antibody. These methods allow measurement of the overall RNA content in cells; however, they do not allow the measurement of the levels of specific mRNAs throughout the cell cycle. Current methods to quantify specific mRNAs generally require the preparation of a large number of cells (5--10 {times} 10{sup 6} cells) to carry out flow cytometric analyses and to isolate RNA for Northern blot analysis or solution hybridization. In this report, the authors describe a method of simultaneously preparing RNA and nuclei for Northern blot and flow cytometric analyses, respectively. The minimum number of nuclei required to obtain flow cytometric data and the effect of conserving nuclei in methanol for several days are also presented.

  14. Results of external quality control study in flow cytometric acute lymphoblastic leukemia diagnostics

    Directory of Open Access Journals (Sweden)

    A. M. Popov

    2016-01-01

    Full Text Available Comparison of interpretation of acute lymphoblastic leukemia (ALL flow cytometric diagnostics data was the aim of the study. Immunophenotyping data obtained from 10 patients with ALL were analysed separately in 26 laboratories from Russian Federation and Kazahstan. Results comparison showed four main type of discordance: B-lineage ALL diagnostics during heavy bone marrow regeneration, great variability of T-ALL interpretation, complexity of ambiguous lineage acute leukemia and, finally, very different report types, unique for each laboratory. All these problems are the serious obstacles for standardization of flow cytometric ALL diagnostics in multicenter setting. Continuation of similar QC rounds following by consecutive discussions with further development of consensus diagnostic algorithm could be the first step for standardization of ALL immunophenotyping in Russian Federation and CIS countries.

  15. Flow cytometric assessment of viability of lactic acid bacteria

    NARCIS (Netherlands)

    Bunthof, C.J.; Bloemen, K.; Breeuwer, P.; Rombouts, F.M.; Abee, T.

    2001-01-01

    The viability of lactic acid bacteria is crucial for their applications as dairy starters and as probiotics. We investigated the usefulness of flow cytometry (FCM) for viability assessment of lactic acid bacteria. The esterase substrate carboxyfluorescein diacetate (cFDA) and the dye exclusion DNA b

  16. Flow cytometric DNA ploidy analysis of ovarian granulosa cell tumors

    NARCIS (Netherlands)

    D. Chadha; C.J. Cornelisse; A. Schabert (A.)

    1990-01-01

    textabstractAbstract The nuclear DNA content of 50 ovarian tumors initially diagnosed as granulosa cell tumors was measured by flow cytometry using paraffin-embedded archival material. The follow-up period of the patients ranged from 4 months to 19 years. Thirty-eight tumors were diploid or near-dip

  17. Detachment and flow cytometric quantification of seagrass-associated bacteria.

    Science.gov (United States)

    Trevathan-Tackett, Stacey; Macreadie, Peter; Ralph, Peter; Seymour, Justin

    2014-07-01

    A new protocol was developed to detach bacteria from seagrass tissue and subsequently enumerate cells using flow cytometry (FCM). A method involving addition of the surfactant Tween 80 and vortexing resulted in maximum detachment efficiency of seagrass attached bacteria, providing a robust protocol for precisely enumerating seagrass-associated bacteria with FCM. Using this approach we detected cell concentrations between 2.0×10(5) and 8.0×10(6)cells mg(-1) DW tissue.

  18. Strategies for immunophenotyping and purifying classical Hodgkin lymphoma cells from lymph nodes by flow cytometry and flow cytometric cell sorting.

    Science.gov (United States)

    Fromm, Jonathan R; Wood, Brent L

    2012-07-01

    Flow cytometry is an established technique to immunophenotype hematopoietic neoplasms. While the diagnosis of classical Hodgkin lymphoma (CHL) has commonly been made using paraffin sections, we have recently demonstrated that the neoplastic Hodgkin and Reed-Sternberg (HRS) cells of CHL can be identified by flow cytometry. Using 6- and 9-color flow cytometric assays, CHL can be immunophenotyped with 85-90% sensitivity and nearly 100% specificity. Analysis of this data requires using established gating strategies to help in the identification of putative HRS cell populations. Interestingly, HRS cells bind to reactive T cells (HRS-T cell rosetting) and this phenomenon can be identified and utilized diagnostically by flow cytometry. In addition, the reactive T cells of CHL show characteristic immunophenotypic changes by flow cytometry and these changes can suggest a diagnosis of CHL. Finally, these principles can be employed to rapidly purify HRS cells using flow cytometric cell sorting. This manuscript provides experimental protocols for immunophenotyping CHL by flow cytometry as well as purifying the HRS cells via flow cytometric cell sorting.

  19. Flow cytometric fluorescence lifetime analysis of DNA binding fluorochromes

    Energy Technology Data Exchange (ETDEWEB)

    Crissman, Harry A.; Cui, H. H. (H. Helen); Steinkamp, J. A.

    2002-01-01

    Most flow cytometry (FCM) applications monitor fluorescence intensity to quantitate the various cellular parameters; however, the fluorescence emission also contains information relative to the fluorescence lifetime. Recent developments in FCM (Pinsky et al., 1993; Steinkamp & Crissman, 1993; Steinkamp et al., 1993), provide for the measurement of fluorescence lifetime which is also commonly referred to as fluorescence decay, or the time interval in which a fluorochrome remains in the excited state. Many unbound fluorochromes have characteristic lifetime values that are determined by their molecular structure; however, when the probe becomes bound, the lifetime value is influenced by a number of factors that affect the probe interaction with a target molecule. Monitoring the changes in the lifetime of the probe yields information relating to the molecular conformation, the functional state or activity of the molecular target. In addition, the lifetime values can be used as signatures to resolve the emissions of multiple fluorochrome labels with overlapping emission spectra that cannot be resolved by conventional FCM methodology. Such strategies can increase the number of fluorochrome combinations used in a flow cytometer with a single excitation source. Our studies demonstrate various applications of lifetime measurements for the analysis of the binding of different fluorochromes to DNA in single cells. Data presented in this session will show the utility of lifetime measurements for monitoring changes in chromatin structure associated with cell cycle progression, cellular differentiation, or DNA damage, such as induced during apoptosis. Several studies show that dyes with specificity for nucleic acids display different lifetime values when bound to DNA or to dsRNA. The Phase Sensitive Flow Cytometer is a multiparameter instrument, capable of performing lifetime measurements in conjunction with all the conventional FCM measurements. Future modifications of this

  20. Flow cytometric data analysis of circulating progenitor cell stability.

    Science.gov (United States)

    Mahar, Ernestine A; Mou, Liping; Hayek, Salim S; Quyyumi, Arshed A; Waller, Edmund K

    2017-02-01

    A recent publication by Mekonnen et al. demonstrated that among women with non-obstructive coronary artery disease, higher levels of circulating progenitor cells in the blood (CPC), were associated with impaired coronary flow reserve [1]. We performed a quality control assessment of the stability of circulating blood progenitor cells in blood samples stored at 4 °C, to determine the time period during which blood samples can be analyzed and yield consistent data for progenitor cell content. Healthy volunteers (n=6) were recruited and underwent phlebotomy, and blood was stored in EDTA tubes at 4 °C. Flow cytometry was performed to quantitate progenitor cell subsets at 0-4 h, 24 h, and 48 h post phlebotomy. All processed samples were fixed with 1% Paraformaldehyde and 1,000,000 total data events were collected. We found no significant differences in PC data for both CD34+ (P=0.68 for one-way ANOVA) and CD34+/CD133+ (P=0.74 for one-way ANOVA).

  1. Flow cytometric data analysis of circulating progenitor cell stability

    Directory of Open Access Journals (Sweden)

    Ernestine A. Mahar

    2017-02-01

    We performed a quality control assessment of the stability of circulating blood progenitor cells in blood samples stored at 4 °C, to determine the time period during which blood samples can be analyzed and yield consistent data for progenitor cell content. Healthy volunteers (n=6 were recruited and underwent phlebotomy, and blood was stored in EDTA tubes at 4 °C. Flow cytometry was performed to quantitate progenitor cell subsets at 0–4 h, 24 h, and 48 h post phlebotomy. All processed samples were fixed with 1% Paraformaldehyde and 1,000,000 total data events were collected. We found no significant differences in PC data for both CD34+ (P=0.68 for one-way ANOVA and CD34+/CD133+ (P=0.74 for one-way ANOVA.

  2. DNA flow cytometric analysis in variable types of hydropic placentas

    Directory of Open Access Journals (Sweden)

    Fatemeh Atabaki pasdar

    2015-05-01

    Full Text Available Background: Differential diagnosis between complete hydatidiform mole, partial hydatidiform mole and hydropic abortion, known as hydropic placentas is still a challenge for pathologists but it is very important for patient management. Objective: We analyzed the nuclear DNA content of various types of hydropic placentas by flowcytometry. Materials and Methods: DNA ploidy analysis was performed in 20 non-molar (hydropic and non-hydropic spontaneous abortions and 20 molar (complete and partial moles, formalin-fixed, paraffin-embedded tissue samples by flow cytometry. The criteria for selection were based on the histopathologic diagnosis. Results: Of 10 cases histologically diagnosed as complete hydatiform mole, 9 cases yielded diploid histograms, and 1 case was tetraploid. Of 10 partial hydatidiform moles, 8 were triploid and 2 were diploid. All of 20 cases diagnosed as spontaneous abortions (hydropic and non-hydropic yielded diploid histograms. Conclusion: These findings signify the importance of the combined use of conventional histology and ploidy analysis in the differential diagnosis of complete hydatidiform mole, partial hydatidiform mole and hydropic abortion.

  3. Ultrastructural and flow cytometric analyses of lipid accumulation in microalgae

    Energy Technology Data Exchange (ETDEWEB)

    Solomon, J.A.; Hand, R.E. Jr.; Mann, R.C.

    1986-12-01

    Lipid accumulation in three species of microalgae was investigated with flow cytometry (FCM) and transmission electron microscopy (TEM). Previous studies using batch cultures of a algae have led to the assumption that lipid accumulation in microalgae is a gradual process requiring at least several days for completion. However, FCM reveals, through changes in the chlorophyll:lipid ratio, that the time span required for individual cells to change metabolic state is short. Simultaneous FCM measurements of chlorophyll and nile red (neutral lipid) fluorescence in individual cells of nitrogen-deficient Isochrysis populations revealed a bimodal population distribution as one stage in the lipid accumulation process. The fact that two discrete populations exist, with few cells in an intermediate stage, suggests rapid response to a liqid trigger. Interpretations of light and electron microscopic observations are consistent with this hypothesis. The time required for an entire population to achieve maximum lipid content is considerably longer than that required for a single cell, due to the variation in response time among cells. In this study high lipid cultures were sometimes obtained by using FCM to separate high lipid cells from the remainder of the population. FCM holds much promise for strain enhancement but considerable developmental work, directed at providing more consistent results, remains to be done. 8 refs., 35 figs.

  4. IVBT-documented platelet function correlates with flow cytometric data.

    Science.gov (United States)

    Hoffmann, J; Bonacker, G; Kretschmer, V; Schulzki, T; Heimanns, J

    1996-12-01

    Thrombocytopenic patients with identical platelet counts often show different bleeding tendencies owing to significant differences in the platelet function. This could be demonstrated by the in vitro bleeding test (IVBT). Using flow cytometry, we tried to find characteristics of platelet antigen expression in order to explain these differences in function. Thirty patients with bone marrow hypoplasia receiving 65 platelet transfusions (mainly from a cell separator) were observed for 3 to 29 days. Size, granulation and fluorescence of platelet-rich plasma (n = 522 samples) were evaluated using monoclonal antibodies against GP IIIb (collagen receptor), GP IIb/IIIa (fibrinogen receptor) and GP Ib (thrombin receptor). We defined separate gates for each antibody using the results from 50 normals and by laying an orthograde cross over the gate to divide the gate into four equal quadrants. The platelet populations were divided into four different groups according to the occlusion time (OT) of the IVBT and the Simplate time (ST). The thrombocytes with the most impaired function (OT > or = 485 s/ST > 30 min) had significantly less platelet fluorescence when marked with antibodies against GP IIIb and GP Ib than those with short OT and ST (OT platelet fluorescence when marked with anti-GP IIIb and anti-GP Ib than thrombocytopenic patients, who had a spontaneous platelet rise beyond 30,000 platelets/microliters a few days later. One day after platelet transfusion, significantly more platelets with high GP IIIb and Ib expression could be found. We were also able to document better transfusion efficacy of platelet concentrates with high GP IIIb and Ib expression. Finally, patients with high bleeding scores showed less GP Ib expression on the platelets than patients with low bleeding scores. In summary, the IVBT-documented platelet function clearly corresponded to an increased expression of the collagen receptor and the thrombin receptor of platelets.

  5. Karyological and flow cytometric evidence of triploid specimens in Bufo viridis (Amphibia Anura

    Directory of Open Access Journals (Sweden)

    D Cavallo

    2010-01-01

    Full Text Available Karyological and flow cytometric (FCM analyses were performed on a group of 14 green toads of the Bufo viridis species from seven Eurasian populations. Both approaches gave concordant results concerning the DNA ploidy level. All the populations examined were represented exclusively by diploid or tetraploid specimens, except one, where triploids were found. Results evidenced an interpopulation variability in DNA content against the same ploidy level, as well as an unusually high number of triploids in a particular reproductive place. The origin of polyploidy and the presence and persistence of a high number of triploids in a particular population are discussed.

  6. A flow cytometric method for characterization of circulating cell-derived microparticles in plasma

    DEFF Research Database (Denmark)

    Nielsen, Morten Hjuler; Beck-Nielsen, Henning; Andersen, Morten Nørgaard;

    2014-01-01

    BACKGROUND AND AIM: Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold...... of detection of a new generation BD FACSAria™ III digital flow cytometer. METHODS: We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (.... The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined. RESULTS: By using a flow cytometric approach we...

  7. Two and three-color fluorescence flow cytometric analysis of immunoidentified viable bacteria.

    Science.gov (United States)

    Barbesti, S; Citterio, S; Labra, M; Baroni, M D; Neri, M G; Sgorbati, S

    2000-07-01

    Traditional culture methods well established in the past and still in use are not able to detect the environmental microorganisms that exist in a viable but not culturable state. A number of different fluorescence-based assays have been developed over the past decade to detect and identify viable bacteria in the environment. We have developed a simple and rapid method for measuring the number and viability of immunolabeled bacteria by means of a two/three color fluorescence flow cytometric analysis. After washing, cultured bacteria in suspension were labeled with a rabbit polyclonal antibody recognizing the wall lipopolysaccharide complex. A secondary biotinylated anti-rabbit polyclonal antibody was added allowing the cells to be labeled with the streptavidin R-phycoerythrin-Cyanine 5 (RPE-Cy5) fluorochrome. Before flow cytometric analysis, bacterial suspensions were stained with SYBR Green I and propidium iodide which stain all of the cells and the non viable ones, respectively. With the appropriate filter sets of both Bryte-HS (Bio-Rad, Hercules, CA) and FACScan (Becton Dickinson, San Jose, CA) flow cytometers, the measurement of separated green (SYBR Green I), orange-red (propidium iodide), and far red (RPE-Cy5) fluorescence was possible, allowing the enumeration of viable immunodetected bacteria. The entire protocol is completed in less than 3 h, offering numerous possibilities for rapid and precise analyses in sanitary, industrial, and environmental microbiology. Copyright 2000 Wiley-Liss, Inc.

  8. Flow cytometric methods to investigate culture heterogeneities for plant metabolic engineering.

    Science.gov (United States)

    Gaurav, Vishal; Kolewe, Martin E; Roberts, Susan C

    2010-01-01

    Plant cell cultures provide an important method for production and supply of a variety of natural products, where conditions can be easily controlled, manipulated, and optimized. Development and optimization of plant cell culture processes require both bioprocess engineering and metabolic engineering approaches. Cultures are generally highly heterogeneous, with significant variability amongst cells in terms of growth, metabolism, and productivity of key metabolites. Taxus cultures produce the important anti-cancer agent Taxol((R)) (i.e., paclitaxel) and have demonstrated significant variability amongst cell populations in culture with regard to paclitaxel accumulation, cell cycle participation, and protein synthesis. To fully understand the link between cellular metabolism and culture behavior and to enable targeted metabolic engineering approaches, cultures need to be studied at a single cell level. This chapter describes the application of plant cell flow cytometric techniques to investigate culture heterogeneity at the single cell level, in order to optimize culture performance through targeted metabolic engineering. Flow cytometric analytical methods are described to study Taxus single cells, protoplasts, and nuclei suspensions with respect to secondary metabolite accumulation, DNA content, cell size, and complexity. Reproducible methods to isolate these single particle suspensions from aggregated Taxus cultures are discussed. Methods to stain both fixed and live cells for a variety of biological markers are provided to enable characterization of cell phenotypes. Fluorescence-activated cell sorting (FACS) methods are also presented to facilitate isolation of certain plant cell culture populations for both analysis and propagation of superior cell lines for use in bioprocesses.

  9. Flow cytometric analysis of microbial contamination in food industry technological lines – initial study

    Directory of Open Access Journals (Sweden)

    Katarzyna Czaczyk

    2012-06-01

    Full Text Available Background. Flow cytometry constitutes an alternative for traditional methods of microorganisms identifi cation and analysis, including methods requiring cultivation step. It enables the detection of pathogens and other microorganisms contaminants without the need to culture microbial cells meaning that the sample (water, waste or food e.g. milk, wine, beer may be analysed directly. This leads to a signifi cant reduction of time required for analysis allowing monitoring of production processes and immediate reaction in case of contamination or any disruption occurs. Apart from the analysis of raw materials or products on different stages of manufacturing process, the fl ow cytometry seems to constitute an ideal tool for the assessment of microbial contamination on the surface of technological lines. Material and methods. In the present work samples comprising smears from 3 different surfaces of technological lines from fruit and vegetable processing company from Greater Poland were analysed directly with fl ow cytometer. The measured parameters were forward and side scatter of laser light signals allowing the estimation of microbial cell contents in each sample. Results. Flow cytometric analysis of the surface of food industry production lines enable the preliminary evaluation of microbial contamination within few minutes from the moment of sample arrival without the need of sample pretreatment. Conclusions. The presented method of fl ow cytometric initial evaluation of microbial state of food industry technological lines demonstrated its potential for developing a robust, routine method for the rapid and laborsaving detection of microbial contamination in food industry.

  10. Flow-Cytometric Isolation of Human Antibodies from a Nonimmune Saccharomyces cerevisiae Surface Display Library

    Energy Technology Data Exchange (ETDEWEB)

    Feldhaus, Michael (BATTELLE (PACIFIC NW LAB)); Siegel, Robert W.(BATTELLE (PACIFIC NW LAB)); Opresko, Lee (BATTELLE (PACIFIC NW LAB)); Coleman, James R.(BATTELLE (PACIFIC NW LAB)); Feldhaus, Jane M.(BATTELLE (PACIFIC NW LAB)); Yeung, Yik A.(Massachusetts Institute Of Tec); Cochran, Jennifer R.(Massachusetts Institute Of Tec); Heinzelman, Peter (Massachusetts Institute Of Tec); Colby, David (Massachusetts Institute Of Tec); Swers, Jeffrey (Massachusetts Institute Of Tec); Graff, Christilyn (Massachusetts Institute Of Tec); Wiley, H Steven (BATTELLE (PACIFIC NW LAB)); Wittrup, K D.(Massachusetts Institute Of Tec)

    2003-02-28

    A nonimmune library of 109 human antibody scFv fragments has been cloned and expressed on the surface of yeast, and nanomolar-affinity scFvs routinely obtained by magnetic bead screening and flow cytometric sorting. The yeast library can be amplified 1010-fold without measurable loss of clonal diversity, enabling effectively indefinite expansion of the library. The expression, stability, and antigen binding properties of more than 50 isolated scFv clones were assessed directly on the yeast cell surface by immunofluorescent labeling and flow cytometry, obviating separate subcloning, expression, and purification steps and thereby expediting the isolation of novel affinity reagents. The ability to use multiplex library screening demonstrates the utility of this approach for high throughput antibody isolation for proteomics applications.

  11. Procarbazine effects on spermatogenesis in golden hamster: a flow cytometric evaluation.

    Science.gov (United States)

    Weissenberg, R; Golan, R; Shochat, L; Lewin, L M

    2002-01-01

    The response of hamster testis to the administration of 450mg/kg procarbazine (PCB) over a period of 4 weeks was evaluated. Flow cytometry was used to investigate changes in cell populations in testicular single cell suspensions and to correlate these changes with those observed in histological sections. PCB caused significant decrease in testicular and epididymal weight and a drastic reduction in haploid cells and spermatogenic arrest, demonstrating variation among the test animals. The results obtained confirm previous observations concerning detrimental effects of PCB upon spermatogenesis in species such as the rat and mouse, though its effect on hamster testis is milder and does not include the germinal stem cells. The histological evaluation of the testis showed a good correlation with flow cytometric evaluation, emphasizing the usefulness of this method in providing quantitative and rapid results.

  12. Multiplex competitive microbead-based flow cytometric immunoassay using quantum dot fluorescent labels

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Hye-Weon; Kim, In S. [School of Environmental Science and Engineering, Gwangju Institute of Science and Technology (GIST), 261 Cheomdan-gwagiro, Buk-gu, Gwangju (Korea, Republic of); Niessner, Reinhard [Chair for Analytical Chemistry, Institute of Hydrochemistry, Technische Universitaet Muenchen, Marchioninistrasse 17, 81377 Muenchen (Germany); Knopp, Dietmar, E-mail: dietmar.knopp@ch.tum.de [Chair for Analytical Chemistry, Institute of Hydrochemistry, Technische Universitaet Muenchen, Marchioninistrasse 17, 81377 Muenchen (Germany)

    2012-10-31

    Highlights: Black-Right-Pointing-Pointer First time, duplex competitive bead-based flow cytometric immunoassay was developed using ODs. Black-Right-Pointing-Pointer Antibody-coated QD detection probes and antigen-immobilized microspheres were synthesized. Black-Right-Pointing-Pointer The two model target analytes were low molecular weight compounds of microbial and chemical origin. Black-Right-Pointing-Pointer The determination of different water types was possible after simple filtration of samples. - Abstract: In answer to the ever-increasing need to perform the simultaneous analysis of environmental hazards, microcarrier-based multiplex technologies show great promise. Further integration with biofunctionalized quantum dots (QDs) creates new opportunities to extend the capabilities of multicolor flow cytometry with their unique fluorescence properties. Here, we have developed a competitive microbead-based flow cytometric immunoassay using QDs fluorescent labels for simultaneous detection of two analytes, bringing the benefits of sensitive, rapid and easy-of-manipulation analytical tool for environmental contaminants. As model target compounds, the cyanobacterial toxin microcystin-LR and the polycyclic aromatic hydrocarbon compound benzo[a]pyrene were selected. The assay was carried out in two steps: the competitive immunological reaction of multiple targets using their exclusive sensing elements of QD/antibody detection probes and antigen-coated microsphere, and the subsequent flow cytometric analysis. The fluorescence of the QD-encoded microsphere was thus found to be inversely proportional to target analyte concentration. Under optimized conditions, the proposed assay performed well within 30 min for the identification and quantitative analysis of the two environmental contaminants. For microcystin-LR and benzo[a]pyrene, dose-response curves with IC{sub 50} values of 5 {mu}g L{sup -1} and 1.1 {mu}g L{sup -1} and dynamic ranges of 0.52-30 {mu}g L{sup -1} and 0

  13. Flow Cytometric Analysis of T, B, and NK Cells Antigens in Patients with Mycosis Fungoides

    Science.gov (United States)

    Yazıcı, Serkan; Bülbül Başkan, Emel; Budak, Ferah; Oral, Barbaros; Adim, Şaduman Balaban; Ceylan Kalin, Zübeyde; Özkaya, Güven; Aydoğan, Kenan; Saricaoğlu, Hayriye; Tunali, Şükran

    2015-01-01

    We retrospectively analyzed the clinicopathological correlation and prognostic value of cell surface antigens expressed by peripheral blood mononuclear cells in patients with mycosis fungoides (MF). 121 consecutive MF patients were included in this study. All patients had peripheral blood flow cytometry as part of their first visit. TNMB and histopathological staging of the cases were retrospectively performed in accordance with International Society for Cutaneous Lymphomas/European Organization of Research and Treatment of Cancer (ISCL/EORTC) criteria at the time of flow cytometry sampling. To determine prognostic value of cell surface antigens, cases were divided into two groups as stable and progressive disease. 17 flow cytometric analyses of 17 parapsoriasis (PP) and 11 analyses of 11 benign erythrodermic patients were included as control groups. Fluorescent labeled monoclonal antibodies were used to detect cell surface antigens: T cells (CD3+, CD4+, CD8+, TCRαβ+, TCRγδ+, CD7+, CD4+CD7+, CD4+CD7−, and CD71+), B cells (HLA-DR+, CD19+, and HLA-DR+CD19+), NKT cells (CD3+CD16+CD56+), and NK cells (CD3−CD16+CD56+). The mean value of all cell surface antigens was not statistically significant between parapsoriasis and MF groups. Along with an increase in cases of MF stage statistically significant difference was found between the mean values of cell surface antigens. Flow cytometric analysis of peripheral blood cell surface antigens in patients with mycosis fungoides may contribute to predicting disease stage and progression. PMID:26788525

  14. Flow Cytometric Analysis of T, B, and NK Cells Antigens in Patients with Mycosis Fungoides

    Directory of Open Access Journals (Sweden)

    Serkan Yazıcı

    2015-01-01

    Full Text Available We retrospectively analyzed the clinicopathological correlation and prognostic value of cell surface antigens expressed by peripheral blood mononuclear cells in patients with mycosis fungoides (MF. 121 consecutive MF patients were included in this study. All patients had peripheral blood flow cytometry as part of their first visit. TNMB and histopathological staging of the cases were retrospectively performed in accordance with International Society for Cutaneous Lymphomas/European Organization of Research and Treatment of Cancer (ISCL/EORTC criteria at the time of flow cytometry sampling. To determine prognostic value of cell surface antigens, cases were divided into two groups as stable and progressive disease. 17 flow cytometric analyses of 17 parapsoriasis (PP and 11 analyses of 11 benign erythrodermic patients were included as control groups. Fluorescent labeled monoclonal antibodies were used to detect cell surface antigens: T cells (CD3+, CD4+, CD8+, TCRαβ+, TCRγδ+, CD7+, CD4+CD7+, CD4+CD7−, and CD71+, B cells (HLA-DR+, CD19+, and HLA-DR+CD19+, NKT cells (CD3+CD16+CD56+, and NK cells (CD3−CD16+CD56+. The mean value of all cell surface antigens was not statistically significant between parapsoriasis and MF groups. Along with an increase in cases of MF stage statistically significant difference was found between the mean values of cell surface antigens. Flow cytometric analysis of peripheral blood cell surface antigens in patients with mycosis fungoides may contribute to predicting disease stage and progression.

  15. Improved flow cytometric assessment reveals distinct microvesicle (cell-derived microparticle signatures in joint diseases.

    Directory of Open Access Journals (Sweden)

    Bence György

    Full Text Available INTRODUCTION: Microvesicles (MVs, earlier referred to as microparticles, represent a major type of extracellular vesicles currently considered as novel biomarkers in various clinical settings such as autoimmune disorders. However, the analysis of MVs in body fluids has not been fully standardized yet, and there are numerous pitfalls that hinder the correct assessment of these structures. METHODS: In this study, we analyzed synovial fluid (SF samples of patients with osteoarthritis (OA, rheumatoid arthritis (RA and juvenile idiopathic arthritis (JIA. To assess factors that may confound MV detection in joint diseases, we used electron microscopy (EM, Nanoparticle Tracking Analysis (NTA and mass spectrometry (MS. For flow cytometry, a method commonly used for phenotyping and enumeration of MVs, we combined recent advances in the field, and used a novel approach of differential detergent lysis for the exclusion of MV-mimicking non-vesicular signals. RESULTS: EM and NTA showed that substantial amounts of particles other than MVs were present in SF samples. Beyond known MV-associated proteins, MS analysis also revealed abundant plasma- and immune complex-related proteins in MV preparations. Applying improved flow cytometric analysis, we demonstrate for the first time that CD3(+ and CD8(+ T-cell derived SF MVs are highly elevated in patients with RA compared to OA patients (p=0.027 and p=0.009, respectively, after Bonferroni corrections. In JIA, we identified reduced numbers of B cell-derived MVs (p=0.009, after Bonferroni correction. CONCLUSIONS: Our results suggest that improved flow cytometric assessment of MVs facilitates the detection of previously unrecognized disease-associated vesicular signatures.

  16. A flow cytometric in vivo chalone assay using retransplanted old murine JB-1 ascites tumour cells.

    Science.gov (United States)

    Barfod, N M

    1981-07-01

    A flow cytometric in vivo chalone assay is described. Transplantation of old JB-1 ascites tumour cells to new hosts induced an influx of tumour cells, with G1 DNA content, to the S phase. This induction could be reversibly and specifically blocked by injections of an ultrafiltrate of old JB-1 ascites fluid. The method described is superior to a previously published in vivo chalone assay using regenerating ascites tumours. Owing to a reduced variability in time of onset of DNA synthesis, a smaller scatter of observations is achieved and thus the number of mice per group may be reduced using the new method. In contrast to the older technique, the present one does not necessitate killing of mice during the observation period.

  17. Standardizing flow cytometric assays in long-term population-based studies

    Science.gov (United States)

    Melzer, Susanne; Bocsi, Jozsef; Tárnok, Attila

    2015-03-01

    Quantification of leukocyte subpopulations and characterization of antigen-expression pattern on the cellular surface can play an important role in diagnostics. The state of cellular immunology on the single-cell level was analyzed by polychromatic flow cytometry in a recent comparative study within the average Leipzig population (LIFE - Leipzig Research Centre for Civilization Diseases). Data of 1699 subjects were recorded over a long-time period of three years (in a total of 1126 days). To ensure compatibility of such huge data sets, quality-controls on many levels (stability of instrumentation, low intra-laboratory variance and reader independent data analysis) are essential. The LIFE study aims to analyze various cytometric pattern to reveal the relationship between the life-style, the environmental effects and the individual health. We therefore present here a multi-step quality control procedure for long-term comparative studies.

  18. Biotinylation of interleukin-2 (IL-2) for flow cytometric analysis of IL-2 receptor expression. Comparison of different methods

    NARCIS (Netherlands)

    M.O. de Jong (Marg); H. Rozemuller (Henk); J.G.J. Bauman (J. G J); J.W.M. Visser (Jan)

    1995-01-01

    textabstractThe main prerequisites for the use of biotinylated ligands to study the expression of growth factor receptors on heterogeneous cell populations, such as peripheral blood or bone marrow, by flow cytometric methods, are that the biotinylated ligand retains its binding ability and that bind

  19. Biotinylation of interleukin-2 (IL-2) for flow cytometric analysis of IL-2 receptor expression. Comparison of different methods

    NARCIS (Netherlands)

    M.O. de Jong (Marg); H. Rozemuller (Henk); J.G.J. Bauman (J. G J); J.W.M. Visser (Jan)

    1995-01-01

    textabstractThe main prerequisites for the use of biotinylated ligands to study the expression of growth factor receptors on heterogeneous cell populations, such as peripheral blood or bone marrow, by flow cytometric methods, are that the biotinylated ligand retains its binding ability and that bind

  20. A short-term in vitro test for tumour sensitivity to adriamycin based on flow cytometric DNA analysis

    DEFF Research Database (Denmark)

    Engelholm, S A; Spang-Thomsen, M; Vindeløv, L L

    1983-01-01

    A new method to test the sensitivity of tumour cells to chemotherapy is presented. Tumour cells were incubated in vitro on agar, and drug-induced cell cycle perturbation was monitored by flow cytometric DNA analysis. In the present study the method was applied to monitor the effect of adriamycin...

  1. Clonal heterogeneity of small-cell anaplastic carcinoma of the lung demonstrated by flow-cytometric DNA analysis

    DEFF Research Database (Denmark)

    Vindeløv, L L; Hansen, H H; Christensen, I J

    1980-01-01

    Flow-cytometric DNA analysis yields information on ploidy and proliferative characteristics of a cell population. The analysis was implemented on small-cell anaplastic carcinoma of the lung using a rapid detergent technique for the preparation of fine-needle aspirates for DNA determination...

  2. Simple flow cytometric detection of haemozoin containing leukocytes and erythrocytes for research on diagnosis, immunology and drug sensitivity testing

    NARCIS (Netherlands)

    Frita, R.; Rebelo, M.; Pamplona, A.; Vigario, A.M.; Mota, M.M.; Grobusch, M.P.; Haenscheid, T.

    2011-01-01

    Background: Malaria pigment (haemozoin, Hz) has been the focus of diverse research efforts. However, identification of Hz-containing leukocytes or parasitized erythrocytes is usually based on microscopy, with inherent limitations. Flow cytometric detection of depolarized Side-Scatter is more accurat

  3. A simple and sensitive flow cytometric assay for determination of the cytotoxic activity of human natural killer cells

    NARCIS (Netherlands)

    Radosevic, Katarina; Radosevic, K.; Garritsen, Henk S.P.; Garritsen, H.S.P.; van Graft, M.; van Graft, Marja; de Grooth, B.G.; Greve, Jan

    1990-01-01

    A new, simple and sensitive flow cytometric assay for the determination of the cytotoxic activity of human natural killer cells is described. The assay is based on the use of two fluorochromes. The target cell population is stained with one fluorochrome (octadecylamine-fluorescein isothiocyanate,

  4. Identification and purification of classical Hodgkin cells from lymph nodes by flow cytometry and flow cytometric cell sorting.

    Science.gov (United States)

    Fromm, Jonathan R; Kussick, Steven J; Wood, Brent L

    2006-11-01

    We demonstrate the feasibility of using flow cytometry (FC) to identify the Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin lymphoma (CHL). Initial flow cytometric studies of the HRS cell line L1236 demonstrated potentially useful antigens for identifying HRS cells. L1236 cells spontaneously bound normal T cells, analogous to the T-cell rosetting of HRS cells seen in tissue sections of CHL, but these interactions could be blocked by using a cocktail of unlabeled antibodies to 4 adhesion molecules. Among 27 lymph nodes involved by CHL, FC enabled HRS cells to be identified in 89%, whereas none of 29 non-CHL neoplasms or 23 reactive lymph nodes demonstrated HRS populations. Of the CHL cases, 82% demonstrated interactions between HRS cells and T cells that could be disrupted with blocking antibodies. Flow cytometric cell sorting experiments demonstrated typical HRS cytomorphologic features among the purified cells. FC may offer an alternative to immunohistochemical analysis in confirming the diagnosis of CHL in certain cases, and, through cell sorting, it provides a means of rapidly isolating pure HRS cells.

  5. Flow cytometric scoring of micronucleated erythrocytes: an efficient platform for assessing in vivo cytogenetic damage.

    Science.gov (United States)

    Dertinger, Stephen D; Torous, Dorothea K; Hayashi, Makoto; MacGregor, James T

    2011-01-01

    The relative simplicity of the micronucleated erythrocyte endpoint has made it amenable to automated scoring approaches. Flow cytometry is one such scoring platform that has been employed successfully. This review describes the evolution and properties of flow cytometry-based scoring of micronucleated erythrocytes. The methodology has become widely applied to rodent blood specimens and the high throughput nature of the technology provides a number of advantages over manual microscopic scoring. For instance, the ability to efficiently survey many dose levels and many more cells per specimen relative to microscopy benefits studies that are designed to identify no observable effect levels or lowest observable effect levels. Furthermore, flow cytometry makes it practical to study species with low spontaneous reticulocyte (RET) counts and micronucleus (MN) frequencies, thereby facilitating integration of blood-based micronucleated reticulocyte (MN-RET) frequency measurements into experiments conducted across species of toxicological interest. This capability enhances genotoxicity assessments that have historically been made in dedicated MN tests performed in one species. Importantly, the feasibility of using MN-RET frequencies in blood from humans as an index of genetic damage in bone marrow opens a critical area of application that had not been practical previously. We conclude with recommendations for additional work that is needed to more fully realise the potential of flow cytometric in vivo MN scoring.

  6. A new spreadsheet method for the analysis of bivariate flow cytometric data

    Directory of Open Access Journals (Sweden)

    Isacke Clare M

    2004-03-01

    Full Text Available Abstract Background A useful application of flow cytometry is the investigation of cell receptor-ligand interactions. However such analyses are often compromised due to problems interpreting changes in ligand binding where the receptor expression is not constant. Commonly, problems are encountered due to cell treatments resulting in altered receptor expression levels, or when cell lines expressing a transfected receptor with variable expression are being compared. To overcome this limitation we have developed a Microsoft Excel spreadsheet that aims to automatically and effectively simplify flow cytometric data and perform statistical tests in order to provide a clearer graphical representation of results. Results To demonstrate the use and advantages of this new spreadsheet method we have investigated the binding of the transmembrane adhesion receptor CD44 to its ligand hyaluronan. In the first example, phorbol ester treatment of cells results in both increased CD44 expression and increased hyaluronan binding. By applying the spreadsheet method we effectively demonstrate that this increased ligand binding results from receptor activation. In the second example we have compared AKR1 cells transfected either with wild type CD44 (WT CD44 or a mutant with a truncated cytoplasmic domain (CD44-T. These two populations do not have equivalent receptor expression levels but by using the spreadsheet method hyaluronan binding could be compared without the need to generate single cell clones or FACS sorting the cells for matching CD44 expression. By this method it was demonstrated that hyaluronan binding requires a threshold expression of CD44 and that this threshold is higher for CD44-T. However, at high CD44-T expression, binding was equivalent to WT CD44 indicating that the cytoplasmic domain has a role in presenting the receptor at the cell surface in a form required for efficient hyaluronan binding rather than modulating receptor activity. Conclusion

  7. The influence of fixation delay on mitotic activity and flow cytometric cell cycle variables.

    Science.gov (United States)

    Bergers, E; Jannink, I; van Diest, P I; Cuesta, M A; Meyer, S; van Mourik, J C; Baak, J P

    1997-01-01

    Proliferation variables such as mitotic activity and the percentage of S-phase cells have been shown to be of prognostic value in many tumors, especially in breast cancer. However, some studies reported a decrease in mitotic activity caused by delay in fixation of the tissue. In contrast, other studies showed that the identifiability of mitotic figures decreases after fixation delay, but the total number of mitotic figures and also the percentage of S-phase cells remain unchanged. Most studies have been done on small numbers of experimental tumors, thus introducing the risk of selection bias. The aim of this study was to reinvestigate the influence of fixation delay on mitotic activity and cell cycle variables assessed by flow cytometry in an adequate number of resected human tissues to reach firmer conclusions. Resection specimens of 19 and 21 cases, respectively, for the mitotic activity estimate and the flow cytometric percentage of S-phase calculation were collected directly from the operating theater using lung, breast, and intestinal cancers and normal intestinal mucosa. The tissues were cut in pieces, and from each specimen, pieces were fixed in 4% buffered formaldehyde (for mitosis counting) as well as snap frozen (for flow cytometry) immediately after excision, as well as after a fixation delay of 1, 2, 4, 6, 8, 18, and 24 hours. Moreover, during the fixation delay, one series from each specimen was kept in the refrigerator and the second at room temperature. Thus, a total of 304 (19 X 16) and 336 (21 X 16) specimens were investigated for the mitotic activity estimate and the percentage of S-phase cells calculation, respectively. With regard to the estimation of the mitotic activity, both clear and doubtful mitotic figures were registered separately, obtaining an "uncorrected" and "corrected" (for doubtful mitotic figures) mitotic activity estimate. The percentage of S-phase cells was obtained by cell cycle analysis of flow cytometric DNA-histograms. The

  8. A novel flow cytometric assay for measurement of In Vivo pulmonary neutrophil phagocytosis

    Directory of Open Access Journals (Sweden)

    Gentry-Nielsen Martha J

    2006-07-01

    Full Text Available Abstract Background Phagocytosis assays are traditionally performed in vitro using polymorphonuclear leukocytes (PMNs isolated from peripheral blood or the peritoneum and heat-killed, pre-opsonized organisms. These assays may not adequately mimic the environment within the infected lung. Our laboratory therefore has developed a flow cytometric in vivo phagocytosis assay that enables quantification of PMN phagocytosis of viable bacteria within the lungs of rats. In these studies, rats are injected transtracheally with lipopolysaccharide (LPS to recruit PMNs to their lungs. They are then infected with live 5(-and 6 carboxyfluorescein diacetate succinimidyl ester (CFDA/SE labeled type 3 Streptococcus pneumoniae. Bronchoalveolar lavage is performed and resident alveolar macrophages and recruited PMNs are labeled with monoclonal antibodies specific for surface epitopes on each cell type. Three color flow cytometry is utilized to identify the cell types, quantify recruitment, and determine uptake of the labeled bacteria. Results The viability of the alveolar macrophages and PMNs isolated from the lavage fluid was >95%. The values of the percentage of PMNs in the lavage fluid as well as the percentage of PMNs associated with CFSE-labeled S. pneumoniae as measured through flow cytometry showed a high degree of correlation with the results from manual counting of cytospin slides. Conclusion This assay is suitable for measuring bacterial uptake within the infected lung. It can be adapted for use with other organisms and/or animal model systems.

  9. A novel flow cytometric hemozoin detection assay for real-time sensitivity testing of Plasmodium falciparum.

    Directory of Open Access Journals (Sweden)

    Maria Rebelo

    Full Text Available Resistance of Plasmodium falciparum to almost all antimalarial drugs, including the first-line treatment with artemisinins, has been described, representing an obvious threat to malaria control. In vitro antimalarial sensitivity testing is crucial to detect and monitor drug resistance. Current assays have been successfully used to detect drug effects on parasites. However, they have some limitations, such as the use of radioactive or expensive reagents or long incubation times. Here we describe a novel assay to detect antimalarial drug effects, based on flow cytometric detection of hemozoin (Hz, which is rapid and does not require any additional reagents. Hz is an optimal parasite maturation indicator since its amount increases as the parasite matures. Due to its physical property of birefringence, Hz depolarizes light, hence it can be detected using optical methods such as flow cytometry. A common flow cytometer was adapted to detect light depolarization caused by Hz. Synchronized in vitro cultures of P. falciparum were incubated for 48 hours with several antimalarial drugs. Analysis of depolarizing events, corresponding to parasitized red blood cells containing Hz, allowed the detection of parasite maturation. Moreover, chloroquine resistance and the inhibitory effect of all antimalarial drugs tested, except for pyrimethamine, could be determined as early as 18 to 24 hours of incubation. At 24 hours incubation, 50% inhibitory concentrations (IC50 were comparable to previously reported values. These results indicate that the reagent-free, real-time Hz detection assay could become a novel assay for the detection of drug effects on Plasmodium falciparum.

  10. Evaluation of Red Cell Membrane Cytoskeletal Disorders Using a Flow Cytometric Method in South Iran

    Directory of Open Access Journals (Sweden)

    Habib Alah Golafshan

    2014-03-01

    Full Text Available OBJECTIVE: The diagnosis of hereditary red blood cell (RBC membrane disorders, and in particular hereditary spherocytosis (HS and Southeast Asian ovalocytosis (SAO, is based on clinical history, RBC morphology, and other conventional tests such as osmotic fragility. However, there are some milder cases of these disorders that are difficult to diagnose. The application of eosin-5’-maleimide (EMA was evaluated for screening of RBC membrane defects along with some other anemias. We used EMA dye, which binds mostly to band 3 protein and to a lesser extent some other membrane proteins, for screening of some membrane defects such as HS. METHODS: Fresh RBCs from hematologically normal controls and patients with HS, SAO, hereditary elliptocytosis, hereditary spherocytosis with pincered cells, severe iron deficiency, thalassemia minor, and autoimmune hemolytic anemia were stained with EMA dye and analyzed for mean fluorescent intensity (MFI using a flow cytometer. RESULTS: RBCs from patients with HS and iron deficiency showed a significant reduction in MFI compared to those from normal controls (p<0.0001 and p<0.001, respectively, while macrocytic RBCs showed a significant increase in MFI (p<0.01. A significant correlation was shown between mean corpuscular volume and MFI, with the exceptions of HS and thalassemia minor. CONCLUSION: Our results showed that the flow cytometric method could be a reliable diagnostic method for screening and confirmation, with higher sensitivity and specificity (95% and 93%, respectively than conventional routine tests for HS patients prior to further specific membrane protein molecular tests.

  11. Flow cytometric analysis of oil palm: a preliminary analysis for cultivars and genomic DNA alteration

    Directory of Open Access Journals (Sweden)

    Warawut Chuthammathat

    2005-12-01

    Full Text Available DNA contents of oil palm (Elaeis guineensis Jacq. cultivars were analyzed by flow cytometry using different external reference plant species. Analysis using corn (Zea mays line CE-777 as a reference plant gave the highest DNA content of oil palm (4.72±0.23 pg 2C-1 whereas the DNA content was found to be lower when using soybean (Glycine max cv. Polanka (3.77±0.09 pg 2C-1 or tomato (Lycopersicon esculentum cv. Stupicke (4.25±0.09 pg 2C-1 as a reference. The nuclear DNA contents of Dura (D109, Pisifera (P168 and Tenera (T38 cultivars were 3.46±0.04, 3.24±0.03 and 3.76±0.04 pg 2C-1 nuclei, respectively, using soybean as a reference. One haploid genome of oil palm therefore ranged from 1.56 to 1.81±109 base pairs. DNA contents from one-year-old calli and cell suspension of oil palm were found to be significantly different from those of seedlings. It thus should be noted that genomic DNA alteration occurred in these cultured tissues. We therefore confirm that flow cytometric analysis could verify cultivars, DNA content and genomic DNA alteration of oil palm using soybean as an external reference standard.

  12. Comparison of multidimensional flow cytometric data by a novel data mining technique

    Science.gov (United States)

    Leary, James F.; Smith, Jacob; Szaniszlo, Peter; Reece, Lisa M.

    2007-02-01

    Most flow/image cytometric data analysis methods look for clusters in the data corresponding to specific cell subpopulations. Comparisons between different cytometry datafiles often use human pattern recognition visualization of all the different combinations of variables ("parameters") two at a time in so-called bivariate scattergrams. Not only is this tedious, but it can miss potential clusters due to projection of higher dimensional dataspaces down onto two dimensional planes making them indiscernible as separate clusters. Novel data mining algorithms, implemented in software allow for the comparison of two or more higher dimensional datafiles without the requirement for reduction of dimensionality for human visualization. Of equal importance is the comparison of higher dimensional clusters which may move around slightly in space, yet still be "similar" according to algorithms which provide measures of similarity. This software, written in C/C++ and currently implemented in software with a Windows graphical user interface, allows for direct reading of FCS2.0 format flow cytometry datafiles of any number of parameters. In a few minutes or less, complex multiparameter data of two or more files can be compared on a personal computer or workstation. The software operates in either supervised or unsupervised mode, depending on whether the user wishes to include prior user knowledge or in a data mining discovery mode. Differences between these files can be exported as sub-datafiles which can be further analyzed using any other software that can read FCS2.0 data format.

  13. A new flow cytometric method for quantitative assessment of lymphocyte mitogenic potentials.

    Science.gov (United States)

    Yamamura, Y; Rodriguez, N; Schwartz, A; Eylar, E; Bagwell, B; Yano, N

    1995-01-01

    A new flow cytometric method was developed to quantitatively assess lymphocyte proliferation simultaneously for different subsets. The cells were stained with a fluorescent dye, PKH-26 and were stimulated with mitogens. The fluorescence intensities (FL2) of proliferating cells were measured by flow cytometry; and each subset was identified by the use of a monoclonal antibody (Mab)-fluorescein-isothiocyanate (FITC) (FL1). FL2 histograms were then analyzed by the cell proliferation model based on the ModFit software (Verity). This new method revealed information which could not be obtained by conventional mitogen assays. For example, the CD4+ and the CD4- T-subsets responded to phytohemagglutinin (PHA) quite differently from each other and it was indicated that activation of one population could significantly alter the response of the other. In addition, even within a subset, all activated cells did not proliferate uniformly. Some cells divided only once while others underwent further cellular division during the same time period. The method is, therefore, invaluable for studying the nature and the extent of interactions between different cellular subsets within a culture.

  14. Flow cytometric evaluation of the intracellular bacterium, Wolbachia pipientis, in mosquito cells

    Science.gov (United States)

    Fallon, Ann M

    2014-01-01

    Wolbachia is an obligate intracellular bacterium (Anaplasmataceae, Rickettisales) that occurs in arthropods and filarial worms, and spreads by vertical transmission in the oocyte cytoplasm. In insects, reproductive distortions associated with Wolbachia, such as cytoplasmic incompatibility in mosquitoes, have potential value for controlling pests, including species that transmit human, animal and plant diseases. Wolbachia strains that propagate as a persistent infection in insect cell lines provide an important resource for developing the genetic tools that will facilitate these applications. Here I describe conditions for flow cytometric evaluation of Wolbachia growth in persistently infected mosquito cells. Cytometry parameters were established using uninfected mosquito cells and Escherichia coli as a surrogate for Wolbachia, and quantitation was correlated with cell counts determined with a Coulter electronic cell counter and bacterial counts based on optical density. The protocol was validated by showing depletion of Wolbachia in medium containing tetracycline and rifampicin, and sensitivity of Wolbachia to treatment of host cells with paraquat, an oxidizing agent, and lumiflavin, an inhibitor of riboflavin uptake. The Wolbachia peak on the flow cytometry histogram was shown to contain Wolbachia by DNA analysis using the polymerase chain reaction, and by infection of naive recipient cells. This approach will streamline investigation of Wolbachia growth in insect cell lines and facilitate identification of culture conditions that select for Wolbachia-infected cells. PMID:25300665

  15. Flow cytometric analysis using SYBR Green I for genome size estimation in coffee.

    Science.gov (United States)

    Ronildo Clarindo, Wellington; Roberto Carvalho, Carlos

    2011-02-01

    Plant genome size has been measured by flow cytometry using propidium iodide as a dye for nuclear DNA staining. However, some authors have reported the occurrence of genome size estimation errors, especially in plants rich in secondary metabolites, such as the coffee tree. In this context, we tested an alternative cytometric protocol using the SYBR Green I as a fluorochrome for stoichiometrically staining nuclear double-stranded DNA in Coffea canephora (2x) and Coffea arabica (4x). The results showed that the respective mean genome size measured from nuclei stained with SYBR Green I and propidium iodide was statistically identical. However, the G(0)/G(1) peaks of nuclei stained with SYBR Green I exhibited lower coefficient variations (1.57-2.85%) compared to those stained with propidium iodide (2.75-4.80%). Coefficient variation statistical data suggest that SYBR Green I is adequate for stoichiometric nuclei staining using this methodology. Our results provide evidence that SYBR Green I can be used in flow cytometry measurements of plants, with the advantages of minimizing errors in nuclear DNA content quantification, staining relatively quicker, with high affinity, and being less mutagenic than propidium iodide.

  16. Flow cytometric sorting coupled with exon capture sequencing identifies somatic mutations in archival lymphoma tissues.

    Science.gov (United States)

    Jiang, Nenggang; Chen, Christopher; Gong, Qiang; Shields, Kristen; Li, Yuping; Chen, YuanYuan; Song, Joo; McKeithan, Timothy W; Chan, Wing C

    2017-08-07

    The enormous number of archived formalin-fixed paraffin-embedded (FFPE) tissues available are a valuable resource of material for research. However, the use of such tissues poses many challenges, among which is the difficulty of isolating different cell populations within the tissue. In this study, we used tissue from two types of non-Hodgkin lymphoma as a model to demonstrate a method we have established and optimized to separate FFPE samples into distinct tumor and nonmalignant populations. Using FFPE reactive tonsil sections, various approaches for antigen retrieval and labeling, and the effectiveness of flow cytometric sorting were tested. We found that, among the 11 cell surface or intracellular antigen markers investigated, CD3ɛ, CD79A, LAT, PD-1, and PAX5 could be successfully labeled after antigen retrieval in Tris-EDTA buffer (pH 8.0) at 65 °C for 60 min, and 1.8-2.7 μg DNA per million cells could be extracted after sorting with DNA quality similar to that of tissue without staining or sorting. To test whether we could perform next-generation sequencing using a custom capture platform on sorted cells, we used three lymphoma cases with FFPE tissues which had been stored for 1 to 4 years. We demonstrated that the DNA from sorted cells was adequate for exon capture sequencing. By comparing the sequencing results between neoplastic and normal populations, somatic mutations could be clearly identified in the tumor population with variant frequencies as low as 11.7%.The corresponding normal fraction clearly helps in the analysis of somatic mutations and the exclusion of artifacts. This study provides an approach using flow cytometric sorting to separate different cellular populations in paraffin-embedded tissues and to unambiguously distinguish somatic mutations from germline variants or artifacts. This approach is also useful in enriching the tumor component in samples with heterogeneous components and low tumor content.Laboratory Investigation advance

  17. FlowFP: A Bioconductor Package for Fingerprinting Flow Cytometric Data

    OpenAIRE

    Wade T. Rogers; Herbert A. Holyst

    2009-01-01

    A new software package called flowFP for the analysis of flow cytometry data is introduced. The package, which is tightly integrated with other Bioconductor software for analysis of flow cytometry, provides tools to transform raw flow cytometry data into a form suitable for direct input into conventional statistical analysis and empirical modeling software tools. The approach of flowFP is to generate a description of the multivariate probability distribution function of flow cytometry data i...

  18. Flow cytometric assessment of specific leucine incorporation in the open Mediterranean

    Science.gov (United States)

    Talarmin, A.; van Wambeke, F.; Catala, P.; Courties, C.; Lebaron, P.

    2011-02-01

    The surface of the Mediterranean Sea is a low-phosphate-low-chlorophyll marine area where marine heterotrophic prokaryotes significantly contribute to the biogeochemical cycles of all biogenic elements such as carbon, notably through the mineralization of dissolved organic compounds. Cell-specific leucine incorporation rates were determined in early summer in the open stratified Mediterranean Sea. The bulk leucine incorporation rate was on average 5 ± 4 pmol leu l-1 h-1 (n=30). Cell-specific 3H-leucine incorporation rates were assayed using flow cytometry coupled to cell sorting. Heterotrophic prokaryotes (Hprok) were divided into cytometric groups according to their side scatter and green fluorescence properties: high nucleic acid containing cells (HNA) with high scatter (HNA-hs) and low scatter (HNA-ls) and low nucleic acid containing cells (LNA). Cell-specific leucine incorporation rates of these cytometric groups ranged from 2 to 54, 0.9 to 11, and 1 to 12 × 10-21 mol cell-1 h-1, respectively. LNA cells represented 45 to 63% of the Hprok abundance, and significantly contributed to the bulk leucine incorporation rates, from 12 to 43%. HNA/LNA ratios of cell-specific leucine incorporation were on average 2.0 ± 0.7 (n=30). In surface layers (from 0 m down to the deep chlorophyll depth, DCM), cell-specific rates of HNA-hs were elevated (7 and 13 times greater than LNA and HNA-ls, respectively). Nevertheless, on average HNA-hs (26%) and LNA (27%) equally contributed to the bulk leucine incorporation in these layers. Prochlorococcus cells were easily sorted near the DCM and displayed cell-specific leucine incorporation rates ranging from 3 to 55 × 10-21 mol leu cell-1 h-1, i.e. as high as HNA-hs'. These sorted groups could therefore be defined as key-players in the process of leucine incorporation into proteins. The mixotrophic features of certain photosynthetic prokaryotes and the high contribution of LNA cells to leucine incorporation within the microbial

  19. Flow cytometric assessment of specific leucine incorporation in the open Mediterranean

    Directory of Open Access Journals (Sweden)

    A. Talarmin

    2011-02-01

    Full Text Available The surface of the Mediterranean Sea is a low-phosphate-low-chlorophyll marine area where marine heterotrophic prokaryotes significantly contribute to the biogeochemical cycles of all biogenic elements such as carbon, notably through the mineralization of dissolved organic compounds. Cell-specific leucine incorporation rates were determined in early summer in the open stratified Mediterranean Sea. The bulk leucine incorporation rate was on average 5 ± 4 pmol leu l−1 h−1 (n=30. Cell-specific 3H-leucine incorporation rates were assayed using flow cytometry coupled to cell sorting. Heterotrophic prokaryotes (Hprok were divided into cytometric groups according to their side scatter and green fluorescence properties: high nucleic acid containing cells (HNA with high scatter (HNA-hs and low scatter (HNA-ls and low nucleic acid containing cells (LNA. Cell-specific leucine incorporation rates of these cytometric groups ranged from 2 to 54, 0.9 to 11, and 1 to 12 × 10-21 mol cell−1 h−1, respectively. LNA cells represented 45 to 63% of the Hprok abundance, and significantly contributed to the bulk leucine incorporation rates, from 12 to 43%. HNA/LNA ratios of cell-specific leucine incorporation were on average 2.0 ± 0.7 (n=30. In surface layers (from 0 m down to the deep chlorophyll depth, DCM, cell-specific rates of HNA-hs were elevated (7 and 13 times greater than LNA and HNA-ls, respectively. Nevertheless, on average HNA-hs (26% and LNA (27% equally contributed to the bulk leucine incorporation in these layers. Prochlorococcus cells were easily sorted near the DCM and displayed cell-specific leucine incorporation rates ranging from 3 to 55 × 10-21 mol leu cell−1 h−1, i.e. as high as HNA-hs'. These sorted groups could therefore be defined as key-players in the process of leucine incorporation into proteins. The

  20. [Flow cytometric analysis of ICRF-193 influence on cell passage through mitosis].

    Science.gov (United States)

    Shatrova, A N; Aksenov, N D; Zenin, V V

    2002-01-01

    Studying the effect of topoisomerase II (topo II) inhibitors on cell passage through mitosis seems to be important for understanding the role of this enzyme during chromosome condensation and segregation. A flow cytometric assay (Zenin et al., 2001) allowed to determine the mitotic index, and to discriminate between not only cells in G2 and M phases (including metaphase and anaphase cells), but also cells in pseudo-G1 with 4c DNA content. It is shown that topo II catalytic inhibitor ICRF-193 blocks G2-M transition in a lymphoblastoid cell line GM-130. Addition of caffeine to cells abrogated a block of their entering mitosis but not the inhibitor action. Cells entered mitosis, which was proven by the presence of chromosomes in the examined specimen, and, bypassing anaphase, appeared in pseudo-G1 with 4c DNA content. We have found that in the presence of ICRF-193 cells, GM-130 and Hep-2 lines, previously blocked by nocodazole when in mitosis and then washed, pass through metaphase, enter anaphase and leave it to pass to pseudo-G1 with the 4c DNA content. Thus, by inhibiting topo II activity ICRF-193 causes abnormal mitotic transition.

  1. Establishment of multiplexed, microsphere-based flow cytometric assay for multiple human tumor markers

    Institute of Scientific and Technical Information of China (English)

    Kai SUN; Qian WANG; Xiao-hui HUANG; Mao-chuan ZHEN; Wen LI; Long-juan ZHANG

    2007-01-01

    Aim: The multiplexed, microsphere-based flow cytometric assay (MFCA) for mul- tiple human tumor markers was established for the early screening and detection of suspected cancer patients. Methods: Covalent coupling of capture antibodies directed against their respective tumor markers to fluorescent microspheres was performed by following the protocols recommended by a commercial corporation with some modifications. The coupling efficiency and cross-reactivity were iden- tified by the Luminex 100 system and associated software. The standard curve was constructed by using serial dilution of recombinant tumor marker standards and was validated by comparison with ELISA for quantifying the tumor markers in serum samples. Results: The identifications revealed that the coupling proce- dures were successful without non-specific cross-reactivity and the standard curve was highly efficient. However, it was necessary to ensure the quality con- trol of the coupling process since slight variations in the coupling procedures could profoundly affect the density of capture reagents coupled to the microspheres and consequently adversely affect the assay precision. In addition to its multi-analyte capability, the MFCA system had definite advantages, such as higher reproducibility, greater dynamic range of measurement, and considerably less preparation time and labor over the conventional "gold standard", which was the ELISA. Conclusion: The successful establishment of the MFCA system for the simultaneous detection of multiple tumor markers will provide the foundation for the further study of clinical applications.

  2. Flow cytometric quantitation of phagocytosis in heparinized complete blood with latex particles and Candida albicans

    Directory of Open Access Journals (Sweden)

    Jesús M. Egido

    1997-12-01

    Full Text Available We report a rapid method for the flow cytometric quantitation of phagocytosis in heparinized complete peripherial blood (HCPB, using commercially available phycoerythrin-conjugated latex particles of 1µm diameter. The method is faster and shows greater reproducibility than Bjerknes' (1984 standard technique using propidium iodide-stained Candida albicans, conventionally applied to the leukocytic layer of peripherial blood but here modified for HCPB. We also report a modification of Bjerknes' Intracellular Killing Test to allow its application to HCPB.Se da cuenta de un método rápido para la cuantización del flujo citométrico de la fagocitosis en sangre periférica completamente heparinizada (HCPB, mediante la utilización de partículas de látex phycoerythrin-conjugadas de 1µm de diámetro disponibles comercialmente. El método es más rápido y presenta mayor reproducibilidad que la técnica estandar de Bjerknes' (1984 utilizando propidium iodide-teñida Candida albicans, aplicada convencionalmente a la capa leucocitica de sangre periférica pero modificada por HCPB. Tambien damos cuenta de una modificación de Bjerknes' Intracellular Killing Test para permitir su aplicación a HCPB.

  3. Flow cytometric analysis of lymphocyte proliferative responses to food allergens in dogs with food allergy.

    Science.gov (United States)

    Fujimura, Masato; Masuda, Kenichi; Hayashiya, Makio; Okayama, Taro

    2011-10-01

    Two different allergy tests, antigen-specific immunoglobulin E quantification (IgE test) and flow cytometric analysis of antigen-specific proliferation of peripheral lymphocytes (lymphocyte proliferation test), were performed to examine differences in allergic reactions to food allergens in dogs with food allergy (FA). Thirteen dogs were diagnosed as FA based on clinical findings and elimination diet trials. Seven dogs clinically diagnosed with canine atopic dermatitis (CAD) were used as a disease control group, and 5 healthy dogs were used as a negative control group. In the FA group, 19 and 33 allergen reactions were identified using the serum IgE test and the lymphocyte proliferation test, respectively. Likewise, in the CAD group, 12 and 6 allergen reactions and in the healthy dogs 3 and 0 allergen reactions were identified by each test, respectively. A significant difference was found between FA and healthy dogs in terms of positive allergen detection by the lymphocyte proliferation test, suggesting that the test can be useful to differentiate FA from healthy dogs but not from CAD. Both tests were repeated in 6 of the dogs with FA after a 1.5- to 5-month elimination diet trial. The IgE concentrations in 9 of 11 of the positive reactions decreased by 20-80%, whereas all the positive reactions in the lymphocyte proliferation test decreased to nearly zero (Pfood allergens may be involved in the pathogenesis of canine FA.

  4. A Flow Cytometric Clonogenic Assay Reveals the Single-Cell Potency of Doxorubicin

    Science.gov (United States)

    Maass, Katie F.; Kulkarni, Chethana; Quadir, Mohiuddin A.; Hammond, Paula T.; Betts, Alison M.; Wittrup, K. Dane

    2015-01-01

    Standard cell proliferation assays use bulk media drug concentration to ascertain the potency of chemotherapeutic drugs; however, the relevant quantity is clearly the amount of drug actually taken up by the cell. To address this discrepancy, we have developed a flow cytometric clonogenic assay to correlate the amount of drug in a single cell with the cell’s ability to proliferate using a cell tracing dye and doxorubicin, a naturally fluorescent chemotherapeutic drug. By varying doxorubicin concentration in the media, length of treatment time, and treatment with verapamil, an efflux pump inhibitor, we introduced 105 – 1010 doxorubicin molecules per cell; then used a dye-dilution assay to simultaneously assess the number of cell divisions. We find that a cell’s ability to proliferate is a surprisingly conserved function of the number of intracellular doxorubicin molecules, resulting in single-cell IC50 values of 4 – 12 million intracellular doxorubicin molecules. The developed assay is a straightforward method for understanding a drug’s single-cell potency and can be used for any fluorescent or fluorescently-labeled drug, including nanoparticles or antibody-drug conjugates. PMID:26344409

  5. Flow cytometric probing of mitochondrial function in equine peripheral blood mononuclear cells

    Directory of Open Access Journals (Sweden)

    Coignoul Freddy

    2007-09-01

    Full Text Available Abstract Background The morphopathological picture of a subset of equine myopathies is compatible with a primary mitochondrial disease, but functional confirmation in vivo is still pending. The cationic dye JC-1 exhibits potential-dependent accumulation in mitochondria that is detectable by a fluorescence shift from green to orange. As a consequence, mitochondrial membrane potential can be optically measured by the orange/green fluorescence intensity ratio. A flow cytometric standardized analytic procedure of the mitochondrial function of equine peripheral blood mononuclear cells is proposed along with a critical appraisal of the crucial questions of technical aspects, reproducibility, effect of time elapsed between blood sampling and laboratory processing and reference values. Results The JC-1-associated fluorescence orange and green values and their ratio were proved to be stable over time, independent of age and sex and hypersensitive to intoxication with a mitochondrial potential dissipator. Unless time elapsed between blood sampling and laboratory processing does not exceed 5 hours, the values retrieved remain stable. Reference values for clinically normal horses are given. Conclusion Whenever a quantitative measurement of mitochondrial function in a horse is desired, blood samples should be taken in sodium citrate tubes and kept at room temperature for a maximum of 5 hours before the laboratory procedure detailed here is started. The hope is that this new test may help in confirming, studying and preventing equine myopathies that are currently imputed to mitochondrial dysfunction.

  6. Antiphospholipid antibody syndrome: the flow cytometric annexin A5 competition assay as a diagnostic tool.

    Science.gov (United States)

    Tomer, A; Bar-Lev, S; Fleisher, S; Shenkman, B; Friger, M; Abu-Shakra, M

    2007-10-01

    The mechanism underlying hypercoagulability in antiphospholipid antibody syndrome (APS) is uncertain. Here, we present a flow-cytometric assay (FCA) based on the hypothesis that anti-platelet-anionic-phospholipid autoantibodies (aPL) interfere with the activity of the natural anticoagulant protein annexin A5, thereby accelerating platelet procoagulant activity. This study assessed the clinical utility of the feasible FCA, which demonstrates the competition of the patient's aPL with the binding of annexin A5 to the platelet-anionic-phospholipids, in the diagnosis of APS. Sixty-two (94%) of 66 APS patients, 20 (51%) of 39 patients with systemic lupus erythematosus and two (4%) of 49 healthy individuals were positive by FCA. Compared with the anticardiolipin (aCL) assay, the relative sensitivity was 82% and the specificity 73.3%. However, 19 (25%) aCL-negative patients were positive by FCA; 12 were positive for lupus-anticoagulant (LA). Compared with LA assay, the relative sensitivity was 85% and the specificity 72.2%. However, 21 (26%) LA-negative patients were FCA-positive, 12 were positive for aCL. The FCA was particularly sensitive for APS patients with arterial (97.0%) and gestational vascular complications (100%) with overall sensitivity of 95% and specificity of 97%. Our findings suggest that the FCA is practical, sensitive and specific for the detection of clinically relevant aPL in the diagnosis of APS.

  7. Dissociation of skeletal muscle for flow cytometric characterization of immune cells in macaques.

    Science.gov (United States)

    Liang, Frank; Ploquin, Aurélie; Hernández, José DelaO; Fausther-Bovendo, Hugues; Lindgren, Gustaf; Stanley, Daphne; Martinez, Aiala Salvador; Brenchley, Jason M; Koup, Richard A; Loré, Karin; Sullivan, Nancy J

    2015-10-01

    The majority of vaccines and several treatments are administered by intramuscular injection. The aim is to engage and activate immune cells, although they are rare in normal skeletal muscle. The phenotype and function of resident as well as infiltrating immune cells in the muscle after injection are largely unknown. While methods for obtaining and characterizing murine muscle cell suspensions have been reported, protocols for nonhuman primates (NHPs) have not been well defined. NHPs comprise important in vivo models for studies of immune cell function due to their high degree of resemblance with humans. In this study, we developed and systematically compared methods to collect vaccine-injected muscle tissue to be processed into single cell suspensions for flow cytometric characterization of immune cells. We found that muscle tissue processed by mechanical disruption alone resulted in significantly lower immune cell yields compared to enzymatic digestion using Liberase. Dendritic cell subsets, monocytes, macrophages, neutrophils, B cells, T cells and NK cells were readily detected in the muscle by the classic human markers. The methods for obtaining skeletal muscle cell suspension established here offer opportunities to increase the understanding of immune responses in the muscle, and provide a basis for defining immediate post-injection vaccine responses in primates.

  8. Successful low dose insemination of flow cytometrically sorted ram spermatozoa in sheep.

    Science.gov (United States)

    de Graaf, S P; Evans, G; Maxwell, W M C; Downing, J A; O'Brien, J K

    2007-12-01

    The fertility of ram spermatozoa that had undergone flow cytometric sorting (MoFlo SX) and cryopreservation was assessed after low-dose insemination of synchronized Merino ewes. Oestrus was synchronized with progestagen-impregnated pessaries, PMSG and GnRH treatment. Ewes (n = 360) were inseminated with 1 x 10(6), 5 x 10(6) or 15 x 10(6) motile sorted frozen-thawed (S(1), S(5), or S(15) respectively) or non-sorted frozen-thawed (C(1), C(5) or C(15) respectively) spermatozoa from three rams. An additional group of ewes were inseminated with 50 x 10(6) motile non-sorted frozen-thawed spermatozoa (C(50)) to provide a commercial dose control. The percentage of ewes lambing after insemination was similar for C(50) (24/38, 63.2%), C(15) (37/54, 68.5%), S(15) (38/57, 66.7%), S(5) (37/56, 66.1%) and S(1) (32/52, 61.5%) groups (p > 0.05), but lower for C(5) (19/48, 39.6%) and C(1) (19/55, 34.5%) treatments (p sheep as a reduction in the minimum effective sperm number will allow a corresponding decrease in the associated cost per dose.

  9. Flow cytometric viability assessment of lactic acid bacteria starter cultures produced by fluidized bed drying.

    Science.gov (United States)

    Bensch, Gerald; Rüger, Marc; Wassermann, Magdalena; Weinholz, Susann; Reichl, Udo; Cordes, Christiana

    2014-06-01

    For starter culture production, fluidized bed drying is an efficient and cost-effective alternative to the most frequently used freeze drying method. However, fluidized bed drying also poses damaging or lethal stress to bacteria. Therefore, investigation of impact of process variables and conditions on viability of starter cultures produced by fluidized bed drying is of major interest. Viability of bacteria is most frequently assessed by plate counting. While reproductive growth of cells can be characterized by the number of colony-forming units, it cannot provide the number of viable-but-nonculturable cells. However, in starter cultures, these cells still contribute to the fermentation during food production. In this study, flow cytometry was applied to assess viability of Lactobacillus plantarum starter cultures by membrane integrity analysis using SYBR®Green I and propidium iodide staining. The enumeration method established allowed for rapid, precise and sensitive determination of viable cell concentration, and was used to investigate effects of fluidized bed drying and storage on viability of L. plantarum. Drying caused substantial membrane damage on cells, most likely due to dehydration and oxidative stress. Nevertheless, high bacterial survival rates were obtained, and granulates contained in the average 2.7 × 10(9) viable cells/g. Furthermore, increased temperatures reduced viability of bacteria during storage. Differences in results of flow cytometry and plate counting suggested an occurrence of viable-but-nonculturable cells during storage. Overall, flow cytometric viability assessment is highly feasible for rapid routine in-process control in production of L. plantarum starter cultures, produced by fluidized bed drying.

  10. An imaging flow cytometric method for measuring cell division history and molecular symmetry during mitosis.

    Science.gov (United States)

    Filby, Andrew; Perucha, Esperanza; Summers, Huw; Rees, Paul; Chana, Prabhjoat; Heck, Susanne; Lord, Graham M; Davies, Derek

    2011-07-01

    Asymmetric cell division is an important mechanism for generating cellular diversity, however, techniques for measuring the distribution of fate-regulating molecules during mitosis have been hampered by a lack of objectivity, quantitation, and statistical robustness. Here we describe a novel imaging flow cytometric approach that is able to report a cells proliferative history and cell cycle position using dye dilution, pH3, and PI staining to then measure the spatial distribution of fluorescent signals during mitosis using CCD-derived imagery. Using Jurkat cells, resolution of the fluorescently labeled populations was comparable to traditional PMT based cytometers thus eliminating the need to sort cells with specific division histories for microscopy. Subdividing mitotic stages by morphology allowed us to determine the time spent in each cell cycle phase using mathematical modeling approaches. Furthermore high sample throughput allowed us to collect statistically relevant numbers of cells without the need to use blocking agents that artificially enrich for mitotic events. The fluorescent imagery was used to measure PKCζ protein and EEA-1+ endosome distribution during different mitotic phases in Jurkat cells. While telophase cells represented the favorable population for measuring asymmetry, asynchronously dividing cells spent approximately 43 seconds in this stage, explaining why they were present at such low frequencies. This necessitated the acquisition of large cell numbers. Interestingly we found that PKCζ was inherited asymmetrically in 2.5% of all telophasic events whereas endosome inheritance was significantly more symmetrical. Furthermore, molecular polarity at early mitotic phases was a poor indicator of asymmetry during telophase highlighting that, though rare, telophasic events represented the best candidates for asymmetry studies. In summary, this technique combines the spatial information afforded by fluorescence microscopy with the statistical

  11. Flow cytometric analysis of lectin binding to in vitro-cultured Perkinsus marinus surface carbohydrates

    Science.gov (United States)

    Gauthier, J.D.; Jenkins, J.A.; La Peyre, Jerome F.

    2004-01-01

    Parasite surface glycoconjugates are frequently involved in cellular recognition and colonization of the host. This study reports on the identification of Perkinsus marinus surface carbohydrates by flow cytometric analyses of fluorescein isothiocyanate-conjugated lectin binding. Lectin-binding specificity was confirmed by sugar inhibition and Kolmogorov-Smirnov statistics. Clear, measurable fluorescence peaks were discriminated, and no parasite autofluorescence was observed. Parasites (GTLA-5 and Perkinsus-1 strains) harvested during log and stationary phases of growth in a protein-free medium reacted strongly with concanavalin A and wheat germ agglutinin, which bind to glucose-mannose and N-acetyl-D-glucosamine (GlcNAc) moieties, respectively. Both P. marinus strains bound with lower intensity to Maclura pomifera agglutinin, Bauhinia purpurea agglutinin, soybean agglutinin (N-acetyl-D-galactosamine-specific lectins), peanut agglutinin (PNA) (terminal galactose specific), and Griffonia simplicifolia II (GlcNAc specific). Only background fluorescence levels were detected with Ulex europaeus agglutinin I (L-fucose specific) and Limulus polyphemus agglutinin (sialic acid specific). The lectin-binding profiles were similar for the 2 strains except for a greater relative binding intensity of PNA for Perkinsus-1 and an overall greater lectin-binding capacity of Perkinsus-1 compared with GTLA-5. Growth stage comparisons revealed increased lectin-binding intensities during stationary phase compared with log phase of growth. This is the first report of the identification of surface glycoconjugates on a Perkinsus spp. by flow cytometry and the first to demonstrate that differential surface sugar expression is growth phase and strain dependent. ?? American Society of Parasitologists 2004.

  12. Flow cytometric method for measuring chromatin fragmentation in fixed sperm from yellow perch (Perca flavescens).

    Science.gov (United States)

    Jenkins, J A; Draugelis-Dale, R O; Pinkney, A E; Iwanowicz, L R; Blazer, V S

    2015-03-15

    Declining harvests of yellow perch, Perca flavescens, in urbanized watersheds of Chesapeake Bay have prompted investigations of their reproductive fitness. The purpose of this study was to establish a flow cytometric technique for DNA analysis of fixed samples sent from the field to provide reliable gamete quality measurements. Similar to the sperm chromatin structure assay, measures were made on the susceptibility of nuclear DNA to acid-induced denaturation, but used fixed rather than live or thawed cells. Nuclei were best exposed to the acid treatment for 1 minute at 37 °C followed by the addition of cold (4 °C) propidium iodide staining solution before flow cytometry. The rationale for protocol development is presented graphically through cytograms. Field results collected in 2008 and 2009 revealed DNA fragmentation up to 14.5%. In 2008, DNA fragmentation from the more urbanized watersheds was significantly greater than from reference sites (P = 0.026) and in 2009, higher percentages of haploid testicular cells were noted from the less urbanized watersheds (P = 0.032) indicating better reproductive condition at sites with less urbanization. For both years, total and progressive live sperm motilities by computer-assisted sperm motion analysis ranged from 19.1% to 76.5%, being significantly higher at the less urbanized sites (P fragmented DNA to be denatured under standard conditions, or 1 minute at 37 °C with 10% buffered formalin-fixed cells. The study of fixed sperm makes possible the restrospective investigation of germplasm fragmentation, spermatogenic ploidy patterns, and chromatin compaction levels from samples translocated over distance and time. The protocol provides an approach that can be modified for other species across taxa.

  13. Flow cytometric method for measuring chromatin fragmentation in fixed sperm from yellow perch (Perca flavescens)

    Science.gov (United States)

    Jenkins, Jill A.; Draugelis-Dale, Rassa O.; Pinkney, Alfred E.; Iwanowicz, Luke R.; Blazer, Vicki

    2015-01-01

    Declining harvests of yellow perch, Perca flavescens, in urbanized watersheds of Chesapeake Bay have prompted investigations of their reproductive fitness. The purpose of this study was to establish a flow cytometric technique for DNA analysis of fixed samples sent from the field to provide reliable gamete quality measurements. Similar to the sperm chromatin structure assay, measures were made on the susceptibility of nuclear DNA to acid-induced denaturation, but used fixed rather than live or thawed cells. Nuclei were best exposed to the acid treatment for 1 minute at 37 °C followed by the addition of cold (4 °C) propidium iodide staining solution before flow cytometry. The rationale for protocol development is presented graphically through cytograms. Field results collected in 2008 and 2009 revealed DNA fragmentation up to 14.5%. In 2008, DNA fragmentation from the more urbanized watersheds was significantly greater than from reference sites (P = 0.026) and in 2009, higher percentages of haploid testicular cells were noted from the less urbanized watersheds (P = 0.032) indicating better reproductive condition at sites with less urbanization. For both years, total and progressive live sperm motilities by computer-assisted sperm motion analysis ranged from 19.1% to 76.5%, being significantly higher at the less urbanized sites (P fragmented DNA to be denatured under standard conditions, or 1 minute at 37 °C with 10% buffered formalin–fixed cells. The study of fixed sperm makes possible the restrospective investigation of germplasm fragmentation, spermatogenic ploidy patterns, and chromatin compaction levels from samples translocated over distance and time. The protocol provides an approach that can be modified for other species across taxa.

  14. Evaluation of the S phase distribution of flow cytometric DNA histograms by autoradiography and computer algorithms.

    Science.gov (United States)

    Sheck, L E; Muirhead, K A; Horan, P K

    1980-09-01

    Cell sorting and tritiated thymidine autoradiography were used to define the distribution of S phase cells in flow cytometric DNA histograms obtained from exponential mouse lymphoma cells (L5178Y). The numbers of labeled S phase cells, autoradiographically determined from cells sorted at 2-channel intervals in the G1/early S and late S/G2M regions of the histogram, were compared with the numbers of computed S phase cells in comparable 2-channel intervals as predicted by several computer algorithms used to extract cell cycle phase distributions from DNA histograms. Polynomial and multirectangle algorithms gave computed estimates of total %S in close agreement with the tritiated thymidine labeling index for the cell population, while multi-Gaussian algorithms underestimated %S. Interval autoradiographic and algorithm studies confirmed these results in that no significant differences were found between the autoradiographic S phase distribution and S phase distributions calculated by the polynomial and multirectangle models. However, S phase cells were significantly underestimated in G1/early S by a constrained multi-Gaussian model and in both G1/early S and late S/G2 by an unconstrained multi-Gaussian model. For the particular cell line (L5178Y), staining protocol (mithramycin following ethanol fixation) and instrumentation (Coulter TPS-2 cell sorter) used in this study, close agreement between computed %S and tritiated thymidine labeling index was found to be a reliable indicator of an algorithm's success in resolving S phase cells in the G1/S and S/G2 transition regions of the DNA histograms.

  15. Flow Cytometric Assessment of Bacterial Abundance in Soils, Sediments and Sludge.

    Science.gov (United States)

    Frossard, Aline; Hammes, Frederik; Gessner, Mark O

    2016-01-01

    Bacterial abundance is a fundamental measure in microbiology, but its assessment is often tedious, especially for soil, and sediment samples. To overcome this limitation, we adopted a time-efficient flow-cytometric (FCM) counting method involving cell detachment and separation from matrix particles by centrifugation in tubes receiving sample suspensions and Histodenz(®) solution. We used this approach to assess bacterial abundances in diverse soils (natural and agricultural), sediments (streams and lakes) and sludge from sand-filters in a drinking water treatment plant and compared the results to bacterial abundances determined by two established methods, epifluorescence microscopy (EM) and adenosine triphosphate (ATP) quantification. Cell abundances determined by FCM and EM correlated fairly well, although absolute cell abundances were generally lower when determined by FCM. FCM also showed significant relations with cell counts converted from ATP concentrations, although estimates derived from ATP determinations were typically higher, indicating the presence of ATP sources other than bacteria. Soil and sediment organic matter (OM) content influenced the goodness of fit between counts obtained with EM and FCM. In particular, bacterial abundance determined by FCM in samples containing less than 10% OM, such as stream sediment, was particularly well correlated with the cell counts assessed by EM. Overall, these results suggest that FCM following cell detachment and purification is a useful approach to increase sample throughput for determining bacterial abundances in soils, sediments and sludge. However, notable scatter and only partial concordance among the FCM and reference methods suggests that protocols require further improvement for assessments requiring high precision, especially when OM contents in samples are high.

  16. Novel nuclei isolation buffer for flow cytometric genome size estimation of Zingiberaceae: a comparison with common isolation buffers.

    Science.gov (United States)

    Sadhu, Abhishek; Bhadra, Sreetama; Bandyopadhyay, Maumita

    2016-11-01

    Cytological parameters such as chromosome numbers and genome sizes of plants are used routinely for studying evolutionary aspects of polyploid plants. Members of Zingiberaceae show a wide range of inter- and intrageneric variation in their reproductive habits and ploidy levels. Conventional cytological study in this group of plants is severely hampered by the presence of diverse secondary metabolites, which also affect their genome size estimation using flow cytometry. None of the several nuclei isolation buffers used in flow cytometry could be used very successfully for members of Zingiberaceae to isolate good quality nuclei from both shoot and root tissues. The competency of eight nuclei isolation buffers was compared with a newly formulated buffer, MB01, in six different genera of Zingiberaceae based on the fluorescence intensity of propidium iodide-stained nuclei using flow cytometric parameters, namely coefficient of variation of the G0/G1 peak, debris factor and nuclei yield factor. Isolated nuclei were studied using fluorescence microscopy and bio-scanning electron microscopy to analyse stain-nuclei interaction and nuclei topology, respectively. Genome contents of 21 species belonging to these six genera were determined using MB01. Flow cytometric parameters showed significant differences among the analysed buffers. MB01 exhibited the best combination of analysed parameters; photomicrographs obtained from fluorescence and electron microscopy supported the superiority of MB01 buffer over other buffers. Among the 21 species studied, nuclear DNA contents of 14 species are reported for the first time. Results of the present study substantiate the enhanced efficacy of MB01, compared to other buffers tested, in the generation of acceptable cytograms from all species of Zingiberaceae studied. Our study facilitates new ways of sample preparation for further flow cytometric analysis of genome size of other members belonging to this highly complex polyploid family.

  17. Effect of intra-cellular trafficking on flow cytometric measurement of neutrophil's oxidative status in iron deficient pregnant females.

    Science.gov (United States)

    Youssef, Soha R; Hendawy, Sherif F; Boshnak, Noha H; Sedhom, Mariana S

    2017-03-27

    Iron deficiency and iron deficiency anemia are prevalent among pregnant women particularly in developing countries. This study aimed to evaluate the iron status among Egyptian pregnant women and its impact on their neutrophil's count and antimicrobial functions. Ninety pregnant females underwent complete blood count, iron profile, flow cytometric studies for neutrophil myeloperoxidase expression & oxidative burst using dihydrorhodamine 123 (DHR) after phorbol-12-myristate-13-acetate (PMA) stimulation as well as neutrophil phagocytic and lytic indices. According to percent saturation 54/90 women (60%) were iron deficient (women were in their third trimester compared to controls. No significant difference was found between the iron deficient & sufficient groups as regards anemia despite a positive correlation between haemoglobin level and percent saturation (P=.02). Both the phagocytic and lytic indices were significantly lower among the cases compared to controls (P=.014 & .002 respectively). Cases and controls were comparable as regards flow cytometric studies of neutrophils' myeloperoxidase and oxidative burst (P>.05). No significant correlation was found between any of the iron profile parameters and the oxidative burst by flow cytometry. Functional microphage assay (phagocytic and lytic indices) may be more relevant and cost effective than flow cytometry assays of myeloperoxidase and oxidative burst in reflecting either iron status or cellular immunity in pregnancy. © 2017 Wiley Periodicals, Inc.

  18. Clonal evolution demonstrated by flow cytometric DNA analysis of a human colonic carcinoma grown in nude mice

    DEFF Research Database (Denmark)

    Vindeløv, L L; Spang-Thomsen, M; Visfeldt, J

    1982-01-01

    A spontaneous change in DNA content of a human colonic carcinoma grown in nude mice was observed fortuitously. The tumor initially had a G1 cell DNA content of 1.3 times that of normal cells. Flow cytometric DNA analysis showed in transplant generation 56 the appearance of a new subpopulation whi...... evolution of a tumor would be less pronounced if old subpopulations often become extinct as new ones emerge. Heterogeneity of human tumors is of clinical importance because the individual subpopulations may have different sensitivity patterns to antineoplastic drugs....

  19. Effects of Hoechst33342 staining on the viability and flow cytometric sex-sorting of frozen-thawed ram sperm.

    Science.gov (United States)

    Quan, Guo Bo; Ma, Yuan; Li, Jian; Wu, Guo Quan; Li, Dong Jiang; Ni, Yi Na; Lv, Chun Rong; Zhu, Lan; Hong, Qiong Hua

    2015-02-01

    Cytometric sorting of frozen-thawed sperm can overcome difficulties caused by the unavailability of sorting facilities on farms where semen is collected from male livestock. In order to optimize the cytometric sex-sorting procedure, effects of Hoechst33342 staining on the viability and cytometric sorting efficiency of frozen-thawed ram sperm were evaluated. The frozen-thawed sperm were stained with Hoechst33342 at various dye concentrations (80 μM, 120 μM, 160 μM, 200 μM, 240 μM, or 320 μM) for 45 min to evaluate effects of dye dose. The frozen-thawed sperm were stained with 160 μM Hoechst33342 for various durations (0 min, 15 min, 30 min, 45 min, 60 min, 75 min, or 90 min) to evaluate effects of staining duration. Sperm motility and moving velocity were analyzed using a computer-assisted sperm analysis system (CASAS). Acrosome status, membrane integrity, and distribution of phosphatidylserine (PS) in Hoechst33342-stained sperm were analyzed using flow cytometry after staining with fluorescein isothiocyanate-labeled lectin from pisum sativum (FITC-PSA), Annexin V, or propidium iodide (PI). The fertility of Hoechst33342-stained sperm was analyzed by in vitro fertilization (IVF). A high-speed cell sorter was used to evaluate effects of Hoechst33342 staining on cytometric sex-sorting of frozen-thawed sperm. The motility, moving velocity, membrane integrity, and PS distribution of Hoechst33342-stained sperm were significantly different from that of immediately thawed sperm (Pram sperm. Results of cytometric sorting indicated that frozen-thawed sperm can be efficiently sorted into two sperm populations with X and Y chromosome when the Hoechst33342 concentration was 160 μM. Moreover, when the staining duration was equal to or longer than 45 min, the frozen-thawed sperm can be successfully sorted in the presence of 160μM Hoechst33342. In conclusion, Hoechst33342 staining can detrimentally influence viability of frozen-thawed ram sperm except acrosome and in vitro

  20. Postlarval muscle growth in fish: a DNA flow cytometric and morphometric analysis.

    Science.gov (United States)

    Alfei, L; Maggi, F; Parvopassu, F; Bertoncello, G; De Vita, R

    1989-01-01

    The mechanism of postlarval fish myotomal growth was investigated in trout (Salmo gairdneri) by means of morphometric and cytofluorometric analysis. The mechanism by which new fibres are added during postlarval growth (hyperplasia) is not fully understood. In histological cross sections these new fibres have a small diameter which give the muscle a "mosaic" appearance. One hypothesis suggested that they could be derived from the proliferative activity of satellite cells. DNA cytofluorometric analysis of nuclei suspensions obtained from trout white myotomal muscle during different developmental stages (eleutherembyronic; alevin; yearling and adult) showed a consistently low S-cytometric phase during all stage in which myofibres of small diameters were present. The percentage of such small fibres, determined by morphometric analysis, suggested that satellite cells are the proliferative population. In fact, their percentages, as determined by morphometric analysis in histological section, bear a linear relationship with the S-cytometric phase percent nuclei (R = 0.927). Only in adults (67 cm in size) there was a significant decrease in the S-cytometric phase. At this stage, in histological sections, the myotomal muscle no longer had a "mosaic" appearance because of the disappearance of the small fibres. It may, therefore, be supposed that in the cm 67 adult specimens, the proliferative population is entering the G0 phase. It is known, in fact, that muscle growth proceeds only by fibre hypertrophy in trout longer than 70 cm in length (Stickland, 1983).

  1. Correlation between histological grading and ploidy status in potentially malignant disorders of the oral mucosa: A flow cytometric analysis

    Directory of Open Access Journals (Sweden)

    T Vijayavel

    2013-01-01

    Full Text Available Background: Histopathological grading of oral dysplastic lesions is the method of choice for evaluating malignant and potentially malignant disorders. Owing to inter- and intra-observer variability, determination of the DNA ploidy status of lesions may serve as an adjunct in the prediction of malignant transformation. Aim: To correlate histopathological grading and ploidy status in potentially malignant and malignant disorders of the oral mucosa. Settings and Design: A pilot study was done with 30 patients (10 patients with oral potentially malignant disorders predominantly leukoplakia, 10 patients with oral malignant lesions and 10 patients with normal mucosa. Materials and Methods: Incisional biopsy was done after isolating the biopsy site with 1% Toluidine blue staining. Two sections of the tissue were removed and sent for histopathological and Flow-cytometric analysis respectively. Histopathological diagnosis was obtained and compared with Flow-cytometric results which were graded as diploid and aneuploid. Further, the S - phase fraction, DNA index were also calculated to evaluate the severity of malignant transformation or malignancy. Statistical Analysis: The results were analyzed using Pearson Chi-Square Test. Results: There exists a significant correlation between histopathology and ploidy status in both potentially malignant and malignant group. (P = 0.002. Conclusion: The data from this study has shown that DNA Ploidy analysis can be used as a valuable tool in assessing the carcinomatous progression of potentially malignant and malignant lesions.

  2. A cluster analysis method for identification of subpopulations of cells in flow cytometric list-mode arrays

    Science.gov (United States)

    Li, Z. K.

    1985-01-01

    A specialized program was developed for flow cytometric list-mode data using an heirarchical tree method for identifying and enumerating individual subpopulations, the method of principal components for a two-dimensional display of 6-parameter data array, and a standard sorting algorithm for characterizing subpopulations. The program was tested against a published data set subjected to cluster analysis and experimental data sets from controlled flow cytometry experiments using a Coulter Electronics EPICS V Cell Sorter. A version of the program in compiled BASIC is usable on a 16-bit microcomputer with the MS-DOS operating system. It is specialized for 6 parameters and up to 20,000 cells. Its two-dimensional display of Euclidean distances reveals clusters clearly, as does its 1-dimensional display. The identified subpopulations can, in suitable experiments, be related to functional subpopulations of cells.

  3. A Protocol for the Comprehensive Flow Cytometric Analysis of Immune Cells in Normal and Inflamed Murine Non-Lymphoid Tissues.

    Science.gov (United States)

    Yu, Yen-Rei A; O'Koren, Emily G; Hotten, Danielle F; Kan, Matthew J; Kopin, David; Nelson, Erik R; Que, Loretta; Gunn, Michael D

    2016-01-01

    Flow cytometry is used extensively to examine immune cells in non-lymphoid tissues. However, a method of flow cytometric analysis that is both comprehensive and widely applicable has not been described. We developed a protocol for the flow cytometric analysis of non-lymphoid tissues, including methods of tissue preparation, a 10-fluorochrome panel for cell staining, and a standardized gating strategy, that allows the simultaneous identification and quantification of all major immune cell types in a variety of normal and inflamed non-lymphoid tissues. We demonstrate that our basic protocol minimizes cell loss, reliably distinguishes macrophages from dendritic cells (DC), and identifies all major granulocytic and mononuclear phagocytic cell types. This protocol is able to accurately quantify 11 distinct immune cell types, including T cells, B cells, NK cells, neutrophils, eosinophils, inflammatory monocytes, resident monocytes, alveolar macrophages, resident/interstitial macrophages, CD11b- DC, and CD11b+ DC, in normal lung, heart, liver, kidney, intestine, skin, eyes, and mammary gland. We also characterized the expression patterns of several commonly used myeloid and macrophage markers. This basic protocol can be expanded to identify additional cell types such as mast cells, basophils, and plasmacytoid DC, or perform detailed phenotyping of specific cell types. In examining models of primary and metastatic mammary tumors, this protocol allowed the identification of several distinct tumor associated macrophage phenotypes, the appearance of which was highly specific to individual tumor cell lines. This protocol provides a valuable tool to examine immune cell repertoires and follow immune responses in a wide variety of tissues and experimental conditions.

  4. Flow cytometric analyses of the viability, surface marker expression and function of lymphocytes from children following cryopreservation.

    Science.gov (United States)

    Chen, Xi; Zhang, Hui; Mou, Wenjun; Qi, Zhan; Ren, Xiaoya; Wang, Guoliang; Jiao, Hong; Kong, Xiaohui; Gui, Jingang

    2016-11-01

    Flow cytometric analysis is important for the investigation and clinical preparation of lymphocytes from children. However, the strict requirement of cell freshness and inter‑assay variability limits the application of this methodology for pediatric investigations. Therefore, it is necessary to identify a reliable cryopreservative method capable of maintaining high cell viability and proper cell function in lymphocytes from children. In the present study, eight commonly‑used cell cyropreservative methods were used, and their effects on cell viability, surface marker expression and cell function were examined. In addition, how these methods affect the distribution of T‑cell receptor Vβ subfamilies were also determined. The results of the present study provided valuable experimental evidence, based on which the optimal method for the cryopreservation of lymphocytes from children in pediatric investigations and clinical applications can be selected.

  5. Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis

    Directory of Open Access Journals (Sweden)

    López-Carro Beatriz

    2009-01-01

    Full Text Available Abstract Homogeneity of cell populations is a prerequisite for the analysis of biochemical and molecular events during male gamete differentiation. Given the complex organization of the mammalian testicular tissue, various methods have been used to obtain enriched or purified cell populations, including flow cell sorting. Current protocols are usually time-consuming and may imply loss of short-lived RNAs, which is undesirable for expression profiling. We describe an optimized method to speed up the preparation of suitable testicular cell suspensions for cytometric analysis of different spermatogenic stages from rodents. The procedure takes only 15 min including testis dissection, tissue cutting, and processing through the Medimachine System (Becton Dickinson. This method could be a substitute for the more tedious and time-consuming cell preparation techniques currently in use.

  6. Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis

    Directory of Open Access Journals (Sweden)

    Rodríguez-Casuriaga Rosana

    2009-03-01

    Full Text Available Abstract Homogeneity of cell populations is a prerequisite for the analysis of biochemical and molecular events during male gamete differentiation. Given the complex organization of the mammalian testicular tissue, various methods have been used to obtain enriched or purified cell populations, including flow cell sorting. Current protocols are usually time-consuming and may imply loss of short-lived RNAs, which is undesirable for expression profiling. We describe an optimized method to speed up the preparation of suitable testicular cell suspensions for cytometric analysis of different spermatogenic stages from rodents. The procedure takes only 15 min including testis dissection, tissue cutting, and processing through the Medimachine System (Becton Dickinson. This method could be a substitute for the more tedious and time-consuming cell preparation techniques currently in use.

  7. Adaptation of ubiquitin-PNA based sperm quality assay for semen evaluation by a conventional flow cytometer and a dedicated platform for flow cytometric semen analysis.

    Science.gov (United States)

    Odhiambo, J F; Sutovsky, M; DeJarnette, J M; Marshall, C; Sutovsky, P

    2011-10-01

    The purpose of semen quality evaluation is to predict the fertility potential of the sample in an objective, rapid and inexpensive manner. However, utilization of sperm quality biomarkers such as ubiquitin and lectin Arachis hypogaea agglutinin (PNA) for flow cytometric semen evaluation might eliminate the need for visual assessment by microscopy. Herein, we demonstrate a robust ubiquitin and PNA-based semen evaluation conducted on a simple, easy to operate, dedicated sperm flow cytometer, EasyCyte Plus (IMV Technologies, L'Aigle, France). Semen samples were collected periodically from two dairy bulls, which were subjected to temporary scrotal insults to induce variable semen quality. Samples were labeled with fluorescently-conjugated anti-ubiquitin antibodies (bind exclusively to the surface of defective sperm) and lectin PNA (binds to acrosomal surface in prematurely capacitated and acrosome-damaged sperm). Fluorescent properties of the samples were measured with a conventional flow cytometer (Becton Dickinson FACScan; Becton Dickinson Corp., Franklin Lakes, NJ, USA) and by the EasyCyte (IMV Technologies) instrument. Data from the two flow cytometers were positively correlated for the percentage of PNA-positive sperm with a damaged acrosome (r = 0.47; P flow cytometric semen evaluation.

  8. Flow cytometric evaluation of physico-chemical impact on Gram-positive and Gram-negative bacteria

    Directory of Open Access Journals (Sweden)

    Antje eFröhling

    2015-09-01

    Full Text Available Since heat sensitivity of fruits and vegetables limits the application of thermal inactivation processes, new emerging inactivation technologies have to be established to fulfil the requirements of food safety without affecting the produce quality. The efficiency of inactivation treatments has to be ensured and monitored. Monitoring of inactivation effects is commonly performed using traditional cultivation methods which have the disadvantage of the time span needed to obtain results.The aim of this study was to compare the inactivation effects of peracetic acid (PAA, ozonated water (O3 and cold atmospheric pressure plasma (CAPP on Gram-positive and Gram-negative bacteria using flow cytometric methods. E. coli cells were completely depolarized after treatment (15 s with 0.25 % PAA at 10 °C, and after treatment (10 s with 3.8 mg l-1 O3 at 12°C. The membrane potential of CAPP treated cells remained almost constant at an operating power of 20 W over a time period of 3 min, and subsequently decreased within 30 s of further treatment. Complete membrane permeabilization was observed after 10 s O3 treatment, but treatment with PAA and CAPP did not completely permeabilize the cells within 2 min and 4 min, respectively. Similar results were obtained for esterase activity. O3 inactivates cellular esterase but esterase activity was detected after 4 min CAPP treatment and 2 min PAA treatment. L. innocua cells and P. carotovorum cells were also permeabilized instantaneously by O3 treatment at concentrations of 3.8 ± 1 mg l-1. However, higher membrane permeabilization of L. innocua and P. carotovorum than of E. coli was observed at CAPP treatment of 20 W. The degree of bacterial damage due to the inactivation processes is highly dependent on treatment parameters as well as on treated bacteria. Important information regarding the inactivation mechanisms can be obtained by flow cytometric measurements and this enables the definition of critical process

  9. Flow cytometric measurement of calcium influx in murine T cell hybrids using Fluo-3 and an organic-anion transport inhibitor.

    Science.gov (United States)

    Baus, E; Urbain, J; Leo, O; Andris, F

    1994-07-12

    A method is described to facilitate flow cytometric analysis of calcium mobilization upon stimulation of murine T cell hybrids. In these transformed cell lines, the accuracy of cytometric measurement of free cytoplasmic calcium with Fluo-3 is compromised by the rapid loss of the intracellular dye. We have found that the addition of sulfinpyrazone, a known organic-anion transporter inhibitor in epithelial cells and in macrophages, severely impairs the leakage of the Fluo-3 probe from the cytoplasmic matrix. Under appropriate conditions, sulfinpyrazone has little effect on the cell physiology and permits the detection of calcium influx in a variety of murine T cell hybrids.

  10. Flow cytometric measurement of RNA synthesis based on bromouridine labelling and combined with measurement of DNA content or cell surface antigen

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J; Larsen, J K

    1993-01-01

    that RNA synthesis increased within the first 24 hours of phytohemagglutinin (PHA) stimulation, reaching a maximum at 48 hours, when cells had entered the cell cycle. Using a new method for flow cytometric dual parameter analysis of BrUrd incorporation and a cell surface antigen, spontaneous RNA synthesis...

  11. Effect of 17 beta-oestradiol on growth curves and flow cytometric DNA distribution of two human breast carcinomas grown in nude mice

    DEFF Research Database (Denmark)

    Brünner, N; Spang-Thomsen, M; Vindeløv, L

    1983-01-01

    The effect of 17 beta-oestradiol on a "receptor positive" and on a "receptor negative" human breast carcinoma grown in nude mice was studied. Experimental growth data were used to determine the effect on tumour growth. Flow cytometric DNA analysis (FCM) performed on tumour tissue obtained...

  12. THE EFFECT OF LABELING INTENSITY, ESTIMATED BY REAL-TIME CONFOCAL LASER SCANNING MICROSCOPY, ON FLOW CYTOMETRIC APPEARANCE AND IDENTIFICATION OF IMMUNOCHEMICALLY LABELED MARINE DINOFLAGELLATES

    NARCIS (Netherlands)

    VRIELING, EG; DRAAIJER, A; VANZEIJL, WJM; PEPERZAK, L; GIESKES, WWC; VEENHUIS, M; Zeijl, Wilhelmus J.M. van

    1993-01-01

    Two different fluorescein isothiocyanate (FITC) conjugates were used to analyze the effect of labeling intensity on the flow cytometric appearance of marine dinoflagellates labeled with antibodies that specifically recognized the outer cell wall. Location of the labeling was revealed by epifluoresce

  13. Flow Cytometric Measurement of [Ca2+]i and pHi in Conjugated Natural Killer Cells and K562 Target Cells during the Cytotoxic Process1,2

    NARCIS (Netherlands)

    van Graft, Marja; van Graft, M.; Kraan, Yvonne M.; Segers-Nolten, Gezina M.J.; Radosevic, K.; Radosevic, Katarina; de Grooth, B.G.; Greve, Jan

    1993-01-01

    We describe a flow cytometric assay that enables one to follow conjugate formation between cytotoxic cells and their target cells during the cytotoxic process. In addition, the internal calcium concentration ([Ca2+]i) and internal pH (pHi) of the conjugated cells can be monitored and directly

  14. Clinical flow cytometric screening of SAP and XIAP expression accurately identifies patients with SH2D1A and XIAP/BIRC4 mutations.

    Science.gov (United States)

    Gifford, Carrie E; Weingartner, Elizabeth; Villanueva, Joyce; Johnson, Judith; Zhang, Kejian; Filipovich, Alexandra H; Bleesing, Jack J; Marsh, Rebecca A

    2014-07-01

    X-linked lymphoproliferative disease is caused by mutations in two genes, SH2D1A and XIAP/BIRC4. Flow cytometric methods have been developed to detect the gene products, SAP and XIAP. However, there is no literature describing the accuracy of flow cytometric screening performed in a clinical lab setting. We reviewed the clinical flow cytometric testing results for 656 SAP and 586 XIAP samples tested during a 3-year period. Genetic testing was clinically performed as directed by the managing physician in 137 SAP (21%) and 115 XIAP (20%) samples. We included these samples for analyses of flow cytometric test accuracy. SH2D1A mutations were detected in 15/137 samples. SAP expression was low in 13/15 (sensitivity 87%, CI 61-97%). Of the 122 samples with normal sequencing, SAP was normal in 109 (specificity 89%, CI 82-94%). The positive predictive values (PPVs) and the negative predictive values (NPVs) were 50% and 98%, respectively. XIAP/BIRC4 mutations were detected in 19/115 samples. XIAP expression was low in 18/19 (sensitivity 95%, CI 73-100%). Of the 96 samples with normal sequencing, 59 had normal XIAP expression (specificity 61%, CI 51-71%). The PPVs and NPVs were 33% and 98%, respectively. Receiver-operating characteristic analysis was able to improve the specificity to 75%. Clinical flow cytometric screening tests for SAP and XIAP deficiencies offer good sensitivity and specificity for detecting genetic mutations, and are characterized by high NPVs. We recommend these tests for patients suspected of having X-linked lymphoproliferative disease type 1 (XLP1) or XLP2. © 2014 Clinical Cytometry Society.

  15. Overview of very small embryonic-like stem cells (VSELs) and methodology of their identification and isolation by flow cytometric methods.

    Science.gov (United States)

    Zuba-Surma, Ewa K; Ratajczak, Mariusz Z

    2010-01-01

    The protocols presented here describe the procedures employed to identify and isolate very small embryonic-like stem cells (VSELs) using flow cytometric technologies including fluorescence-activated cell sorting (FACS). We describe the recommended steps in detail for their successful identification and isolation from adult tissues. These protocols were initially established to isolate such cells from murine bone marrow (BM) and human cord blood (CB) and may also be employed to isolate these primitive cells from other adult organs and embryonic tissues. Here, we focus on some critical parameters/key points required for the successful identification and purification of these rare cells by employing classical flow cytometry. In the last part of this unit, we also discuss a novel flow cytometric tool, ImageStream, an imaging flow cytometer, which allows better identification and morphological analysis of sorted cells.

  16. CD33 monoclonal antibody conjugated Au cluster nano-bioprobe for targeted flow-cytometric detection of acute myeloid leukaemia

    Energy Technology Data Exchange (ETDEWEB)

    Retnakumari, Archana; Jayasimhan, Jasusri; Chandran, Parwathy; Menon, Deepthy; Nair, Shantikumar; Mony, Ullas; Koyakutty, Manzoor, E-mail: manzoork@aims.amrita.edu, E-mail: ullasmony@aims.amrita.edu [Amrita Centre for Nanoscience and Molecular Medicine, Amrita Institute of Medical Science, Cochin 682 041 (India)

    2011-07-15

    Protein stabilized gold nanoclusters (Au-NCs) are biocompatible, near-infrared (NIR) emitting nanosystems having a wide range of biomedical applications. Here, we report the development of a Au-NC based targeted fluorescent nano-bioprobe for the flow-cytometric detection of acute myeloid leukaemia (AML) cells. Au-NCs with {approx} 25-28 atoms showing bright red-NIR fluorescence (600-750 nm) and average size of {approx} 0.8 nm were prepared by bovine serum albumin assisted reduction-cum-stabilization in aqueous phase. The protein protected clusters were conjugated with monoclonal antibody against CD33 myeloid antigen, which is overexpressed in {approx} 99.2% of the primitive population of AML cells, as confirmed by immunophenotyping using flow cytometry. Au-NC-CD33 conjugates having average size of {approx} 12 nm retained bright fluorescence over an extended duration of {approx} a year, as the albumin protein protects Au-NCs against degradation. Nanotoxicity studies revealed excellent biocompatibility of Au-NC conjugates, as they showed no adverse effect on the cell viability and inflammatory response. Target specificity of the conjugates for detecting CD33 expressing AML cells (KG1a) in flow cytometry showed specific staining of {approx} 95.4% of leukaemia cells within 1-2 h compared to a non-specific uptake of {approx} 8.2% in human peripheral blood cells (PBMCs) which are CD33{sup low}. The confocal imaging also demonstrated the targeted uptake of CD33 conjugated Au-NCs by leukaemia cells, thus confirming the flow cytometry results. This study demonstrates that novel nano-bioprobes can be developed using protein protected fluorescent nanoclusters of Au for the molecular receptor targeted flow cytometry based detection and imaging of cancer cells.

  17. A Simple Flow Cytometric Method to Measure Glucose Uptake and Glucose Transporter Expression for Monocyte Subpopulations in Whole Blood.

    Science.gov (United States)

    Palmer, Clovis S; Anzinger, Joshua J; Butterfield, Tiffany R; McCune, Joseph M; Crowe, Suzanne M

    2016-08-12

    Monocytes are innate immune cells that can be activated by pathogens and inflammation associated with certain chronic inflammatory diseases. Activation of monocytes induces effector functions and a concomitant shift from oxidative to glycolytic metabolism that is accompanied by increased glucose transporter expression. This increased glycolytic metabolism is also observed for trained immunity of monocytes, a form of innate immunological memory. Although in vitro protocols examining glucose transporter expression and glucose uptake by monocytes have been described, none have been examined by multi-parametric flow cytometry in whole blood. We describe a multi-parametric flow cytometric protocol for the measurement of fluorescent glucose analog 2-NBDG uptake in whole blood by total monocytes and the classical (CD14(++)CD16(-)), intermediate (CD14(++)CD16(+)) and non-classical (CD14(+)CD16(++)) monocyte subpopulations. This method can be used to examine glucose transporter expression and glucose uptake for total monocytes and monocyte subpopulations during homeostasis and inflammatory disease, and can be easily modified to examine glucose uptake for other leukocytes and leukocyte subpopulations within blood.

  18. HLA-targeted flow cytometric sorting of blood cells allows separation of pure and viable microchimeric cell populations.

    Science.gov (United States)

    Drabbels, Jos J M; van de Keur, Carin; Kemps, Berit M; Mulder, Arend; Scherjon, Sicco A; Claas, Frans H J; Eikmans, Michael

    2011-11-10

    Microchimerism is defined by the presence of low levels of nonhost cells in a person. We developed a reliable method for separating viable microchimeric cells from the host environment. For flow cytometric cell sorting, HLA antigens were targeted with human monoclonal HLA antibodies (mAbs). Optimal separation of microchimeric cells (present at a proportion as low as 0.01% in artificial mixtures) was obtained with 2 different HLA mAbs, one targeting the chimeric cells and the other the background cells. To verify purity of separated cell populations, flow-sorted fractions of 1000 cells were processed for DNA analysis by HLA-allele-specific and Y-chromosome-directed real-time quantitative PCR assays. After sorting, PCR signals of chimeric DNA markers in the positive fractions were significantly enhanced compared with those in the presort samples, and they were similar to those in 100% chimeric control samples. Next, we demonstrate applicability of HLA-targeted FACS sorting after pregnancy by separating chimeric maternal cells from child umbilical cord mononuclear cells. Targeting allelic differences with anti-HLA mAbs with FACS sorting allows maximal enrichment of viable microchimeric cells from a background cell population. The current methodology enables reliable microchimeric cell detection and separation in clinical specimens.

  19. Laser-based flow cytometric analysis of genotoxicity of humans exposed to ionizing radiation during the Chernobyl accident

    Science.gov (United States)

    Jensen, Ronald H.; Bigbee, William L.; Langlois, Richard G.; Grant, Stephen G.; Pleshanov, Pavel G.; Chirkov, Andre A.; Pilinskaya, Maria A.

    1991-05-01

    An analytical technique has been developed that allows laser-based flow cytometric measurement of the frequency of red blood cells that have lost allele-specific expression of a cell surface antigen due to genetic toxicity in bone marrow precursor cells. Previous studies demonstrated a correlation of such effects with the exposure of each individual to mutagenic phenomena, such as ionizing radiation, and the effects can persist for the lifetime of each individual. During the emergency response to the nuclear power plant accidert at Chemobyl, Ukraine, USSR, a number of people were exposed to whole body doses of ioniing radiation. Some of these individuals were tested with this laser-based assay and found to express a dose-dependent increase in the frequency of variant red blood cells that appears to be a persistent biological effect. This effect is similar to that which was previously observed in individuals who were exposed to ionizing radiation at Hiroshima in 1945 because of the A-bomb explosion. All data indicate that this assay might well be used as a biodosimeter to estimate radiation dose and also as an element to be used for estimating the risk of each individual to develop cancer due to radiation exposure.

  20. A whole blood flow cytometric determination of platelet activation by unfractionated and low molecular weight heparin in vitro.

    Science.gov (United States)

    Klein, Bernd; Faridi, Andreé; von Tempelhoff, G F; Heilmann, Lothar; Mittermayer, Christian; Rath, Werner

    2002-12-15

    The influence of unfractionated (Heparin-Natrium) and low-molecular heparin (Fragmin(R)) on platelet activation in whole blood was investigated by FACS analysis in vitro using antibodies against glycoprotein (gp) IIb/IIIa (CD 41), GMP 140 (CD 62P), gp 53 (CD 63) and fibrinogen. Samples were also labeled with anti-gp Ib (CD 42b). Neither unfractionated heparin (UFH) nor low molecular weight heparin (LMWH) led to significant (i.e., p<0.05) changes in fluorescence intensities of platelets labeled with anti-gp IIb/IIIa or anti-gp 53. Significant platelet activation due to unfractionated heparin could be observed by labeling with anti-GMP 140 (UFH: p=0.009; LMWH: p=0.16). The proportion of platelets with surface-bound fibrinogen was significantly increased (UFH: p=0.00006; LMWH: p=0.008). After incubation with heparins, activation ability of platelets by adenosine diphosphate (ADP) was significantly increased. The potentiating action of unfractionated heparin was larger. Therefore, flow cytometric results of platelet activation in patients receiving heparin should be interpreted carefully.

  1. Estimation of the Whitefly Bemisia tabaci Genome Size Based on k-mer and Flow Cytometric Analyses

    Directory of Open Access Journals (Sweden)

    Wenbo Chen

    2015-07-01

    Full Text Available Whiteflies of the Bemisia tabaci (Hemiptera: Aleyrodidae cryptic species complex are among the most important agricultural insect pests in the world. These phloem-feeding insects can colonize over 1000 species of plants worldwide and inflict severe economic losses to crops, mainly through the transmission of pathogenic viruses. Surprisingly, there is very little genomic information about whiteflies. As a starting point to genome sequencing, we report a new estimation of the genome size of the B. tabaci B biotype or Middle East-Asia Minor 1 (MEAM1 population. Using an isogenic whitefly colony with over 6500 haploid male individuals for genomic DNA, three paired-end genomic libraries with insert sizes of ~300 bp, 500 bp and 1 Kb were constructed and sequenced on an Illumina HiSeq 2500 system. A total of ~50 billion base pairs of sequences were obtained from each library. K-mer analysis using these sequences revealed that the genome size of the whitefly was ~682.3 Mb. In addition, the flow cytometric analysis estimated the haploid genome size of the whitefly to be ~690 Mb. Considering the congruency between both estimation methods, we predict the haploid genome size of B. tabaci MEAM1 to be ~680–690 Mb. Our data provide a baseline for ongoing efforts to assemble and annotate the B. tabaci genome.

  2. Flow Cytometric Evaluation of Human Neutrophil Apoptosis During Nitric Oxide Generation In Vitro: The Role of Exogenous Antioxidants

    Directory of Open Access Journals (Sweden)

    Zofia Sulowska

    2005-01-01

    in vitro. The effect of exogenous supply of NO donors such as SNP, SIN-1, and GEA-3162 on the course of human neutrophil apoptosis and the role of extracellular antioxidants in this process was investigated. Isolated from peripheral blood, neutrophils were cultured in the presence or absence of NO donor compounds and antioxidants for 8, 12, and 20 hours. Apoptosis of neutrophils was determined in vitro by flow cytometric analysis of cellular DNA content and Annexin V protein binding to the cell surface. Exposure of human neutrophils to GEA-3162 and SIN-1 significantly accelerates and enhances their apoptosis in vitro in a time-dependent fashion. In the presence of SNP, intensification of apoptosis has not been revealed until 12 hours after the culture. The inhibition of GEA-3162- and SIN-1-mediated neutrophil apoptosis by superoxide dismutase (SOD but not by catalase (CAT was observed. Our results show that SOD and CAT can protect neutrophils against NO-donors-induced apoptosis and suggest that the interaction of NO and oxygen metabolites signals may determine the destructive or protective role of NO donor compounds during apoptotic neutrophil death.

  3. Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug-induced cell cycle perturbations in vitro

    DEFF Research Database (Denmark)

    Engelholm, S A; Spang-Thomsen, M; Vindeløv, L L

    1986-01-01

    A method based on detection of drug-induced cell cycle perturbation by flow cytometric DNA analysis has previously been described in Ehrlich ascites tumors as a way to estimate chemosensitivity. The method is extended to test human small-cell carcinoma of the lung. Three tumors with different...... sensitivities to melphalan in nude mice were used. Tumors were disaggregated by a combined mechanical and enzymatic method and thereafter have incubated with different doses of melphalan. After incubation the cells were plated in vitro on agar, and drug induced cell cycle changes were monitored by flow...... cytometric DNA analysis. Melphalan produced a dose-related S phase accumulation in the two sensitive tumors, whereas no changes in the cell cycle distribution were found in the resistant tumor. The size of S phase accumulation correlated to the chemosensitivity in vivo. For low concentrations of melphalan...

  4. Improved flow cytometric detection of minimal residual disease in childhood acute lymphoblastic leukemia

    NARCIS (Netherlands)

    Denys, B.; van der Sluijs-Gelling, A. J.; Homburg, C.; van der Schoot, C. E.; de Haas, V.; Philippe, J.; Pieters, R.; van Dongen, J. J. M.; van der Velden, V. H. J.

    2013-01-01

    Most current treatment protocols for acute lymphoblastic leukemia (ALL) include minimal residual disease (MRD) diagnostics, generally based on PCR analysis of rearranged antigen receptor genes. Although flow cytometry (FCM) can be used for MRD detection as well, discordant FCM and PCR results are ob

  5. Flow cytometric measurement of RNA synthesis using bromouridine labelling and bromodeoxyuridine antibodies

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J; Christiansen, J

    1993-01-01

    Nuclear RNA synthesis can be analysed by flow cytometry of cells labelled with 5-bromouridine (BrUrd) and stained with anti-bromodeoxyuridine (BrdUrd) antibody and FITC-conjugated secondary antibody. A panel of 5 different commercially available anti-BrdUrd antibodies was tested on cells of a HL-...

  6. Microbial Eco-Physiology of the human intestinal tract: a flow cytometric approach

    NARCIS (Netherlands)

    Amor, Ben K.

    2004-01-01

    This thesis describes a multifaceted approach to further enhance our view of the complex human intestinal microbial ecosystem. This approach combines me advantages of flow cyrometry (FCM), a single cell and high-throughput technology, and molecular techniques that have proven themselves to be invalu

  7. Microbial Eco-Physiology of the human intestinal tract: a flow cytometric approach

    NARCIS (Netherlands)

    Amor, Ben K.

    2004-01-01

    This thesis describes a multifaceted approach to further enhance our view of the complex human intestinal microbial ecosystem. This approach combines me advantages of flow cyrometry (FCM), a single cell and high-throughput technology, and molecular techniques that have proven themselves to be

  8. Microbial Eco-Physiology of the human intestinal tract: a flow cytometric approach

    NARCIS (Netherlands)

    Amor, Ben K.

    2004-01-01

    This thesis describes a multifaceted approach to further enhance our view of the complex human intestinal microbial ecosystem. This approach combines me advantages of flow cyrometry (FCM), a single cell and high-throughput technology, and molecular techniques that have proven themselves to be invalu

  9. Comparison of monoclonal antibodies reactive with lymphocyte subsets in routinely fixed paraffin-embedded material: flow cytometric analyses, immunoperoxidase staining and influence of fixatives.

    Directory of Open Access Journals (Sweden)

    Yoshino,Tadashi

    1990-10-01

    Full Text Available We have attempted to clarify the characteristics of monoclonal antibodies (MAbs detecting lymphocyte subsets in fixed materials. We examined by means of flow cytometric technique influences of fixatives and reactivity with malignant lymphomas (MLs. Specific markers for T-cells were UCHL1 and OPD4, which reacted especially with helper/inducer T-cells. MT1 recognized almost all of T-cells from peripheral blood and tonsils, but reacted with a part of B-MLs. As for B-cell markers, L26 was the most reliable marker for B-MLs. L26 and MB1 antigens could not be detected on living cells flow cytometrically. LN1 reacted with a part of T-cells as well as B-cells, but fluorescent intensity of the former was apparently stronger than that of the latter. Although LN2 antigen was located mainly in the cytoplasm close to the nuclear membrane immunohistochemically, it could be detected on living cells flow cytometrically. LN2 positive cells belonged to B-cells in peripheral blood and tonsils. When fixed for relatively short time, B5 and buffered formalin were better for examining MAbs than non-buffered formalin and ethanol.

  10. A sensitive flow cytometric methodology for studying the binding of L. chagasi to canine peritoneal macrophages

    Directory of Open Access Journals (Sweden)

    Mosser David M

    2005-05-01

    Full Text Available Abstract Background The Leishmania promastigote-macrophage interaction occurs through the association of multiple receptors on the biological membrane surfaces. The success of the parasite infection is dramatically dependent on this early interaction in the vertebrate host, which permits or not the development of the disease. In this study we propose a novel methodology using flow cytometry to study this interaction, and compare it with a previously described "in vitro" binding assay. Methods To study parasite-macrophage interaction, peritoneal macrophages were obtained from 4 dogs and adjusted to 3 × 106 cells/mL. Leishmania (Leishmania chagasi parasites (stationary-phase were adjusted to 5 × 107 cells/mL. The interaction between CFSE-stained Leishmania chagasi and canine peritoneal macrophages was performed in polypropylene tubes to avoid macrophage adhesion. We carried out assays in the presence or absence of normal serum or in the presence of a final concentration of 5% of C5 deficient (serum from AKR/J mice mouse serum. Then, the number of infected macrophages was counted in an optical microscope, as well as by flow citometry. Macrophages obtained were stained with anti-CR3 (CD11b/CD18 antibodies and analyzed by flow citometry. Results Our results have shown that the interaction between Leishmania and macrophages can be measured by flow cytometry using the fluorescent dye CFSE to identify the Leishmania, and measuring simultaneously the expression of an important integrin involved in this interaction: the CD11b/CD18 (CR3 or Mac-1 β2 integrin. Conclusion Flow cytometry offers rapid, reliable and sensitive measurements of single cell interactions with Leishmania in unstained or phenotypically defined cell populations following staining with one or more fluorochromes.

  11. Current International Flow Cytometric Practices for the Detection and Monitoring of Paroxysmal Nocturnal Haemoglobinuria (PNH) clones: A UK NEQAS Survey.

    Science.gov (United States)

    Fletcher, Matthew; Whitby, Liam; Whitby, Alison; Barnett, David

    2016-03-02

    Background Paroxysmal Nocturnal Haemoglobinuria (PNH) is a rare acquired genetic disorder, with an incidence of approximately 1.3 new cases per million population per year. Evidence from the UK National External Quality Assessment Service for Leucocyte Immunophenotyping (UK NEQAS LI) programme suggested major discrepancies on how PNH testing is undertaken. To investigate this we surveyed laboratories in the UK NEQAS LI PNH programme and report here the findings. Method A questionnaire was distributed to all centres registered in UK NEQAS LI flow cytometry programmes (n=1587). Comprising several subsections, it covered the majority of clinical flow cytometric practices. Participants completed a general section and then the subsections relevant to their laboratory repertoire. One subsection contained 34 questions regarding practices in PNH clone detection. Results A total of 105 laboratories returned results for the PNH section; the results demonstrated lack of consensus in all areas of PNH testing. Variation was seen in gating and testing strategies, sensitivity levels and final reporting of test results. Several incorrect practices were highlighted such as inappropriate antibody selection and failure to wash the red blood cells (RBCs) prior to analysis. Conclusion Despite the availability of consensus guidelines there appears to be no agreement in the detection and monitoring of PNH. We found only fourteen centres using methods compatible with the International Clinical Cytometry Society guidelines. Of specific note we found that no two laboratories used the same method. This technical variation could lead to incorrect diagnoses, highlighting the need for better adoption and understanding of consensus practices. This article is protected by copyright. All rights reserved.

  12. Flow cytometric gating for spleen monocyte and DC subsets: differences in autoimmune NOD mice and with acute inflammation.

    Science.gov (United States)

    Dong, Matthew B; Rahman, M Jubayer; Tarbell, Kristin V

    2016-05-01

    The role of antigen presenting cells (APCs) in the pathogenesis of autoimmune and other inflammatory diseases is now better understood due to advances in multicolor flow cytometry, gene expression analysis of APC populations, and functional correlation of mouse to human APC populations. A simple but informative nomenclature of conventional and plasmacytoid dendritic cell subsets (cDC1, cDC2, pDC) and monocyte-derived populations incorporates these advances, but accurate subset identification is critical. Ambiguous gating schemes and alterations of cell surface markers in inflammatory condition can make comparing results between studies difficult. Both acute inflammation, such as TLR-ligand stimulation, and chronic inflammation as found in mouse models of autoimmunity can alter DC subset gating. Here, we address these issues using in vivo CpG stimulation as an example of acute inflammation and the non-obese diabetic (NOD) mouse as a model of chronic inflammation.We provide a flow cytometric antibody panel and gating scheme that differentiate 2 monocytic and 3DC subsets in the spleen both at steady state and after CpG stimulation. Using this method, we observed differences in the composition of NOD DCs that have been previously reported, and newly identified increases in the number of NOD monocyte-derived DCs. Finally, we established a protocol for DC phosphoflow to measure the phosphorylation state of intracellular proteins, and use it to confirm functional differences in the identified subsets. Therefore, we present optimized methods for distinguishing monocytic and DC populations with and without inflammation and/or autoimmunity associated with NOD mice.

  13. High-throughput flow cytometric screening of combinatorial chemistry bead libraries for proteomics and drug discovery

    Science.gov (United States)

    Leary, James F.; Reece, Lisa M.; Yang, Xian-Bin; Gorenstein, David

    2005-04-01

    For proteomics drug discovery applications, combinatorial microbead thioaptamer libraries (one thioaptamer sequence per bead) are being created by split synthesis method, creating a "proteomics library" of protein capture beads which can be analyzed by high-throughput screening methods in this case, flow cytometry and cell sorting. Thioaptamers, oligonucleotides with thiophosphate backbone substitutions, function like antibodies in terms of recognizing specific protein sequences but have a number of advantages over antibody libraries. These proteomics beads can then be analyzed by high-speed flow cytometry and sorted to single-bead level depending on relative fluorescence brightness of fluorescently-labeled proteins, or for a specific protein from all of the molecules of cell subpopulations being analyzed. The thioaptamer sequences on a given bead showing high affinity for that protein can then be sequenced. Alternatively, the protein-capturing beads can be analyzed by MALDI-TOF mass spectrometry for analysis of the bound proteins. The beads can be thought of as equivalent to single-element positions of a proteomics chip arrays but with the advantage of being able to much more rapidly analyze hundreds of millions of possible amino acid sequences/epitopes on the basis of thioaptamer sequence affinities to select single sequences of interest. Additionally, those beads can be manipulated and isolated at the single bead level by high-throughput flow cytometry/cell sorting for subsequent sequencing of the thioaptamer sequences.

  14. Flow cytometric determination of osmotic behaviour of animal erythrocytes toward their engineering for drug delivery

    Directory of Open Access Journals (Sweden)

    Kostić Ivana T.

    2015-01-01

    Full Text Available Despite the fact that the methods based on the osmotic properties of the cells are the most widely used for loading of drugs in human and animal erythrocytes, data related to the osmotic properties of erythrocytes derived from animal blood are scarce. This work was performed with an aim to investigate the possibility of use the flow cytometry as a tool for determination the osmotic behaviour of porcine and bovine erythrocytes, and thus facilitate the engineering of erythrocytes from animal blood to be drug carriers. The method of flow cytometry successfully provided the information about bovine and porcine erythrocyte osmotic fragility, and made the initial steps in assessment of erythrocyte shape in a large number of erythrocytes. Although this method is not able to confirm the swelling of pig erythrocytes, it indicated to the differences in pig erythrocytes that had basic hematological parameters inside and outside the reference values. In order to apply/use the porcine and bovine erythrocytes as drug carriers, the method of flow cytometry, confirming the presence of osmotically different fractions of red blood cells, indicated that various amounts of the encapsulated drug in porcine and bovine erythrocytes can be expected.

  15. Flow Cytometric Analysis of Particle-bound Bet v 1 Allergen in PM10.

    Science.gov (United States)

    Süring, Katrin; Bach, Sabine; Höflich, Conny; Straff, Wolfgang

    2016-11-19

    Flow cytometry is a method widely used to quantify suspended solids such as cells or bacteria in a size range from 0.5 to several tens of micrometers in diameter. In addition to a characterization of forward and sideward scatter properties, it enables the use of fluorescent labeled markers like antibodies to detect respective structures. Using indirect antibody staining, flow cytometry is employed here to quantify birch pollen allergen (precisely Bet v 1)-loaded particles of 0.5 to 10 µm in diameter in inhalable particulate matter (PM10, particle size ≤10 µm in diameter). PM10 particles may act as carriers of adsorbed allergens possibly transporting them to the lower respiratory tract, where they could trigger allergic reactions. So far the allergen content of PM10 has been studied by means of enzyme linked immunosorbent assays (ELISAs) and scanning electron microscopy. ELISA measures the dissolved and not the particle-bound allergen. Compared to scanning electron microscopy, which can visualize allergen-loaded particles, flow cytometry may additionally quantify them. As allergen content of ambient air can deviate from birch pollen count, allergic symptoms might perhaps correlate better with allergen exposure than with pollen count. In conjunction with clinical data, the presented method offers the opportunity to test in future experiments whether allergic reactions to birch pollen antigens are associated with the Bet v 1 allergen content of PM10 particles >0.5 µm.

  16. Simple flow cytometric detection of haemozoin containing leukocytes and erythrocytes for research on diagnosis, immunology and drug sensitivity testing

    Directory of Open Access Journals (Sweden)

    Grobusch Martin P

    2011-03-01

    Full Text Available Abstract Background Malaria pigment (haemozoin, Hz has been the focus of diverse research efforts. However, identification of Hz-containing leukocytes or parasitized erythrocytes is usually based on microscopy, with inherent limitations. Flow cytometric detection of depolarized Side-Scatter is more accurate and its adaptation to common bench top flow cytometers might allow several applications. These can range from the ex-vivo and in-vitro detection and functional analysis of Hz-containing leukocytes to the detection of parasitized Red-Blood-Cells (pRBCs to assess antimalarial activity. Methods A standard benchtop flow cytometer was adapted to detect depolarized Side-Scatter. Synthetic and Plasmodium falciparum Hz were incubated with whole blood and PBMCs to detect Hz-containing leukocytes and CD16 expression on monocytes. C5BL/6 mice were infected with Plasmodium berghei ANKA or P. berghei NK65 and Hz-containing leukocytes were analysed using CD11b and Gr1 expression. Parasitized RBC from infected mice were identified using anti-Ter119 and SYBR green I and were analysed for depolarized Side Scatter. A highly depolarizing RBC population was monitored in an in-vitro culture incubated with chloroquine or quinine. Results A flow cytometer can be easily adapted to detect depolarized Side-Scatter and thus, intracellular Hz. The detection and counting of Hz containing leukocytes in fresh human or mouse blood, as well as in leukocytes from in-vitro experiments was rapid and easy. Analysis of CD14/CD16 and CD11b/Gr1 monocyte expression in human or mouse blood, in a mixed populations of Hz-containing and non-containing monocytes, appears to show distinct patterns in both types of cells. Hz-containing pRBC and different maturation stages could be detected in blood from infected mice. The analysis of a highly depolarizing population that contained mature pRBC allowed to assess the effect of chloroquine and quinine after only 2 and 4 hours, respectively

  17. Flow cytometric assessment of Streptococcus mutans viability after exposure to blue light-activated curcumin.

    Science.gov (United States)

    Manoil, Daniel; Filieri, Anna; Gameiro, Cécile; Lange, Norbert; Schrenzel, Jacques; Wataha, John C; Bouillaguet, Serge

    2014-09-01

    Streptococcus mutans biofilms are considered as primary causative agents of dental caries. Photodynamic antimicrobial chemotherapy (PACT) has been recently proposed as a strategy for inactivating dental biofilms. This study aimed to investigate the effect of blue light-activated curcumin on S. mutans viability and to explore its potential as a new anti-caries therapeutic agent. The effect of different concentrations and incubation times of photo-activated curcumin on the survival of S. mutans in planktonic and biofilm models of growth was assessed by flow cytometry. Streptococcus mutans in planktonic suspensions or biofilms formed on hydroxyapatite disks were incubated for 5 or 10min with curcumin prior to blue light activation. Bacteria were labeled with SYTO 9 and propidium iodide before viability was assessed by flow cytometry. Results were statistically analyzed using one-way ANOVA and Tukey multiple comparison intervals (α=0.05). For planktonic cultures, 0.2μM of light-activated curcumin significantly reduced S. mutans viability (p<0.05). For biofilm cultures, light-activated curcumin at concentration of 40-60μM only suppressed viability by 50% (p<0.05). Independently of the mode of growth, incubation time has no significant effect on PACT efficiency. This study indicates that blue light-activated curcumin can efficiently inactivate planktonic cultures of S. mutans whereas biofilms were more resistant to treatment. Flow cytometry allowed the detection of bacteria with damaged membranes that were unable to replicate and grow after cell sorting. Further studies seem warranted to optimize the efficacy of light-activated curcumin against S. mutans biofilms. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Flow cytometric determination of genome size in European sunbleak Leucaspius delineatus (Heckel, 1843).

    Science.gov (United States)

    Filipiak, Marta; Tylko, Grzegorz; Kilarski, Wincenty

    2012-04-01

    The aim of this study was to compare DNA content in hepatocyte and erythrocyte nuclei of the European sunbleak, Leucaspius delineatus, in relation to nuclear and cell size by means of flow cytometry and fluorescence microscopy. The DNA standards, chicken and rainbow trout erythrocytes, were prepared in parallel with both cell types, with initial separation of liver cells in pepsin solution followed by cell filtering. Standards and investigated cells were stained with a mixture of propidium iodide, citric acid, and Nonidet P40 in the presence of RNAse, and fluorescence of at least 50,000 nuclei was analyzed by flow cytometry. Average cell size was determined by flow cytometry, using fresh cell suspension in relation to latex beads of known diameter. The size of nuclei was examined on the basis of digital micrographs obtained by fluorescence microscopy after nuclei staining with DAPI. The sunbleak's erythrocyte nuclei contain 2.25 ± 0.06 pg of DNA, whereas the hepatocyte nuclei contain 2.46 ± 0.06 pg of DNA. This difference in DNA content was determined spectroscopically using isolated DNA from the two cell types. The modal diameters of the erythrocytes and hepatocytes were estimated to be 5.1 ± 0.2 and 22.3 ± 5.0 μm, respectively, and the corresponding modal dimensions of their nuclei (measured as surface area) were 15.2 and 21.4 μm(2), respectively. The nucleoplasmic index, as calculated from diameters estimated from surface area of nuclear profiles, was 2.51 for the erythrocytes compared with 0.08 for hepatocytes.

  19. Flow cytometric analysis of T lymphocyte proliferation in vivo by EdU incorporation.

    Science.gov (United States)

    Sun, Xiaojing; Zhang, Chunpan; Jin, Hua; Sun, Guangyong; Tian, Yue; Shi, Wen; Zhang, Dong

    2016-12-01

    Monitoring T lymphocyte proliferation, especially in vivo, is essential for the evaluation of adaptive immune reactions. Flow cytometry-based proliferation assays have advantages in measuring cell division of different T lymphocyte subsets at the same time by multicolor labelling. In this study, we aimed to establish the use of 5-Ethynyl-2'-deoxyuridine (EdU) incorporation in vivo to monitor T lymphocyte proliferation by flow cytometry with an adoptive transfer model. We found that fixation followed by permeabilization preserved T cell surface antigens and had no obvious effects on the fluorescence intensity of APC, PE, PE-Cy7, FITC and PerCP-Cy5.5 when the concentration of the permeabilization reagents was optimized. However, the click reaction resulted in a significant decrease in the fluorescence intensity of PE and PE-Cy7, and surface staining after the click reaction improved the fluorescence intensity. Thus, an extra step of blocking with PBS with 3% FBS between the click reaction and cell surface staining is needed. Furthermore, the percentage of EdU-positive cells increased in a dose-dependent manner, and the saturated dose of EdU was 20mg/kg. Intraperitoneal and intravenous injection had no differences in lymphocyte proliferation detection with EdU in vivo. In addition, T cell proliferation measured by EdU incorporation was comparable to BrdU but was lower than CFSE labelling. In conclusion, we optimized the protocols for EdU administration in vivo and staining in vitro, providing a feasible method for the measurement of T lymphocyte proliferation with EdU incorporation by flow cytometry in vivo.

  20. Quantitative assessment of BAX transcript and flow cytometric expression in acute myeloid leukemia: a prospective study.

    Science.gov (United States)

    Sharawat, Surender Kumar; Raina, Vinod; Kumar, Lalit; Sharma, Atul; Bakhshi, Radhika; Vishnubhatla, Sreenivas; Gupta, Ritu; Bakhshi, Sameer

    2014-10-01

    Quantitative assessment of BAX transcripts and protein in acute myeloid leukemia (AML). We quantitatively evaluated BAX gene transcripts by real-time polymerase chain reaction (TaqMan probe chemistry) and protein expression by flow cytometry. Consecutive 112 AML patients with a median age of 16 (1-59) years were recruited in the study. By flow cytometry, the percentage expression was in linear correlation with relative median fluorescent intensity (RMFI; R = 0.4425; P BAX with its RMFI (R = -0.0559; P = 0.586). The expression of the BAX at both protein and transcript level was significantly higher in AML patients as compared with normal control. RMFI of the BAX were higher in the cohort with lower white blood cell count (P = 0.029). None of the other baseline characteristics correlated with either the BAX transcript or the RMFI. BAX expression did not correlate with complete remission rate, event free, disease free, and overall survival. BAX gene expression in AML was evaluated first time with two different methods but did not correlate with the survival outcome.

  1. Flow cytometric measurement of the cellular propagation of TDP-43 aggregation.

    Science.gov (United States)

    Zeineddine, Rafaa; Whiten, Daniel R; Farrawell, Natalie E; McAlary, Luke; Hanspal, Maya A; Kumita, Janet R; Wilson, Mark R; Yerbury, Justin J

    2017-05-04

    Amyotrophic lateral sclerosis is a devastating neuromuscular degenerative disease characterized by a focal onset of motor neuron loss, followed by contiguous outward spreading of pathology including TAR DNA-binding protein of 43 kDa (TDP-43) aggregates. Previous work suggests that TDP-43 can move between cells. Here we used a novel flow cytometry technique (FloIT) to analyze TDP-43 inclusions and propagation. When cells were transfected to express either mutant G294A TDP-43 fused to GFP or wild type TDP-43fused to tomato red and then co-cultured, flow cytometry detected intact cells containing both fusion proteins and using FloIT detected an increase in the numbers of inclusions in lysates from cells expressing wild type TDP-43-tomato. Furthermore, in this same model, FloIT analyses detected inclusions containing both fusion proteins. These results imply the transfer of TDP-43 fusion proteins between cells and that this process can increase aggregation of wild-type TDP-43 by a mechanism involving co-aggregation with G294A TDP-43.

  2. Optimization of flow cytometric detection and cell sorting of transgenic Plasmodium parasites using interchangeable optical filters.

    Science.gov (United States)

    Vorobjev, Ivan A; Buchholz, Kathrin; Prabhat, Prashant; Ketman, Kenneth; Egan, Elizabeth S; Marti, Matthias; Duraisingh, Manoj T; Barteneva, Natasha S

    2012-09-05

    Malaria remains a major cause of morbidity and mortality worldwide. Flow cytometry-based assays that take advantage of fluorescent protein (FP)-expressing malaria parasites have proven to be valuable tools for quantification and sorting of specific subpopulations of parasite-infected red blood cells. However, identification of rare subpopulations of parasites using green fluorescent protein (GFP) labelling is complicated by autofluorescence (AF) of red blood cells and low signal from transgenic parasites. It has been suggested that cell sorting yield could be improved by using filters that precisely match the emission spectrum of GFP. Detection of transgenic Plasmodium falciparum parasites expressing either tdTomato or GFP was performed using a flow cytometer with interchangeable optical filters. Parasitaemia was evaluated using different optical filters and, after optimization of optics, the GFP-expressing parasites were sorted and analysed by microscopy after cytospin preparation and by imaging cytometry. A new approach to evaluate filter performance in flow cytometry using two-dimensional dot blot was developed. By selecting optical filters with narrow bandpass (BP) and maximum position of filter emission close to GFP maximum emission in the FL1 channel (510/20, 512/20 and 517/20; dichroics 502LP and 466LP), AF was markedly decreased and signal-background improve dramatically. Sorting of GFP-expressing parasite populations in infected red blood cells at 90 or 95% purity with these filters resulted in 50-150% increased yield when compared to the standard filter set-up. The purity of the sorted population was confirmed using imaging cytometry and microscopy of cytospin preparations of sorted red blood cells infected with transgenic malaria parasites. Filter optimization is particularly important for applications where the FP signal and percentage of positive events are relatively low, such as analysis of parasite-infected samples with in the intention of gene

  3. Optimization of flow cytometric detection and cell sorting of transgenic Plasmodium parasites using interchangeable optical filters

    Directory of Open Access Journals (Sweden)

    Vorobjev Ivan A

    2012-09-01

    Full Text Available Abstract Background Malaria remains a major cause of morbidity and mortality worldwide. Flow cytometry-based assays that take advantage of fluorescent protein (FP-expressing malaria parasites have proven to be valuable tools for quantification and sorting of specific subpopulations of parasite-infected red blood cells. However, identification of rare subpopulations of parasites using green fluorescent protein (GFP labelling is complicated by autofluorescence (AF of red blood cells and low signal from transgenic parasites. It has been suggested that cell sorting yield could be improved by using filters that precisely match the emission spectrum of GFP. Methods Detection of transgenic Plasmodium falciparum parasites expressing either tdTomato or GFP was performed using a flow cytometer with interchangeable optical filters. Parasitaemia was evaluated using different optical filters and, after optimization of optics, the GFP-expressing parasites were sorted and analysed by microscopy after cytospin preparation and by imaging cytometry. Results A new approach to evaluate filter performance in flow cytometry using two-dimensional dot blot was developed. By selecting optical filters with narrow bandpass (BP and maximum position of filter emission close to GFP maximum emission in the FL1 channel (510/20, 512/20 and 517/20; dichroics 502LP and 466LP, AF was markedly decreased and signal-background improve dramatically. Sorting of GFP-expressing parasite populations in infected red blood cells at 90 or 95% purity with these filters resulted in 50-150% increased yield when compared to the standard filter set-up. The purity of the sorted population was confirmed using imaging cytometry and microscopy of cytospin preparations of sorted red blood cells infected with transgenic malaria parasites. Discussion Filter optimization is particularly important for applications where the FP signal and percentage of positive events are relatively low, such as analysis

  4. Flow cytometric applicability to evaluate UV inactivation of phytoplankton in marine water samples.

    Science.gov (United States)

    Olsen, Ranveig Ottoey; Hess-Erga, Ole-Kristian; Larsen, Aud; Thuestad, Gunnar; Tobiesen, August; Hoell, Ingunn Alne

    2015-07-15

    Disinfection of microbes is of importance to prevent the spread of pathogens and non-indigenous species in the environment. Here we test the applicability of using flow cytometry (FCM) to evaluate inactivation of the phytoplankter Tetraselmis suecica after UV irradiation and labeling with the esterase substrate 5-carboxyfluorescein diacetate acetoxymethyl ester (CFDA-AM). Non-irradiated and UV irradiated samples were analyzed with the plate count technique and FCM for 24 days. The numbers of colony forming units were used as a standard to develop a FCM protocol. Our protocol readily distinguishes live and dead cells, but challenges were encountered when determining whether UV damaged cells are dying or repairable. As damaged cells can represent a risk to aquatic organisms and/or humans, this was taken into account when developing the FCM protocol. In spite of the above mentioned challenges we argue that FCM represents an accurate and rapid method to analyze T. suecica samples.

  5. Determination of micro-litre volumes with high accuracy for flow cytometric blood cell counting

    Science.gov (United States)

    Reitz, S.; Kummrow, A.; Kammel, M.; Neukammer, J.

    2010-07-01

    We have gravimetrically calibrated the volumes dispensed by 1 mL syringes in the range between 1 µL and 100 µL using ultra-pure water. Protocols are based on series of consecutive difference measurements of masses in order to precisely compensate for evaporation, being the most important disturbing quantity. We determined expanded uncertainties of volume measurements for glass syringes of typically 0.2% (expansion factor 2) when dispensing volumes of 10 µL. For polypropylene syringes, selected with respect to the manufacturer, expanded uncertainties of 0.25% (expansion factor 2) were observed. Calibrated syringes were applied for measuring concentrations of blood cells in a flow cytometer demonstrating the capability to determine reference measurement values. Since the direct interaction of blood cells and syringe walls may lead to cell adhesion, glass syringes as well as (disposable) polypropylene syringes were calibrated.

  6. Flow cytometric assay detecting cytotoxicity against human endogenous retrovirus antigens expressed on cultured multiple sclerosis cells

    DEFF Research Database (Denmark)

    Møller-Larsen, A; Brudek, T; Petersen, T

    2013-01-01

    as control antibody. Without antibodies this system is suitable for analyses of natural killer cell activity. In optimization of the assay we have used effector lymphocytes from healthy donors. The most effective effector cells are CD56(+) cells. CD8(+) T cells also express CD107a in ADCC. Using the adapted......Damage of target cells by cytotoxicity, either mediated by specific lymphocytes or via antibody-dependent reactions, may play a decisive role in causing the central nervous system (CNS) lesions seen in multiple sclerosis (MS). Relevant epitopes, antibodies towards these epitopes and a reliable...... assay are all mandatory parts in detection and evaluation of the pertinence of such cytotoxicity reactions. We have adapted a flow cytometry assay detecting CD107a expression on the surface of cytotoxic effector cells to be applicable for analyses of the effect on target cells from MS patients...

  7. Flow cytometric analysis of lymphocytes and lymphocyte subpopulations in induced sputum from patients with asthma

    Directory of Open Access Journals (Sweden)

    Yutaro Shiota

    2000-01-01

    Full Text Available Study objectives were to compare the numbers of lymphocytes and lymphocyte subpopulations in induced sputum from asthmatic patients and from healthy subjects, and to determine the effect of inhaled anti-asthmatic steroid therapy on these cell numbers. Hypertonic saline inhalation was used to non-invasively induce sputum samples in 34 patients with bronchial asthma and 21 healthy subjects. The sputum samples were reduced with dithioerythritol and absolute numbers of lymphocytes and lymphocyte subpopulations were assessed by direct immunofluorescence and flow cytometry. To assess the effect of beclomethasone dipropionate (BDP on induced sputum, numbers of lymphocytes and lymphocyte subpopulations in sputum also were evaluated after 4 weeks of BDP inhalation treatment in seven asthmatic patients. An adequate sample was obtained in 85.3% of patients with asthma and in 79.2% of the healthy subjects. Induced sputum from patients with asthma had increased numbers of lymphocytes (P = 0.009; CD4+ cells (P = 0.044; CD4+ cells-bearing interleukin-2 receptor (CD25; P = 0.016; and CD4+ cells bearing human histocompatibility leukocyte antigen (HLA-DR (P = 0.033. CD8+ cells were not increased in asthmatic patients. In patients treated with inhaled steroids, numbers of lymphocytes, CD4+ cells, CD25-bearing CD4+ cells and HLA-DR-bearing CD4+ cells in sputum decreased from pretreatment numbers (P = 0.016, 0.002, 0.003 and 0.002, respectively. Analysis of lymphocytes in induced sputum by flow cytometry is useful in assessing bronchial inflammation, and activated CD4+ lymphocytes may play a key role in the pathogenesis of airway inflammation in bronchial asthma.

  8. Assessment of a five-color flow cytometric assay for verifying automated white blood cell differentials

    Institute of Scientific and Technical Information of China (English)

    HUANG Chun-mei; YU Lian-hui; PU Cheng-wei; WANG Xin; WANG Geng; SHEN Li-song; WANG Jian-zhong

    2013-01-01

    Background White blood cell (WBC) counts and differentials performed using an automated cell counter typically require manual microscopic review.However,this last step is time consuming and requires experienced personnel.We evaluated the clinical efficiency of using flow cytometry (FCM) employing a six-antibody/five-color reagent for verifying automated WBC differentials.Methods A total of 56 apparently healthy samples were assessed using a five-color flow cytometer to verify the normal reference ranges of WBC differentials.WBC differentials of 622 samples were also determined using both a cell counter and FCM.These results were then confirmed using manual microscopic methods.Results The probabilities for all of the parameters of WBC differentials exceeded the corresponding normal reference ranges by no more than 7.5%.The resulting WBC differentials were well correlated between FCM and the cell counter (r >0.88,P <0.001),except in the case of basophils.Neutrophils,lymphocytes,and eosinophils were well correlated between FCM and standard microscopic cytology assessment (r >0.80,P <0.001).The sensitivities of FCM for identification of immature granulocytes and blast cells (72.03% and 22.22%,respectively) were higher than those of the cell counter method (44.92% and 11.11%,respectively).The specificities of FCM were all above 85%,substantially better than those of the cell counter method.Conclusion These five-color FCM assays could be applied to accurately verify abnormal results of automated assessment of WBC differentials.

  9. Flow cytometric assessment of microbial abundance in the near-field area of seawater reverse osmosis concentrate discharge

    KAUST Repository

    Van Der Merwe, Riaan

    2014-06-01

    The discharge of concentrate and other process waters from seawater reverse osmosis (SWRO) plant operations into the marine environment may adversely affect water quality in the near-field area surrounding the outfall. The main concerns are the increase in salt concentration in receiving waters, which results in a density increase and potential water stratification near the outfall, and possible increases in turbidity, e.g., due to the discharge of filter backwash waters. Changes in ambient water quality may affect microbial abundance in the area, for example by hindering the photosynthesis process or disrupting biogenesis. It is widely accepted that marine biodiversity is lower in more extreme conditions, such as high salinity environments. As aquatic microbial communities respond very rapidly to changes in their environment, they can be used as indicators for monitoring ambient water quality. The objective of this study was to assess possible changes in microbial abundance as a result of concentrate discharge into the near-field area (<. 25. m) surrounding the outfall of the King Abdullah University of Science and Technology (KAUST) SWRO plant. Flow cytometric (FCM) analysis was conducted in order to rapidly determine microbial abundance on a single-cell level in 107 samples, taken by diving, from the discharge area, the intake area and two control sites. FCM analysis combined the measurement of distinct scatter of cells and particles, autofluorescence of cyanobacteria and algae, and fluorescence after staining of nucleic acids with SYBR® Green for a total bacterial count. The results indicate that changes in microbial abundance in the near-field area of the KAUST SWRO outfall are minor and appear to be the result of a dilution effect rather than a direct impact of the concentrate discharge. © 2014 Elsevier B.V.

  10. Histopathologic and Flow-Cytometric Analysis of Neoplastic and Benign “background” Tissue in Breast Carcinoma Resections

    Directory of Open Access Journals (Sweden)

    Daniel W. Visscher

    1998-01-01

    Full Text Available Two-color, multiparametric synthesis phase fraction (SPF analysis of cytokeratin-labeled epithelial cells was flow cytometrically performed on both benign (SPFb and malignant tissue samples (if available, SPFt from 132 mastectomy/lumpectomy specimens. These data were then correlated with clinicopathologic features, including (1 tumor differentiation, (2 the proportion of tumor comprised of duct carcinoma-in situ (DCIS, and (3 the histology of accompanying benign breast tissue, classified by predominant microscopic pattern as intact, normal terminal duct lobular units (NTDLU, 34% of cases, atrophic (AT, 33% of cases, proliferative fibrocystic (PFC, 26% of cases, and non-proliferative fibrocystic (NPFC, 7% of cases. SPFt was inversely correlated with extent of DCIS (DCIS =0 – 20% tumor volume – 12.7% mean SPFt, vs. DCIS >20% tumor volume – 6.4% mean SPFt, p = 0.001. SPFt also correlated with the histology of background benign breast tissue (NTDLU – 14.8% mean SPFt vs. AT – 6.9% mean SPFt vs. PFC – 12.7% mean SPFt, p = 0.05 but it did not correlate with patient age or SPFb (overall mean =0.73%. SPFb was correlated with patient age (>56 yr – 0.59% mean SPFb vs. < yr – 0.84% mean SPFb, p = 0.02, with background histology (NTDLU – 1.1% mean SPFb vs. AT – 0.43% mean SPFb vs. PFC – 0.70% mean SPFb, p < 0.02 and with the grade of the neoplasm (well/moderate – 0.58% mean vs. poorly differentiated – 0.85% mean, p = 0.04. Patients having a background of PFC were significantly older than patients with a background of NTDLU (45.2 yr vs. 60.2 yr, p = 0.01.

  11. Performance of the flow cytometric E-screen assay in screening estrogenicity of pure compounds and environmental samples

    Energy Technology Data Exchange (ETDEWEB)

    Vanparys, Caroline, E-mail: caroline.vanparys@ua.ac.be [Laboratory of Ecophysiology, Biochemistry and Toxicology, University of Antwerp, Antwerp (Belgium); Depiereux, Sophie; Nadzialek, Stephanie [Research Unit in Organismal Biology (URBO), University of Namur (FUNDP), Namur (Belgium); Robbens, Johan; Blust, Ronny [Laboratory of Ecophysiology, Biochemistry and Toxicology, University of Antwerp, Antwerp (Belgium); Kestemont, Patrick [Research Unit in Organismal Biology (URBO), University of Namur (FUNDP), Namur (Belgium); De Coen, Wim [Laboratory of Ecophysiology, Biochemistry and Toxicology, University of Antwerp, Antwerp (Belgium); European Chemicals Agency (ECHA), Helsinki (Finland)

    2010-09-15

    In vitro estrogenicity screens are believed to provide a first prioritization step in hazard characterization of endocrine disrupting chemicals. When applied to complex environmental matrices or mixture samples, they have been indicated valuable in estimating the overall estrogen-mimicking load. In this study, the performance of an adapted format of the classical E-screen or MCF-7 cell proliferation assay was profoundly evaluated to rank pure compounds as well as influents and effluents of sewage treatment plants (STPs) according to estrogenic activity. In this adapted format, flow cytometric cell cycle analysis was used to allow evaluation of the MCF-7 cell proliferative effects after only 24 h of exposure. With an average EC{sub 50} value of 2 pM and CV of 22%, this assay appears as a sensitive and reproducible system for evaluation of estrogenic activity. Moreover, estrogenic responses of 17 pure compounds corresponded well, qualitatively and quantitatively, with other in vitro and in vivo estrogenicity screens, such as the classical E-screen (R{sup 2} = 0.98), the estrogen receptor (ER) binding (R{sup 2} = 0.84) and the ER transcription activation assay (R{sup 2} = 0.87). To evaluate the applicability of this assay for complex samples, influents and effluents of 10 STPs covering different treatment processes, were compared and ranked according to estrogenic removal efficiencies. Activated sludge treatment with phosphorus and nitrogen removal appeared most effective in eliminating estrogenic activity, followed by activated sludge, lagoon and filter bed. This is well in agreement with previous findings based on chemical analysis or biological activity screens. Moreover, ER blocking experiments indicated that cell proliferative responses were mainly ER mediated, illustrating that the complexity of the end point, cell proliferation, compared to other ER screens, does not hamper the interpretation of the results. Therefore, this study, among other E-screen studies

  12. Evaluation of Prognostic Factors Following Flow-Cytometric DNA Analysis after Cytokeratin Labelling: II. Cervical and Endometrial Cancer

    Directory of Open Access Journals (Sweden)

    Pauline Wimberger

    2002-01-01

    Full Text Available In gynecologic oncology valid prognostic factors are necessary to define biologically similar subgroups for analysis of therapeutic efficacy. This study is the first published prospective study concerning prognostic significance of DNA ploidy and S‐phase fraction in cervical and endometrial cancer following enrichment of tumor cells by cytokeratin labelling. Epithelial cells were labeled by FITC‐conjugated cytokeratin antibody (CK 5, 6, 8, and CK 17 prior to flow cytometric cell cycle analysis in 91 specimens of cervical cancer and 73 samples of endometrial cancer. In cervical cancer neither DNA‐ploidy nor S‐phase fraction were relevant prognostic parameters. But CV of the G0G1‐peak showed prognostic relevance in cervical cancer cells, even in multivariate analysis. This interesting observation, however, seems to have no therapeutic consequence due to the small discrimination capacity of CV. In endometrial carcinoma, gross DNA‐aneuploidy (DNA‐index > 1.3 and a high percentage of proliferating cells (>75th percentile were univariate and multivariate highly significant prognostic factors for recurrence‐free survival. Especially DNA‐aneuploidy (DI>1.3 is one of the most important independent molecular biological prognostic factors. While diagnostic curettage we could identify risk patients even preoperatively by determination of the prognostic factors like histologic tumor type, grading, cervical involvement and DNA‐ploidy. Thereby these patients could be treated primarily in an oncologic center. In conclusion, our investigations showed that the determination of DNA‐ploidy should be done in endometrial carcinoma. In cervical cancer no clinical significance for determination of DNA‐parameters was found.

  13. Antibody affinity maturation through combining display of two-chain paired antibody and precision flow cytometric sorting.

    Science.gov (United States)

    Sun, Shuang; Yang, Xiao; Wang, Haifeng; Zhao, Yun; Lin, Yan; Ye, Chen; Fang, Xiangdong; Hang, Haiying

    2016-07-01

    Recombination of antibody light and heavy chain libraries greatly increases the size of a two-chain paired antibody library, thus easing the construction of large antibody libraries. Here, light and heavy chain variable domains paired by a coiled coil were applied to a bacterial inner membrane display system. However, the probability of the correct pairing of light and heavy chains through random recombination after each round of flow cytometric sorting and cloning was very low in the presence of mostly unmatched light and heavy chain genes, resulting in inefficient enrichment; a target antibody clone in the ratio of 1:100,000 negative control spheroplasts was unable to be enriched by six rounds of sorting and cloning by a conventional sorting strategy (sorting the top 1 %). By just sorting the top 0.000025 % of spheroplasts, we succeeded in enriching the target antibody clone mixed with negative control spheroplasts in a ratio of 1:10(8) by just one round of sorting and cloning. Furthermore, using this gating strategy, we efficiently enriched for an antibody clone with an affinity slightly better than the parent antibody clone from mixed spheroplasts which were present in the ratio of 1 better affinity clone to 10 parent clones to 10(6) negative control clones after just two rounds of sorting and cloning, suggesting that this gating strategy is highly sensitive in distinguishing between clones with a small difference in affinity and also enriching for clones with a higher affinity. Taken together, the combination of the display of a two-chain paired antibody library and the use of stringent gating has significantly increased the efficiency of the antibody maturation system.

  14. Relevance of Flow Cytometric Auto-Crossmatch to the Post-transplant Course of Kidney Transplant Recipients.

    Science.gov (United States)

    Demir, E; Yeğit, O; Erol, A; Akgül, S U; Çalışkan, B; Bayraktar, A; Çalışkan, Y; Türkmen, A; Savran, F O; Sever, M S

    2017-04-01

    The crossmatch test is essential prior to kidney transplantation (tx) to confirm compatibility between the donor and the recipient. However, its results can be misleading due to "undetectable antibodies" in the recipient's serum. To establish if undetectable autoantibodies are responsible for a positive result, an auto-crossmatch test can be performed. In this study, we aim to determine the long-term prognostic value of auto-flow cytometric auto-crossmatch (FCXM) test on kidney survival in kidney tx recipients. The primary outcome variable was reduced renal function. Secondary endpoints were incidence of biopsy-confirmed chronic antibody-mediated rejection (CAMR) and recurrent glomerulonephritis (GN). There were no differences regarding initial serum creatinine levels between the study and control groups (P = .441). Patients who had positive auto-B FCXM had a significantly reduced renal function compared with the control group (P = .016). Four patients developed biopsy-confirmed CAMR in the study group and 1 patient in the control group (P = .047). Five patients had biopsy-confirmed recurrent GN in the GN study group, and only 1 patient had recurrent GN in the GN control group (P = .026). Kidney transplant recipients with positive auto-FCXM test had significantly reduced renal function and a higher incidence of recurrent GN and CAMR compared with the control group. The findings of this study suggest a potential role of auto-antibody causing positive auto-FCXM test result, meanwhile increasing the risk of CAMR, recurrent GN, and new-onset diabetes after tx. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis.

    Science.gov (United States)

    Koopman, G; Reutelingsperger, C P; Kuijten, G A; Keehnen, R M; Pals, S T; van Oers, M H

    1994-09-01

    Apoptosis, or programmed cell death, is a general mechanism for removal of unwanted cells from the immune system. It is characterized by chromatin condensation, a reduction in cell volume, and endonuclease cleavage of DNA into oligonucleosomal length fragments. Apoptosis is also accompanied by a loss of membrane phospholipid asymmetry, resulting in the exposure of phosphatidylserine at the surface of the cell. Expression of phosphatidylserine at the cell surface plays an important role in the recognition and removal of apoptotic cells by macrophages. Here we describe a new method for the detection of apoptotic cells by flow cytometry, using the binding of fluorescein isothiocyanate-labeled annexin V to phosphatidylserine. When Burkitt lymphoma cell lines and freshly isolated germinal center B cells are cultured under apoptosis inducing conditions, all cells showing chromatin condensation strongly stain with annexin V, whereas normal cells are annexin V negative. Moreover, DNA fragmentation is only found in the annexin V-positive cells. The nonvital dye ethidium bromide was found to stain a subpopulation of the annexin V-positive apoptotic cells, increasing with time. Our results indicate that the phase in apoptosis that is characterized by chromatin condensation coincides with phosphatidylserine exposure. Importantly, it precedes membrane damage that might lead to release from the cells of enzymes that are harmful to the surrounding tissues. Annexin V may prove important in further unravelling the regulation of apoptosis.

  16. Flow cytometric bacterial cell counts challenge conventional heterotrophic plate counts for routine microbiological drinking water monitoring.

    Science.gov (United States)

    Van Nevel, S; Koetzsch, S; Proctor, C R; Besmer, M D; Prest, E I; Vrouwenvelder, J S; Knezev, A; Boon, N; Hammes, F

    2017-04-15

    Drinking water utilities and researchers continue to rely on the century-old heterotrophic plate counts (HPC) method for routine assessment of general microbiological water quality. Bacterial cell counting with flow cytometry (FCM) is one of a number of alternative methods that challenge this status quo and provide an opportunity for improved water quality monitoring. After more than a decade of application in drinking water research, FCM methodology is optimised and established for routine application, supported by a considerable amount of data from multiple full-scale studies. Bacterial cell concentrations obtained by FCM enable quantification of the entire bacterial community instead of the minute fraction of cultivable bacteria detected with HPC (typically water samples per day, depending on the laboratory and selected staining procedure(s). Moreover, many studies have shown FCM total (TCC) and intact (ICC) cell concentrations to be reliable and robust process variables, responsive to changes in the bacterial abundance and relevant for characterising and monitoring drinking water treatment and distribution systems. The purpose of this critical review is to initiate a constructive discussion on whether FCM could replace HPC in routine water quality monitoring. We argue that FCM provides a faster, more descriptive and more representative quantification of bacterial abundance in drinking water.

  17. Improved sensitivity in flow cytometric intracellular ionized calcium measurement using fluo-3/Fura Red fluorescence ratios.

    Science.gov (United States)

    Novak, E J; Rabinovitch, P S

    1994-10-01

    Measurement of changes in intracellular ionized calcium concentrations ([Ca2+]i) has proved to be of wide use in the study of cellular responses to activating stimuli. The fluorescent dye Indo-1 has successfully been used in flow cytometry for this purpose, and when used as a ratiometric indicator it provides optimum sensitivity and accuracy. Unfortunately, this dye requires ultraviolet (UV) excitation which is often not available. We show here that similar results can be obtained using a ratio of green to red fluorescence from the simultaneous loading of the dyes Fura Red and fluo-3. Both Fura Red and fluo-3 are excited using the commonly available blue 488 nm laser line. With appropriate concentrations of the two dyes, the magnitude of response with the fluo-3/Fura Red ratio is greater than that achieved with indo-1, while the intercellular variation in measurement is similar to that seen with indo-1. Analyses can be simultaneously combined with immunofluorescent detection of PE-labeled antibodies to enable [Ca2+]i measurement within cell subsets.

  18. Neuropathological similarities and differences between schizophrenia and bipolar disorder: a flow cytometric postmortem brain study.

    Directory of Open Access Journals (Sweden)

    Yoshitaka Hayashi

    Full Text Available Recent studies suggest that schizophrenia (SCH and bipolar disorder (BPD may share a similar etiopathology. However, their precise neuropathological natures have rarely been characterized in a comprehensive and quantitative fashion. We have recently developed a rapid, quantitative cell-counting method for frozen unfixed postmortem brains using a flow cytometer. In the present study, we not only counted stained nuclei, but also measured their sizes in the gray matter of frontopolar cortices (FPCs and inferior temporal cortices (ITCs from patients with SCH or BPD, as well as in that from normal controls. In terms of NeuN(+ neuronal nuclei size, particularly in the reduced densities of small NeuN(+ nuclei, we found abnormal distributions present in the ITC gray matter of both patient groups. These same abnormalities were also found in the FPCs of SCH patients, whereas in the FPCs of BPD patients, a reduction in oligodendrocyte lineage (olig2(+ cells was much more common. Surprisingly, in the SCH FPC, normal left-greater-than-right asymmetry in neural nuclei densities was almost completely reversed. In the BPD FPC, this asymmetry, though not obvious, differed significantly from that in the SCH FPC. These findings indicate that while similar neuropathological abnormalities are shared by patients with SCH or BPD, differences also exist, mainly in the FPC, which may at least partially explain the differences observed in many aspects in these disorders.

  19. Evaluation of chromatin condensation in human spermatozoa: a flow cytometric assay using acridine orange staining.

    Science.gov (United States)

    Golan, R; Shochat, L; Weissenberg, R; Soffer, Y; Marcus, Z; Oschry, Y; Lewin, L M

    1997-01-01

    The quality of sperm chromatin is an important factor in fertilization and is especially critical where one spermatozoon is artificially selected for fertilizing an egg (as in intracytoplasmic sperm injection). In this study, flow cytometry after staining of human spermatozoa with Acridine Orange was used to study chromatin structure. A method is described for estimating the percentage of cells in a human sperm sample that have completed epididymal maturation in regard to chromatin condensation. Of the 121 samples of the semen that were examined, nine contained a higher percentage of hypocondensed spermatozoa and six samples contained elevated amounts of hypercondensed spermatozoa. In addition to aberrancies in chromatin condensation other defects showed up as satellite populations of spermatozoa with higher than normal ratios of red/green fluorescence after Acridine Orange staining. Such defects were found in 15 semen samples. The use of swim-up and Percoll gradient centrifugation methods was shown to improve the percentage of spermatozoa with normal chromatin structure in some samples with poor initial quality.

  20. Establishing the flow cytometric assessment of myeloid cells in kidney ischemia/reperfusion injury.

    Science.gov (United States)

    Williams, Timothy M; Wise, Andrea F; Alikhan, Maliha A; Layton, Daniel S; Ricardo, Sharon D

    2014-03-01

    Polychromatic flow cytometry is a powerful tool for assessing populations of cells in the kidney through times of homeostasis, disease and tissue remodeling. In particular, macrophages have been identified as having central roles in these three settings. However, because of the plasticity of myeloid cells it has been difficult to define a specific immunophenotype for these cells in the kidney. This study developed a gating strategy for identifying and assessing monocyte and macrophage subpopulations, along with neutrophils and epithelial cells in the healthy kidney and following ischemia/reperfusion (IR) injury in mice, using antibodies against CD45, CD11b, CD11c, Ly6C, Ly6G, F4/80, CSF-1R (CD115), MHC class II, mannose receptor (MR or CD206), an alternatively activated macrophage marker, and the epithelial cell adhesion marker (EpCAM or CD326). Backgating analysis and assessment of autofluorescence was used to extend the knowledge of various cell types and the changes that occur in the kidney at various time-points post-IR injury. In addition, the impact of enzymatic digestion of kidneys on cell surface markers and cell viability was assessed. Comparisons of kidney myeloid populations were also made with those in the spleen. These results provide a useful reference for future analyses of therapies aimed at modulating inflammation and enhancing endogenous remodeling following kidney injury.

  1. Development of a novel flow cytometric approach to evaluate fish sperm chromatin using fixed samples

    Science.gov (United States)

    Jenkins, Jill A.

    2013-01-01

    The integrity of the paternal DNA is essential for the accurate transmission of genetic information, yet fertilization is not inhibited by chromatin breakage. Some methods are available for the sensitive detection of DNA damage and can be applied in studies of environmental toxicology, carcinogenesis, aging, and assisted reproduction techniques in both clinical and experimental settings. Because semen samples obtained from remote locations undergo chromatin damage prior to laboratory assessment, the present study was undertaken to evaluate treatments for effective chromatin staining in the development of a DNA fragmentation assay using fixed milt from yellow perch (Perca flavescens). Similar to the sperm chromatin structure assay (SCSA), susceptibility of nuclear DNA to acid-induced denaturation was measured by flow cytometry (FCM). Use of 10% buffered formalin for milt fixation allowed easier peak discrimination than 4% paraformaldehyde. The effects of time and temperature of incubation in 0.08 N HCl were evaluated in order to determine the ideal conditions for promoting DNA decondensation and making strand breaks more available for staining and detection by FCM. The best results were obtained with incubation at 37°C for 1 minute, followed by cold propidium iodide staining for 30 minutes.

  2. Flow cytometric functional analysis of multidrug resistance by Fluo-3: a comparison with rhodamine-123.

    Science.gov (United States)

    Koizumi, S; Konishi, M; Ichihara, T; Wada, H; Matsukawa, H; Goi, K; Mizutani, S

    1995-09-01

    Using four cell lines including drug-sensitive K562/Parent cells, P-glycoprotein (Pgp)-mediated multidrug resistant (MDR) K562/VCR, K562/ADR and revertant K562/ADR-R cells, two fluorescent agents, Fluo-3 and rhodamine-123 (Rh-123), were compared as indicators in a functional assay of MDR. Cells were incubated with 4 microM Fluo-3 or 1 microM Rh-123 for 45 min and then the intracellular accumulation of the agent was measured using a flow cytometer. Verapamil (20 microM) or cepharanthine (biscoclaurine alkaloid, 10 microM) was added just before the fluorescent agents. Efflux patterns were also studied 60 min after incubation with or without verapamil and cepharanthine. Increased intracellular accumulation and a delayed efflux pattern of Fluo-3 by verapamil and cepharanthine were demonstrated in multidrug resistant K562/VCR and K562/ADR cells, indicating that Fluo-3 is another good indicator of MDR. However, a similar, but lower, increase in uptake and a delayed efflux pattern of Fluo-3 by verapamil and cepharanthine were also demonstrated even in Pgp-non-overexpressed K562/Parent cells. In contrast, accumulation of Rh-123 was not affected by verapamil and cepharanthine. To further study the Pgp dependency of Fluo-3, another cell line, K562/NC16 expressing minimum MDR1 mRNA, was cloned. Increased uptake and a delayed efflux pattern of Fluo-3, but not Rh-123, with verapamil or cepharanthine were again demonstrated in K562/NC16 cells, indicating that intracellular accumulation of Fluo-3 may be non-specifically influenced by verapamil and cepharanthine at very low levels of Pgp-related MDR, while the influx and efflux patterns of Rh-123 may be specifically affected by Pgp overexpression.

  3. Monitoring microbiological changes in drinking water systems using a fast and reproducible flow cytometric method

    KAUST Repository

    Prest, Emmanuelle I E C

    2013-12-01

    Flow cytometry (FCM) is a rapid, cultivation-independent tool to assess and evaluate bacteriological quality and biological stability of water. Here we demonstrate that a stringent, reproducible staining protocol combined with fixed FCM operational and gating settings is essential for reliable quantification of bacteria and detection of changes in aquatic bacterial communities. Triplicate measurements of diverse water samples with this protocol typically showed relative standard deviation values and 95% confidence interval values below 2.5% on all the main FCM parameters. We propose a straightforward and instrument-independent method for the characterization of water samples based on the combination of bacterial cell concentration and fluorescence distribution. Analysis of the fluorescence distribution (or so-called fluorescence fingerprint) was accomplished firstly through a direct comparison of the raw FCM data and subsequently simplified by quantifying the percentage of large and brightly fluorescent high nucleic acid (HNA) content bacteria in each sample. Our approach enables fast differentiation of dissimilar bacterial communities (less than 15min from sampling to final result), and allows accurate detection of even small changes in aquatic environments (detection above 3% change). Demonstrative studies on (a) indigenous bacterial growth in water, (b) contamination of drinking water with wastewater, (c) household drinking water stagnation and (d) mixing of two drinking water types, univocally showed that this FCM approach enables detection and quantification of relevant bacterial water quality changes with high sensitivity. This approach has the potential to be used as a new tool for application in the drinking water field, e.g. for rapid screening of the microbial water quality and stability during water treatment and distribution in networks and premise plumbing. © 2013 Elsevier Ltd.

  4. Flow Cytometric Analysis of Leishmania Reactive CD4+/CD8+ Lymphocyte Proliferation in Cutaneous Leishmaniasis

    Directory of Open Access Journals (Sweden)

    H Keshavarz

    2008-12-01

    Full Text Available Background: Determination of the division history of T cells in vitro is helpful in the study of effector mechanisms against infections. Technique described here uses the intracellular fluorescent label carboxyfluorescein diacetate succinimidyl ester (CFSE to monitor the proliferation. Methods: In a cross sectional study, blood samples were collected from 7 volunteers with history of cutaneous leishmania­sis (CL and one healthy control from endemic areas in Isfahan province who referred to the Center for Research and Training in Skin Diseases and Leprosy (CRTSDL, then CD4+/CD8+ lymphocytes and CD14+ monocytes were isolated from peri­pheral blood mononuclear cells (PBMC using mAbs and magnetic nanoparticles. CFSE labeled CD4+ or CD8+ lympho­cytes cultured with autologous monocytes in the presence of PHA, SLA, live Leishmania major or as control with­out sti­mulation. Cells were harvested after 7 days and were analyzed using flow cytometry. Results: Five consecutive divisions were monitored separately. Stimulation of CD4+ or CD8+ lymphocytes from CL sub­jects with SLA showed a significant difference in proliferation comparing with unstimulated cells (P< 0.05. The signifi­cant difference in the percentages of CD4+ cells stimulated with SLA was revealed at different divisions for each subject. In CD8+ lymphocyte, significant stronger stimulation of SLA was evident later in the proliferation process. The mean number of divisions in both CD4+/CD8+ lymphocytes stimulated with SLA was significantly greater than when stimulated with live L. major (P=0.007 / P=0.012, respectively Conclusion: The percentage of divided cells might be calculated separately in each division. The cells remained active following CFSE staining and there is possibility of functional analysis simultaneously.

  5. Monitoring microbiological changes in drinking water systems using a fast and reproducible flow cytometric method.

    Science.gov (United States)

    Prest, E I; Hammes, F; Kötzsch, S; van Loosdrecht, M C M; Vrouwenvelder, J S

    2013-12-01

    Flow cytometry (FCM) is a rapid, cultivation-independent tool to assess and evaluate bacteriological quality and biological stability of water. Here we demonstrate that a stringent, reproducible staining protocol combined with fixed FCM operational and gating settings is essential for reliable quantification of bacteria and detection of changes in aquatic bacterial communities. Triplicate measurements of diverse water samples with this protocol typically showed relative standard deviation values and 95% confidence interval values below 2.5% on all the main FCM parameters. We propose a straightforward and instrument-independent method for the characterization of water samples based on the combination of bacterial cell concentration and fluorescence distribution. Analysis of the fluorescence distribution (or so-called fluorescence fingerprint) was accomplished firstly through a direct comparison of the raw FCM data and subsequently simplified by quantifying the percentage of large and brightly fluorescent high nucleic acid (HNA) content bacteria in each sample. Our approach enables fast differentiation of dissimilar bacterial communities (less than 15 min from sampling to final result), and allows accurate detection of even small changes in aquatic environments (detection above 3% change). Demonstrative studies on (a) indigenous bacterial growth in water, (b) contamination of drinking water with wastewater, (c) household drinking water stagnation and (d) mixing of two drinking water types, univocally showed that this FCM approach enables detection and quantification of relevant bacterial water quality changes with high sensitivity. This approach has the potential to be used as a new tool for application in the drinking water field, e.g. for rapid screening of the microbial water quality and stability during water treatment and distribution in networks and premise plumbing. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. A numerical analysis model for interpretation of flow cytometric studies of ex vivo phagocytosis.

    Directory of Open Access Journals (Sweden)

    Ted S Strom

    Full Text Available The study of ex vivo phagocytosis via flow cytometry requires that one distinguish experimentally between uptake and adsorption of fluorescently labeled targets by phagocytes. Removal of the latter quantity from the analysis is the most common means of analyzing such data. Because the probability of phagocytosis is a function of the probability of adsorption, and because partially quenched fluorescence after uptake often overlaps with that of negative controls, this approach is suboptimal at best. Here, we describe a numerical analysis model which overcomes these limitations. We posit that the random adsorption of targets to macrophages, and subsequent phagocytosis, is a function of three parameters: the ratio of targets to macrophages (m, the mean fluorescence intensity imparted to the phagocyte by the internalized target (alpha, and the probability of phagocytosis per adsorbed target (p. The potential values of these parameters define a parameter space and their values at any point in parameter space can be used to predict the fraction of adsorption(+ and [adsorption(-, phagocytosis(+] cells that might be observed experimentally. By systematically evaluating the points in parameter space for the latter two values and comparing them to experimental data, the model arrives at sets of parameter values that optimally predict such data. Using activated THP-1 cells as macrophages and platelets as targets, we validate the model by demonstrating that it can distinguish between the effects of experimental changes in m, alpha, and p. Finally, we use the model to demonstrate that platelets from a congenitally thrombocytopenic WAS patient show an increased probability of ex vivo phagocytosis. This finding correlates with other evidence that rapid in vivo platelet consumption contributes significantly to the thrombocytopenia of WAS. Our numerical analysis method represents a useful and innovative approach to multivariate analysis.

  7. Flow cytometric bacterial cell counts challenge conventional heterotrophic plate counts for routine microbiological drinking water monitoring

    KAUST Repository

    Van Nevel, S.

    2017-02-08

    Drinking water utilities and researchers continue to rely on the century-old heterotrophic plate counts (HPC) method for routine assessment of general microbiological water quality. Bacterial cell counting with flow cytometry (FCM) is one of a number of alternative methods that challenge this status quo and provide an opportunity for improved water quality monitoring. After more than a decade of application in drinking water research, FCM methodology is optimised and established for routine application, supported by a considerable amount of data from multiple full-scale studies. Bacterial cell concentrations obtained by FCM enable quantification of the entire bacterial community instead of the minute fraction of cultivable bacteria detected with HPC (typically < 1% of all bacteria). FCM measurements are reproducible with relative standard deviations below 3% and can be available within 15 min of samples arriving in the laboratory. High throughput sample processing and complete automation are feasible and FCM analysis is arguably less expensive than HPC when measuring more than 15 water samples per day, depending on the laboratory and selected staining procedure(s). Moreover, many studies have shown FCM total (TCC) and intact (ICC) cell concentrations to be reliable and robust process variables, responsive to changes in the bacterial abundance and relevant for characterising and monitoring drinking water treatment and distribution systems. The purpose of this critical review is to initiate a constructive discussion on whether FCM could replace HPC in routine water quality monitoring. We argue that FCM provides a faster, more descriptive and more representative quantification of bacterial abundance in drinking water.

  8. Flow cytometric sorting of fecal bacteria after in situ hybridization with polynucleotide probes.

    Science.gov (United States)

    Bruder, Lena M; Dörkes, Marcel; Fuchs, Bernhard M; Ludwig, Wolfgang; Liebl, Wolfgang

    2016-10-01

    The gut microbiome represents a key contributor to human physiology, metabolism, immune function, and nutrition. Elucidating the composition and genetics of the gut microbiota under various conditions is essential to understand how microbes function individually and as a community. Metagenomic analyses are increasingly used to study intestinal microbiota. However, for certain scientific questions it is sufficient to examine taxon-specific submetagenomes, covering selected bacterial genera in a targeted manner. Here we established a new variant of fluorescence in situ hybridization (FISH) combined with fluorescence-activated cell sorting (FACS), providing access to the genomes of specific taxa belonging to the complex community of the intestinal microbiota. In contrast to standard oligonucleotide probes, the RNA polynucleotide probe used here, which targets domain III of the 23S rRNA gene, extends the resolution power in environmental samples by increasing signal intensity. Furthermore, cells hybridized with the polynucleotide probe are not subjected to harsh pretreatments, and their genetic information remains intact. The protocol described here was tested on genus-specifically labeled cells in various samples, including complex fecal samples from different laboratory mouse types that harbor diverse intestinal microbiota. Specifically, as an example for the protocol described here, RNA polynucleotide probes could be used to label Enterococcus cells for subsequent sorting by flow cytometry. To detect and quantify enterococci in fecal samples prior to enrichment, taxon-specific PCR and qPCR detection systems have been developed. The accessibility of the genomes from taxon-specifically sorted cells for subsequent molecular analyses was demonstrated by amplification of functional genes. Copyright © 2016 Elsevier GmbH. All rights reserved.

  9. Flow-cytometric study of vital cellular functions in Escherichia coli during solar disinfection (SODIS).

    Science.gov (United States)

    Berney, Michael; Weilenmann, Hans-Ulrich; Egli, Thomas

    2006-06-01

    The effectiveness of solar disinfection (SODIS), a low-cost household water treatment method for developing countries, was investigated with flow cytometry and viability stains for the enteric bacterium Escherichia coli. A better understanding of the process of injury or death of E. coli during SODIS could be gained by investigating six different cellular functions, namely: efflux pump activity (Syto 9 plus ethidium bromide), membrane potential [bis-(1,3-dibutylbarbituric acid)trimethine oxonol; DiBAC4(3)], membrane integrity (LIVE/DEAD BacLight), glucose uptake activity (2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose; 2-NBDG), total ATP concentration (BacTiter-Glo) and culturability (pour-plate method). These variables were measured in E. coli K-12 MG1655 cells that were exposed to either sunlight or artificial UVA light. The inactivation pattern of cellular functions was very similar for both light sources. A UVA light dose (fluence) of 80 % of the cells was observed at a fluence of approximately 1500 kJ m(-2), and the cytoplasmic membrane of bacterial cells became permeable at a fluence of >2500 kJ m(-2). Culturable counts of stressed bacteria after anaerobic incubation on sodium pyruvate-supplemented tryptic soy agar closely correlated with the loss of membrane potential. The results strongly suggest that cells exposed to >1500 kJ m(-2) solar UVA (corresponding to 530 W m(-2) global sunlight intensity for 6 h) were no longer able to repair the damage and recover. Our study confirms the lethal effect of SODIS with cultivation-independent methods and gives a detailed picture of the 'agony' of E. coli when it is stressed with sunlight.

  10. Color encoded microbeads-based flow cytometric immunoassay for polycyclic aromatic hydrocarbons in food

    Energy Technology Data Exchange (ETDEWEB)

    Meimaridou, Anastasia, E-mail: anastasia.meimaridou@wur.nl [RIKILT-Institute of Food Safety, Wageningen UR, P.O. Box 230, 6700 AE Wageningen (Netherlands); Haasnoot, Willem; Noteboom, Linda; Mintzas, Dimitrios [RIKILT-Institute of Food Safety, Wageningen UR, P.O. Box 230, 6700 AE Wageningen (Netherlands); Pulkrabova, Jana; Hajslova, Jana [Department of Food Chemistry and Analysis, Institute of Chemical Technology Prague, Technicka 3, 166 28 Prague 6 (Czech Republic); Nielen, Michel W.F. [RIKILT-Institute of Food Safety, Wageningen UR, P.O. Box 230, 6700 AE Wageningen (Netherlands); Wageningen University, Laboratory of Organic Chemistry, Dreijenplein 8, 6703 HB Wageningen (Netherlands)

    2010-07-05

    Food contamination caused by chemical hazards such as persistent organic pollutants (POPs) is a worldwide public health concern and requires continuous monitoring. The chromatography-based analysis methods for POPs are accurate and quite sensitive but they are time-consuming, laborious and expensive. Thus, there is a need for validated simplified screening tools, which are inexpensive, rapid, have automation potential and can detect multiple POPs simultaneously. In this study we developed a flow cytometry-based immunoassay (FCIA) using a color-encoded microbeads technology to detect benzo[a]pyrene (BaP) and other polycyclic aromatic hydrocarbons (PAHs) in buffer and food extracts as a starting point for the future development of rapid multiplex assays including other POPs in food, such as polychlorinated biphenyls (PCBs) and polybrominated diphenyl ethers (PBDEs). A highly sensitive assay for BaP was obtained with an IC{sub 50} of 0.3 {mu}g L{sup -1} using a monoclonal antibody (Mab22F12) against BaP, similar to the IC{sub 50} of a previously described enzyme-linked immunosorbent assay (ELISA) using the same Mab. Moreover, the FCIA was 8 times more sensitive for BaP compared to a surface plasmon resonance (SPR)-based biosensor immunoassay (BIA) using the same reagents. The selectivity of the FCIAs was tested, with two Mabs against BaP for 25 other PAHs, including two hydroxyl PAH metabolites. Apart from BaP, the FCIAs can detect PAHs such as indenol[1,2,3-cd]pyrene (IP), benz[a]anthracene (BaA), and chrysene (CHR) which are also appointed by the European Food Safety Authority (EFSA) as suitable indicators of PAH contamination in food. The FCIAs results were in agreement with those obtained with gas chromatography-mass spectrometry (GC-MS) for the detection of PAHs in real food samples of smoked carp and wheat flour and has great potential for the future routine application of this assay in a simplex or multiplex format in combination with simplified extraction

  11. Flow-cytometric determination of genotoxic effects of exposure to petroleum in mink and sea otters

    Science.gov (United States)

    Bickham, J.W.; Mazet, J.A.; Blake, J.; Smolen, M.J.; Lou, Y.; Ballachey, B.E.

    1998-01-01

    Three experiments were conducted to investigate the genotoxic effects of crude oil on mink and sea otters, In the first experiment, the effects on mink of chronic exposure to weathered Prudhoe Bay crude oil were studied, Female mink were fed a diet that included weathered crude oil for a period of 3 weeks prior to mating, during pregnancy and until weaning. Kits were exposed through lactation and by diet after weaning until 4 months of age. Kidney and liver tissues of the kits were examined using flow cytometry (FCM) and it was found that the genome size was increased in kidney samples from the experimental group compared to the control group. This effect was probably due to some type of DNA amplification and it could have been inherited from the exposed mothers or have been a somatic response to oil exposure in the pups, No evidence of clastogenic effects, as measured by the coefficient of variation (CV) of the G(1) peak, was found in kidney or liver tissue. In the second experiment, yearling female mink were exposed either by diet or externally to crude oil or bunker C fuel oil. Evidence for clastogenic damage was found in spleen tissue for the exposure groups, but not in kidney tissue. No evidence of increased genome size was observed. In the third experiment, blood was obtained from wild-caught sea otters in Prince William Sound. The sea otters represented two populations: one from western Prince William Sound that was potentially exposed to oil from the Exxon Valdez oil spill and a reference population from eastern Prince William Sound that did not receive oil from the spill. The spill had occurred 1.5 years prior to obtaining the blood samples. Although the mean CVs did not differ between the populations, the exposed population had a significantly higher variance of CV measurements and five out of 15 animals from the exposed population had CVs higher than the 95% confidence limits of the reference population, It is concluded that FCM is a sensitive indicator

  12. Flow Cytometric Quantification of Peripheral Blood Cell β-Adrenergic Receptor Density and Urinary Endothelial Cell-Derived Microparticles in Pulmonary Arterial Hypertension.

    Directory of Open Access Journals (Sweden)

    Jonathan A Rose

    Full Text Available Pulmonary arterial hypertension (PAH is a heterogeneous disease characterized by severe angiogenic remodeling of the pulmonary artery wall and right ventricular hypertrophy. Thus, there is an increasing need for novel biomarkers to dissect disease heterogeneity, and predict treatment response. Although β-adrenergic receptor (βAR dysfunction is well documented in left heart disease while endothelial cell-derived microparticles (Ec-MPs are established biomarkers of angiogenic remodeling, methods for easy large clinical cohort analysis of these biomarkers are currently absent. Here we describe flow cytometric methods for quantification of βAR density on circulating white blood cells (WBC and Ec-MPs in urine samples that can be used as potential biomarkers of right heart failure in PAH. Biotinylated β-blocker alprenolol was synthesized and validated as a βAR specific probe that was combined with immunophenotyping to quantify βAR density in circulating WBC subsets. Ec-MPs obtained from urine samples were stained for annexin-V and CD144, and analyzed by a micro flow cytometer. Flow cytometric detection of alprenolol showed that βAR density was decreased in most WBC subsets in PAH samples compared to healthy controls. Ec-MPs in urine was increased in PAH compared to controls. Furthermore, there was a direct correlation between Ec-MPs and Tricuspid annular plane systolic excursion (TAPSE in PAH patients. Therefore, flow cytometric quantification of peripheral blood cell βAR density and urinary Ec-MPs may be useful as potential biomarkers of right ventricular function in PAH.

  13. The effects of orange juice clarification on the physiology of Escherichia coli; growth-based and flow cytometric analysis.

    Science.gov (United States)

    Anvarian, Amir H P; Smith, Madeleine P; Overton, Tim W

    2016-02-16

    Orange juice (OJ) is a food product available in various forms which can be processed to a greater or lesser extent. Minimally-processed OJ has a high consumer perception but presents a potential microbiological risk due to acid-tolerant bacteria. Clarification of OJ (such as removal of cloud) is a common processing step in many OJ products. However, many of the antimicrobial components of OJ such as essential oils are present in the cloud fraction. Here, the effect of clarification by filtration on the viability and physiology of Escherichia coli K-12 was tested using total viable count (TVC) and flow cytometric (FCM) analysis. The latter technique was also used to monitor intracellular pH during incubation in OJ. Removal of the OJ cloud fraction was shown to have dramatic effects on bacterial viability and physiology during storage at a range of incubation temperatures. For instance, at 4 °C, a significantly lower number of healthy cells and a significantly higher number of injured cells were observed in 0.22 μm-filtered OJ at 24h post-inoculation, compared to filtered OJ samples containing particles between 0.22 μm and 11 μm in size. Similarly, there was a significant difference between the number of healthy bacteria in the 0.7 μm-filtered OJ and both 0.22 μm-filtered and 1.2 μm-filtered OJ after 24 hour incubation at 22.5 °C. This indicated that OJ cloud between 0.7 μm and 0.22 μm in size might have an adverse effect on the viability of E. coli K-12. Furthermore, FCM allowed the rapid analysis of bacterial physiology without the requirement for growth on agar plates, and revealed the extent of the viable but non-culturable (VBNC) population. For example, at 4 °C, while the FCM viable count did not substantially decrease until 48 h, decreases in TVC were observed between 0 and 48 hour incubation, due to a subset of injured bacteria entering the VBNC state, hence being unable to grow on agar plates. This study highlights the application of FCM in

  14. Flow cytometric 96-well microplate-based in vitro micronucleus assay with human TK6 cells: protocol optimization and transferability assessment.

    Science.gov (United States)

    Bryce, Steven M; Avlasevich, Svetlana L; Bemis, Jeffrey C; Tate, Matthew; Walmsley, Richard M; Saad, Frédéric; Van Dijck, Kris; De Boeck, Marlies; Van Goethem, Freddy; Lukamowicz-Rajska, Magdalena; Elhajouji, Azeddine; Dertinger, Stephen D

    2013-04-01

    An automated approach for scoring in vitro micronuclei (MN) has been described in which flow cytometric analysis is combined with compound exposure, processing, and sampling in a single 96-well plate (Bryce SM et al. [2010]: Mutat Res 703:191-199). The current report describes protocol optimization and an interlaboratory assessment of the assay's transferability and reproducibility. In a training phase, the methodology was refined and collaborating laboratories were qualified by repeatedly testing three compounds. Second, a set of 32 chemicals comprised of reference genotoxicants and presumed non-genotoxicants was tested at each of four sites. TK6 cells were exposed to 10 closely spaced compound concentrations for 1.5- to 2-cell population doublings, and were then stained and lysed for flow cytometric analysis. MN frequencies were determined by evaluating ≥ 5,000 cells per replicate well, and several indices of cytotoxicity were acquired. The prevalence of positive results varied according to the MN-fold increase used to signify a genotoxic result, as well as the endpoint used to define a cytotoxicity limit. By varying these parameters, assay sensitivity and specificity values ranged from 82 to 98%, and 86 to 97%, respectively. In a third phase, one laboratory tested a further six genotoxicants and five non-genotoxic apoptosis inducers. In these experiments assay specificity was markedly improved when top concentration selection was based on two cytotoxicity endpoints-relative survival and quantification of ethidium monoazide-positive events. Collectively, the results indicate that the miniaturized assay is transferable across laboratories. The 96-well format consumes considerably less compound than conventional in vitro MN test methods, and the high information content provided by flow cytometry helps guard against irrelevant positive results arising from overt toxicity.

  15. Development of a flow cytometric bead immunoassay and its assessment as a possible aid to potency evaluation of enterotoxaemia vaccines

    Directory of Open Access Journals (Sweden)

    Angela Buys

    2014-02-01

    Full Text Available Enterotoxaemia, an economically important disease of sheep, goats and calves, is caused by systemic effects of the epsilon toxin produced by the anaerobic bacterium Clostridium perfringens type D. The only practical means of controlling the occurrence of enterotoxaemia is to immunise animals by vaccination. The vaccine is prepared by deriving a toxoid from the bacterial culture filtrate and the potency of the vaccine is tested with the in vivo mouse neutralisation test (MNT. Due to ethical, economic and technical reasons, alternative in vitro assays are needed. In this study an indirect cytometric bead immunoassay (I-CBA was developed for use in vaccine potency testing and the results were compared with those obtained using an indirect enzyme-linked immunosorbent assay (I-ELISA and the MNT. Sera were collected from guinea pigs immunised with three different production batches of enterotoxaemia vaccine and the levels of anti-epsilon toxin antibodies were determined. Although the intra- and inter-assay variability was satisfactory, epsilon antitoxin levels determined by both the I-ELISA and indirect cytometric bead immunoassay (I-CBA tests were higher than those of the MNT assay. In contrast to the MNT, all of the serum samples were identified as having antitoxin levels above the required minimum (not less than 5 U/mL. These results indicate that the respective in vitro tests in their current formats are not yet suitable alternatives to the in vivo MNT. The growing demand for a more humane, cost-effective and efficient method for testing the potency of enterotoxaemia vaccines, however, provides a strong impetus for further optimisation and standardisation of the I-CBA assay but further analytical research is required.

  16. Transmission Electron Microscopic Morphological Study and Flow Cytometric Viability Assessment of Acinetobacter baumannii Susceptible to Musca domestica cecropin

    Directory of Open Access Journals (Sweden)

    Shuiqing Gui

    2014-01-01

    Full Text Available Multidrug-resistant (MDR Acinetobacter baumannii infections are difficult to treat owing to the extremely limited armamentarium. Expectations about antimicrobial peptides' use as new powerful antibacterial agents have been raised on the basis of their unique mechanism of action. Musca domestica cecropin (Mdc, a novel antimicrobial peptide from the larvae of Housefly (Musca domestica, has potently active against Gram-positive and Gram-negative bacteria standard strain. Here we evaluated the antibacterial activity of Mdc against clinical isolates of MDR-A. baumannii and elucidate the related antibacterial mechanisms. The minimal inhibitory concentration (MIC of Mdc was 4 μg/mL. Bactericidal kinetics of Mdc revealed rapid killing of A. baumannii (30 min. Flow cytometry using viability stain demonstrated that Mdc causes A. baumannii membrane permeabilization in a concentration- and time-dependent process, which correlates with the bactericidal action. Moreover, transmission electron microscopic (TEM examination showed that Mdc is capable of disrupting the membrane of bacterial cells, resulting in efflux of essential cytoplasmic components. Overall, Mdc could be a promising antibacterial agent for MDR-A. baumannii infections.

  17. Flow cytometric method for in situ preparation of standard materials of a small defined number of microbial cells with colony-forming potentiality.

    Science.gov (United States)

    Matsuoka, Hideaki; Nakano, Koichiro; Takatani, Norimasa; Yoshida, Tomonori; Igimi, Shizunobu; Saito, Mikako

    2014-01-01

    Standard materials of a small defined number of cells with colony-forming potentiality are essential for the rational validation of food microbiological methods. An in situ flow cytometric method using viable staining with 6-carboxyfluorescein diacetate (CFDA) and tryptic soy agar (TSA) was previously proposed and its feasibility was demonstrated with five strains. In this study, this method was applied to 16 strains to support its broad applicability. The cell sorting gate was previously determined based on the CFDA stainability alone. Now the structural properties of cells designated by forward and side-scattering intensities have been introduced as the second gating criteria. Under the optimum gate condition, 100 cells have been selected and sorted on TSA. Consequently, a 95% or higher colony-forming rate has been attained for every strain. A successful application to microaerophilic Campylobacter spp. is especially of great importance because it suggests further broader applicability.

  18. Fluorescence in situ hybridization and sequential catalysed reporter deposition (2C-FISH for the flow cytometric sorting of freshwater ultramicrobacteria

    Directory of Open Access Journals (Sweden)

    Stefan M Neuenschwander

    2015-03-01

    Full Text Available Flow cytometric sorting is a powerful tool to physically separate cells within mixed microbial communities. If combined with phylogenetic staining (fluorescence in situ hybridization, FISH it allows to specifically sort defined genotypic microbial populations from complex natural samples. However, the targeted enrichment of freshwater ultramicrobacteria, such as members of the LD12 clade of Alphaproteobacteria (SAR11-IIIb, is still challenging. Current FISH protocols, even in combination with signal amplification by catalysed reporter deposition (CARD, are not sufficiently sensitive for the distinction of these bacteria from background noise by flow cytometry, presumably due to their low ribosome content and small cell sizes. We, therefore, modified a CARD based flow sorting protocol with the aim of increasing its sensitivity to a level sufficient for ultramicrobacteria. This was achieved by a second signal amplification step mediated by horseradish peroxidase labelled antibodies targeted to the fluorophores that were previously deposited by CARD-FISH staining. The protocol was tested on samples from an oligo-mesotrophic lake. Ultramicrobacteria affiliated with LD12 Alphaproteobacteria could be successfully sorted to high purity by flow cytometry. The ratios of median fluorescence signal to background ranged around 20, and hybridization rates determined by flow cytometry were comparable to those obtained by fluorescence microscopy. Potential downstream applications of our modified cell staining approach range from the analysis of microdiversity within 16S rRNA-defined populations to that of functional properties, such as the taxon-specific incorporation rates of organic substrates.

  19. Detection of P-glycoprotein with a rapid flow cytometric functional assay using Fluo-3: evaluation of sensitivity, specificity and feasibility in multiparametric analysis.

    Science.gov (United States)

    Van Acker, K L; De Greef, C; Eggermont, J; Zhang, P; Vandenberghe, P; Boogaerts, M A

    1995-08-01

    The specificity and sensitivity of a flow cytometric assay simultaneously measuring expression and transport function of the multidrug resistance associated P-glycoprotein (Pgp) was evaluated. The monoclonal antibody (mAb), MRK16 was used to detect phenotypic Pgp expression while Fluo-3-AM was used as a fluorescent substrate in a Pgp functional transport assay. The specificity of the functional assay was examined in two vinblastine selected human leukemic cell lines (K562/VLB2.5 and CCRF-CEM/VLB50) with acquired Pgp overexpression. Downmodulation of Pgp function in these cell lines could be demonstrated with different substances (verapamil, vinblastine, trifluoperazine, cyclosporin A, progesterone and quinidine) and was proven to be consistently higher in the vinblastine selected cells than in their non-selected drug sensitive counterparts. Unexpectedly, modulator activity was also observed in drug sensitive K562 and CCRF-CEM cell lines despite the inability to detect Pgp in those cells by MRK16 flow cytometrically. Low level expression of the MDR1 gene encoding Pgp in sensitive K562 cells was however demonstrated with a sensitive RT-PCR procedure. The small effect of Pgp modulators in non-drug selected cells could therefore be attributed to low level basal expression of Pgp and illustrates the sensitivity of the functional assay. Also, the effect of various Pgp modulators on Pgp function was more pronounced in a subpopulation of Pgp expressing lymphocytes than in lymphocytes which did not express Pgp. Finally, a correlation was found between discrete variations in Pgp expression and Pgp function of CD4+ lymphocytes, underscoring the feasibility of the functional assay in a triple parametric procedure. The triple parametric assay holds promise to detect Pgp expression and function in clinical samples containing mixtures of malignant and non-malignant cells.

  20. Flow cytometric analysis with a fluorescently labeled formyl peptide receptor ligand as a new method to study the pharmacological profile of the histamine H2 receptor.

    Science.gov (United States)

    Werner, Kristin; Kälble, Solveig; Wolter, Sabine; Schneider, Erich H; Buschauer, Armin; Neumann, Detlef; Seifert, Roland

    2015-10-01

    The histamine H2 receptor (H2R) is a Gs protein-coupled receptor. Its activation leads to increases in the second messenger adenosine-3',5'-cyclic monophosphate (cAMP). Presently, several systems are established to characterize the pharmacological profile of the H2R, mostly requiring radioactive material, animal models, or human blood cells. This prompted us to establish a flow cytometric analysis with a fluorescently labeled formyl peptide receptor (FPR) ligand in order to investigate the H2R functionally and pharmacologically. First, we stimulated U937 promonocytes, which mature in a cAMP-dependent fashion upon H2R activation, with histamine (HA) or selective H2R agonists and measured increases in cAMP concentrations by mass spectrometry. Next, indicative for the maturation of U937 promonocytes, we assessed the FPR expression upon incubation with HA or H2R agonists. FPR expression was measured either indirectly by formyl peptide-induced changes in intracellular calcium concentrations ([Ca(2+)]i) or directly with the fluorescein-labeled FPR ligand fNleLFNleYK-Fl. HA and H2R agonists concentration-dependently induced FPR expression, and potencies and efficacies of fMLP-induced increases in [Ca(2+)]i and FPR density correlated linearly. Accordingly, flow cytometric analysis of FPR expression constitutes a simple, inexpensive, sensitive, and reliable method to characterize the H2R pharmacologically. Furthermore, we evaluated FPR expression at the mRNA level. Generally, quantitative real-time polymerase chain reaction confirmed functional data. Additionally, our study supports the concept of functional selectivity of the H2R, since we observed dissociations in the efficacies of HA and H2R agonists in cAMP accumulation and FPR expression.

  1. Detection of chromosome aneuploidy in breast lesions with fluorescence in situ hybridization: Comparison of whole nuclei to thin tissue sections and correlation with flow cytometric DNA analysis

    Energy Technology Data Exchange (ETDEWEB)

    Visscher, D.W.; Wallis, T.; Ritchie, C.A. [Wayne State Univ., Detroit, MI (United States)

    1995-09-01

    We compared flow-cytometric DNA histogram pattern to counts of 4 fluorescent-labelled centromeric probes (chromosomes 1, 7, 8, and 17) in whole nuclei (WN) and in nuclei from formalin-fixed deparaffinized thin tissue section (TS) in 25 breast lesions. In benign lesions, signal gains (i.e., trisomic nuclei) were never observed in greater than 10% of nuclei from either WN or TS preparations. Loss of signal in benign breast lesions, however, varied considerably (0-43%) between individual case and between chromosome probes. The mean incidence of signal loss in WN of benign lesions ranged from 8.9% (chromosome 7) to 14.4 % (chromosome 1) of nuclei. These signal loss frequencies exceeded those of benign lymphoid control cells. In three benign lesions, signal loss in WN (with one probe) was observed in at least 25% of nuclei. Signal losses in benign TS, on average, were 50-150% greater than in matched WN preparations (chromosome 1: 21.7%, chromosome 7: 21.5%). Malignant lesions generally, but not always, displayed fewer monosomic nuclei and more trisomic nuclei in compared to TS, compatible with a slicing (i.e., nuclear truncation) artifact. Signal counts in carcinomas correlated well with flow cytometric DNA index; however, they were also characterized by evidence of genetic instability, manifest as signal gains in a subset of nuclei (10-25%) with individual probes in diploid range cases, as well as intratumoral heterogeneity, reflected as discrepancies in probe counts between WN and TS samples. We conclude that signal losses with centromeric probes are largely, but not entirely, explained by nuclear slicing. The minimum signal loss threshold for establishment of monosomy using interphase cytogenetics is thus unclear, even in WN. Signal gains indicative of trisomy, in contrast, are reliably associated with malignancy and may reflect gross DNA aneuploidy as well as genetic instability. 10 refs., 1 fig., 3 tabs.

  2. Flow Cytometric DNA Analysis Using Cytokeratin Labeling for Identification of Tumor Cells in Carcinomas of the Breast and the Female Genital Tract

    Directory of Open Access Journals (Sweden)

    Rainer Kimmig

    2001-01-01

    Full Text Available Flow cytometric assessment of DNA‐ploidy and S‐phase fraction in malignant tumors is compromised by the heterogeneity of cell subpopulations derived from the malignant and surrounding connective tissue, e.g., tumor, stromal and inflammatory cells. To evaluate the effect on quality of DNA cell cycle analysis and determination of DNA ploidy, cytokeratin labeling of epithelial cells was used for tumor cell enrichment in breast, ovarian, cervical and endometrial cancer prior to DNA analysis. In a prospective study, tumor cell subpopulations of 620 malignant tumors were labeled by a FITC‐conjugated cytokeratin antibody (CK 5, 6, CK18 and CK 5, 6, 8 and CK 17, respectively prior to flow cytometric cell cycle analysis. Compared to total cell analysis, detection rate of DNA‐aneuploid tumors following cytokeratin labeling was increased from 62% to 76.5% in breast cancer, from 68% to 77% in ovarian cancer, from 60% to 80% in cervical cancer and from 30% to 53% in endometrial cancer. Predominantly in DNA‐diploid tumors, a significantly improved detection of S‐phase fraction of the tumor cells was shown due to the elimination of contaminating nonproliferating “normal cells”. S‐phase fraction following tumor cell enrichment was increased by 10% (mean following cytokeratin staining in ovarian and endometrial cancer, by 30% in breast cancer and even by 70% in cervical cancer compared to total cell analysis. Thus, diagnostic accuracy of DNA‐analysis was enhanced by cytokeratin labeling of tumor cells for all tumor entities investigated.

  3. Dose-dependent effect of 17 beta-estradiol determined by growth curves and flow cytometric DNA analysis of a human breast carcinoma (T61) grown in nude mice

    DEFF Research Database (Denmark)

    Brünner, N; Spang-Thomsen, M; Vindeløv, L

    1985-01-01

    of the treatment was evaluated using growth curves and flow cytometric DNA analysis. The treatment induced a dose-dependent growth delay and dose-dependent changes in the cell cycle distribution. The cell cycle changes comprised a decrease in the G1 phase, an accumulation of cells in the S phase, and an increasing...

  4. Identification of microbes from the surfaces of food-processing lines based on the flow cytometric evaluation of cellular metabolic activity combined with cell sorting.

    Science.gov (United States)

    Juzwa, W; Duber, A; Myszka, K; Białas, W; Czaczyk, K

    2016-09-01

    In this study the design of a flow cytometry-based procedure to facilitate the detection of adherent bacteria from food-processing surfaces was evaluated. The measurement of the cellular redox potential (CRP) of microbial cells was combined with cell sorting for the identification of microorganisms. The procedure enhanced live/dead cell discrimination owing to the measurement of the cell physiology. The microbial contamination of the surface of a stainless steel conveyor used to process button mushrooms was evaluated in three independent experiments. The flow cytometry procedure provided a step towards monitoring of contamination and enabled the assessment of microbial food safety hazards by the discrimination of active, mid-active and non-active bacterial sub-populations based on determination of their cellular vitality and subsequently single cell sorting to isolate microbial strains from discriminated sub-populations. There was a significant correlation (r = 0.97; p flow cytometry, despite there being differences in the absolute number of cells detected. The combined approach of flow cytometric CRP measurement and cell sorting allowed an in situ analysis of microbial cell vitality and the identification of species from defined sub-populations, although the identified microbes were limited to culturable cells.

  5. Flow cytometric analysis of p21 protein expression on irradiated human lymphocytes; Analise por citometria de fluxo da expressao da proteina p21 em linfocitos humanos irradiados

    Energy Technology Data Exchange (ETDEWEB)

    Santos, N.F.G.; Amaral, A., E-mail: neyliane@gmail.com [Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil). Departamento de Energia Nuclear. Laboratorio de Modelagem e Biodosimetria Aplicada; Freitas-Silva, R. [Universidade Federal de Pernambuco (UFPE), Garanhuns, PE (Brazil). Departamento de Ciencias Naturais e Exatas; Pereira, V.R.A. [Fundacao Oswaldo Cruz (FIOCRUZ), Recife, PE (Brazil). Centro de Pesquisas Aggeu Magalhaes. Departamento de Imunologia. Lab. de Imunoparasitologia; Tasat, D.R. [Universidad Nacional de General San Martin, Buenos Aires (Argentina). Escuela de Ciencia y Tecnologia. Laboratorio de Biologia Celular del Pulmon

    2013-08-15

    Cell cycle blockage in G1 is a mechanism p21 protein-regulated and coupled to DNA damage response to permit genetic content analysis, damage repair and cell death. Analysis of proteins that participates of this response has progressed with new analytic tools, and data contributes to comprehension of radioinduced molecular events as well as to new approaches on practices that employ ionizing radiation. On this perspective, the aim of this research was to evaluate, by flow cytometry, p21 expression on irradiated human lymphocytes, maintained under different experimental conditions. Peripheral blood samples from 10 healthy subjects were irradiated with doses of 0 (non-irradiated), 1, 2 and 4 Gy. Lymphocytes were processed to analysis on ex vivo (no cultured) condition and after 24; 48 and 72 hours culture, with and without phytohemagglutinin stimulation. p21 protein expression levels were measured by flow cytometry, as percentage values. Results indicate that flow cytometric assay allows detection of changes on p21 expression, since it was detected significant increase on phytohemagglutinin-stimulated samples, for all times, against basal expression (ex vivo). However, it was not observed significant alterations on p21 protein radioinduced levels, for all doses, times and culture conditions analyzed. These results not indicate so p21 protein as bioindicator of ionizing radiation exposure. Nevertheless, data confirmation may to require analysis of a more numerous population. (author)

  6. Development of flow cytometric protocol for nuclear DNA content estimation and determination of chromosome number in Pongamia pinnata L., a valuable biodiesel plant.

    Science.gov (United States)

    Ramesh, Aadi Moolam; Basak, Supriyo; Choudhury, Rimjhim Roy; Rangan, Latha

    2014-01-01

    The potentiality of Pongamia pinnata L. as a sustainable source of feedstock for the biodiesel industry is dependent on an extensive knowledge of the genome structure of the plant. Flow cytometry, with propidium iodide (PI) as the DNA stain, was used to estimate the nuclear DNA content of P. pinnata, with respect to Zea mays 'CE-777' as standard. The internal and pseudo-internal standardization was followed on account of the inhibitory effect of secondary compounds on PI intercalation. The antioxidants (PVP-40 and β-mercaptoethanol) were added to the nuclear isolation buffer for the reduction of inhibitory effect of P. pinnata cytosol. Nuclear DNA content estimation was done for P. pinnata leaves from different altitudes (37-117 m height from sea level) of Assam. Flow cytometry analysis indicated that the nuclear DNA content of P. pinnata is 2.66 pg with predicted 1C value of 1,300 Mb using Z. mays as standard. Coefficient of variation in flow cytometric analysis was within the limit of 5 % indicating that the results were reliable. Somatic chromosome numbers were counted from root-tip cells and was found to be 2n = 22 corresponding to the diploid level (x = 11). A decreasing trend in the nuclear DNA content was observed for the species of different altitudes.

  7. Establishment of flow cytometric in micronucleus assay in vitro%流式细胞术检测体外微核方法的建立

    Institute of Scientific and Technical Information of China (English)

    欧红梅; 周长慧; 涂宏刚; 黄鹏程; 常艳

    2015-01-01

    OBJECTIVE:Establish the flow cytometric 96-well microplate-basedin vitro micronucleus assay in CHO-K1 cells,and explore the possibility of this method for early genetic toxicity screening during drug discovery. MEHTODS:The test included treatment with and without metabolic activation. For the treatment with metabolic activation,CHO-K1 cells were treated with three different concentrations of cyclophosphamide in the S9 mixmedium for 4 h,then incubated with S9-free fresh medium for 20 h. For the treatment without metabolic activation,cells were incubated with three different concentrations of mitomycin C continuously for 24 h. In all cases,after a total of 24 h since initiation of the treatment,cells were processed for microscopic scoring or flow cytometric MN analysis. A flow cytometric method for scoring MN used EMA and SYTOX Green to label the cells in 96-well microplate,and then compared with cytokinesis-block micronucleus assay in cell culture disks based on microscopy.RESULTS:Mitomycin C and cyclophosphamide at different concerntrations caused statistically significant and dose-dependent increasess in micronucleus assay . Non-parametric Spearman's coefficients (rs) is 1.000.CONCLUSION:Similar to literature published,mitomycin C and cyclophosphamide induced positive results in flow cytometric based in vitro micronucleus assay. So the method of flow cytometric 96-well microplate-based in vitro micronucleus assay in CHO-K1 cells was established. The concordance between microscopic scoring and flow cytometricwas good,therefore this method is promising for screening and evaluating genetic toxicity of chemicals.%目的:建立96孔板流式细胞术体外微核自动化检测的方法,并探讨其用于药物早期遗传毒性筛选和遗传毒性评价的可能性。方法:试验分为+S9短时处理组(4 h)和-S9持续处理组(24 h),分别选择3个不同浓度的环磷酰胺和丝裂霉素C处理CHO-K1细胞,24 h后收获细胞。采用EMA和SYTOX Green

  8. Flow cytometric assay to assess short-term effects of personal care products on the marine microalga Tetraselmis suecica.

    Science.gov (United States)

    Seoane, Marta; Esperanza, Marta; Rioboo, Carmen; Herrero, Concepción; Cid, Ángeles

    2017-03-01

    Large quantities of personal care products (PCPs) are used daily and many of their chemical ingredients are subsequently released into marine environments. Cultures of the marine microalga Tetraselmis suecica were exposed for 24 h to three emerging compounds included in the main classes of PCPs: the UV filter benzophenone-3 (BP-3), the disinfectant triclosan (TCS) and the fragrance tonalide (AHTN). Concentrations tested, expressed as cellular quota (pg cell(-1)), ranged from 5 to 40 for BP-3, from 2 to 16 for TCS and from 1.2 to 2.4 for AHTN. A small cytometric panel was carried out to evaluate key cytotoxicity biomarkers including inherent cell properties, growth and metabolic activity and cytoplasmic membrane properties. BP-3 caused a significant increase in growth rate, metabolic activity and chlorophyll a fluorescence from 10 pg cell(-1). However, growth and esterase activity decreased in cells exposed to all TCS and AHTN concentrations, except the lowest ones. Also these two compounds provoked a significant swelling of cells, more pronounced in the case of TCS-exposed cells. Although all treated cells remained viable, changes in membrane potential were observed. BP-3 and AHTN caused a significant depolarization of cells from 10 to 1.6 pg cell(-1), respectively; however all TCS concentrations assayed caused a noticeable hyperpolarization of cells. Metabolic activity and cytoplasmic membrane potential were the most sensitive parameters. It can be concluded that the toxicological model used and the toxicological parameters evaluated are suitable to assess the toxicity of these emerging contaminants.

  9. Flow cytometric analysis of kappa and lambda light chain expression in endoscopic biopsy specimens before the diagnosis of B-cell lymphoma.

    Science.gov (United States)

    Oka, Satoko; Muroi, Kazuo; Sato, Kazuya; Fujiwara, Shin-ichiro; Oh, Iekuni; Matsuyama, Tomohiro; Ohmine, Ken; Suzuki, Takahiro; Ozaki, Katsutoshi; Mori, Masaki; Nagai, Tadashi; Fukushima, Noriyoshi; Fukushima, Noriyoshi; Tanaka, Akira; Ozawa, Keiya

    2012-01-01

    Forty-eight patients with gastrointestinal (GI) tract B-cell lymphoma (BCL) were analyzed retrospectively. The diagnosis was based on the histological examination of specimens obtained by endoscopic biopsy. Before the diagnosis was made, single-color flow cytometry was performed to analyze the expression of light chains and B-cell antigens including CD10 in the specimens. Restricted light chain (RLC) expression, a marker of B-cell clonality, was defined as κ and λ ratios of either more than 3.0 or less than 0.5. The specimens from 30 patients (62.5%) showed RLC expression. No RLC expression or RLC expression not examined was divided into two groups : those showing CD10 positivity in more than 20% of cells (4 patients, 8.3%) and those showing no positivity (14 patients, 29.2%). The cell number analyzed in the latter group was significantly smaller than that in the other two groups. Abnormal karyotypes were found in the specimens from 8 patients (16.7%). These results indicate that the flow cytometric analysis of endoscopic biopsy specimens is useful when BCL is suspected if an adequate number of cells are obtained.

  10. Scales and kinetics of granular flows.

    Science.gov (United States)

    Goldhirsch, I.

    1999-09-01

    When a granular material experiences strong forcing, as may be the case, e.g., for coal or gravel flowing down a chute or snow (or rocks) avalanching down a mountain slope, the individual grains interact by nearly instantaneous collisions, much like in the classical model of a gas. The dissipative nature of the particle collisions renders this analogy incomplete and is the source of a number of phenomena which are peculiar to "granular gases," such as clustering and collapse. In addition, the inelasticity of the collisions is the reason that granular gases, unlike atomic ones, lack temporal and spatial scale separation, a fact manifested by macroscopic mean free paths, scale dependent stresses, "macroscopic measurability" of "microscopic fluctuations" and observability of the effects of the Burnett and super-Burnett "corrections." The latter features may also exist in atomic fluids but they are observable there only under extreme conditions. Clustering, collapse and a kinetic theory for rapid flows of dilute granular systems, including a derivation of boundary conditions, are described alongside the mesoscopic properties of these systems with emphasis on the effects, theoretical conclusions and restrictions imposed by the lack of scale separation. (c) 1999 American Institute of Physics.

  11. Experimental Studies on Turbulence Kinetic Energy in Confined Vortex Flows

    Institute of Scientific and Technical Information of China (English)

    L.Yan; G.H.Vatistas; 等

    2000-01-01

    Turbulence kinetic energies in confined vortex flows have been studied.The studies were based on the experiments performed in a vortex chamber,In the experiments,a Laser Doppler Anemometry(LDA) was used to perform flow measurements inside the vortex chamber,which provided the data for the kinetic energy analysis.The studies concentrated on the influences of the contraction ratio and the inlet air flow rate on the kinetic energy,and analyzed the characteristics of the kinetic energy in the confined vortex flows,including the distributions of the tangential component,radial component and total turbulence kinetic energy,In the paper,both the experimental techniques and the experimental results were presented.Based on a similarity analyis and the experimental data,an empirical scaling formula was proposed so that the tangential component of the turbulence kinetic energy was dependent only on the parameter of the contraction ratio.

  12. Flow cytometric immunobead assay for fast and easy detection of PML-RARA fusion proteins for the diagnosis of acute promyelocytic leukemia.

    Science.gov (United States)

    Dekking, E H A; van der Velden, V H J; Varro, R; Wai, H; Böttcher, S; Kneba, M; Sonneveld, E; Koning, A; Boeckx, N; Van Poecke, N; Lucio, P; Mendonça, A; Sedek, L; Szczepański, T; Kalina, T; Kanderová, V; Hoogeveen, P; Flores-Montero, J; Chillón, M C; Orfao, A; Almeida, J; Evans, P; Cullen, M; Noordijk, A L; Vermeulen, P M; de Man, M T; Dixon, E P; Comans-Bitter, W M; van Dongen, J J M

    2012-09-01

    The PML-RARA fusion protein is found in approximately 97% of patients with acute promyelocytic leukemia (APL). APL can be associated with life-threatening bleeding complications when undiagnosed and not treated expeditiously. The PML-RARA fusion protein arrests maturation of myeloid cells at the promyelocytic stage, leading to the accumulation of neoplastic promyelocytes. Complete remission can be obtained by treatment with all-trans-retinoic acid (ATRA) in combination with chemotherapy. Diagnosis of APL is based on the detection of t(15;17) by karyotyping, fluorescence in situ hybridization or PCR. These techniques are laborious and demand specialized laboratories. We developed a fast (performed within 4-5 h) and sensitive (detection of at least 10% malignant cells in normal background) flow cytometric immunobead assay for the detection of PML-RARA fusion proteins in cell lysates using a bead-bound anti-RARA capture antibody and a phycoerythrin-conjugated anti-PML detection antibody. Testing of 163 newly diagnosed patients (including 46 APL cases) with the PML-RARA immunobead assay showed full concordance with the PML-RARA PCR results. As the applied antibodies recognize outer domains of the fusion protein, the assay appeared to work independently of the PML gene break point region. Importantly, the assay can be used in parallel with routine immunophenotyping for fast and easy diagnosis of APL.

  13. A flow cytometric comparison of Indo-1 to fluo-3 and Fura Red excited with low power lasers for detecting Ca(2+) flux.

    Science.gov (United States)

    Bailey, Sheree; Macardle, Peter J

    2006-04-20

    Indo-1 and high-power water-cooled lasers have been the standard for flow cytometric based Ca(2+) flux measurements. With advances in technology and the availability of low-power air-cooled lasers, there is interest in alternative protocols. Here, we have compared Indo-1 with the combination of fluo-3 and Fura Red calcium indicator dyes using low-power air-cooled lasers as the excitation source. The reagents were examined in parallel to detect Ca(2+) flux in peripheral blood T lymphocytes and in a T lymphoblastoid cell line. Ca(2+) flux was detected with a FACSVantage SE equipped with an Omnichrome Series 74 Helium-Cadmium, or a Spectra Physics 177-G1202 Argon ion air-cooled laser. Following determination of optimal loading conditions, Ca(2+) flux was examined in response to membrane receptor stimulation or intracellular Ca(2+) mobilization. Dose dependent Ca(2+) flux to anti-CD3 and thapsigargin was detected with either Indo-1 or with fluo-3 and Fura Red. The profile of the Ca(2+) flux detected by Indo-1 or with fluo-3 and Fura Red appeared similar, with the combination of fluo-3 and Fura Red more sensitive under the particular test conditions. The results clearly demonstrated that Indo-1 could be usefully excited with a low-power air-cooled laser. The alternative use of fluo-3 and Fura Red does not require the availability of a UV capable laser and produced equivalent data.

  14. Identification of New Rat Bone Marrow-Derived Population of Very Small Stem Cell with Oct-4A and Nanog Expression by Flow Cytometric Platforms

    Directory of Open Access Journals (Sweden)

    Anna Labedz-Maslowska

    2016-01-01

    Full Text Available Very small embryonic-like stem cells (VSELs represent a unique rare population of adult stem cells (SCs sharing several structural, genetic, biochemical, and functional properties with embryonic SCs and have been identified in several adult murine and human tissues. However, rat bone marrow- (BM- derived SCs closely resembling murine or human VSELs have not been described. Thus, we employed multi-instrumental flow cytometric approach including classical and imaging cytometry and we established that newly identified population of nonhematopoietic cells expressing CD106 (VCAM-I antigen contains SCs with very small size, expressing markers of pluripotency (Oct-4A and Nanog on both mRNA and protein levels that indicate VSEL population. Based on our experience in both murine and human VSEL isolation procedures by fluorescence-activated cell sorting (FACS, we also optimized sorting protocol for separation of CD45−/Lin−/CD106+ rat BM-derived VSELs from wild type and eGFP-expressing rats, which are often used as donor animals for cell transplantations in regenerative studies in vivo. Thus, this is a first study identifying multiantigenic phenotype and providing sorting protocols for isolation VSELs from rat BM tissue for further examining of their functional properties in vitro as well as regenerative capacity in distinct in vivo rat models of tissue injury.

  15. Response of Syngonium podophyllum L. ‘White Butterfly’ shoot cultures to alternative media additives and gelling agents, and flow cytometric analysis of regenerants

    Directory of Open Access Journals (Sweden)

    JAIME A. TEIXEIRA DA SILVA

    2015-05-01

    Full Text Available Abstract. Teixeira da Silva JA. 2015. Response of Syngonium podophyllum L. ‘White Butterfly’ shoot cultures to alternative media additives and gelling agents, and flow cytometric analysis of regenerants. Nusantara Bioscience 7: 26-32. Syngonium podophyllum L. (arrowhead vine is a popular leafy indoor pot plant whose tissue culture has been established, primarily through in vitro shoot culture, but several interesting aspects have not yet been explored. In this study, cv. ‘White Butterfly’ was used to investigate the response of shoot formation to alternative gelling agents and media additives. Gellan gum (Gelrite® at 2 g/L resulted in greater leaf production, plantlet fresh weight and higher chlorophyll content (SPAD value than all other gelling agents tested, including agar, Bacto agar, phytagel, oatmeal agar, potato dextrose agar, barley starch and corn starch, when on a basal Hyponex® (NPK = 6.5: 6: 19; 3 g/L medium. Several alternative liquid medium additives tested (low and full fat milk, Coca-Cola®, coffee, Japanese green, Oolong and Darjeeling teas negatively impacted plant growth, stunted roots and decreased chlorophyll content (SPAD value of leaves. Plant growth on medium with refined sucrose or table sugar responded similarly. Poor growth was observed when crude extract from a high rebaudioside-containing stevia (Stevia rebaudiana Bertoni line - an artificial sweetener - was used. Leaf tissue from the control did not show any endopolyploidy but low levels of endopolyploidy (8C were detected in some treatments.

  16. The level of heparin-induced antibodies in correlation with the result of the flow cytometric functional assay in the patients with suspected HIT.

    Science.gov (United States)

    Maličev, Elvira; Maček Kvanka, Marjeta; Klemenc, Polona; Rožman, Primož

    2017-09-13

    Heparin can induce the formation of antibodies against a heparin complex with a platelet factor 4 (PF4), leading to platelet activation and the development of heparin-induced thrombocytopaenia (HIT). Because screening ELISA does not discriminate between platelet activating and non-activating anti-heparin/PF4 antibodies, each positive result is confirmed by an additional functional assay. We analysed 1004 sera of patients with suspected HIT. Optical density (OD) values of ELISA-positive results were correlated with the risk for a positive result with our functional flow cytometric assay. Only 10.7% were ELISA positive and 59.8% of those were positive with the functional assay. The positive functional assay was found in 23.4% of patients with OD2.0. Although our results showed that higher ELISA OD values increasethe possibility of the presence of platelet-activating anti-heparin/PF4 antibodies - , there is no need for improving ELISA cut-off value for positive result. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  17. Flow cytometric assessment of circulating platelet and erythrocytes microparticles in young thalassemia major patients: relation to pulmonary hypertension and aortic wall stiffness.

    Science.gov (United States)

    Tantawy, Azza A G; Adly, Amira A M; Ismail, Eman A R; Habeeb, Nevin M

    2013-06-01

    Heart disease is the leading cause of mortality and morbidity in β-thalassemia major (β-TM). Aggregability of abnormal red cells and membrane-derived microparticles (MPs) stemming from activated platelets and erythrocytes are responsible for thrombotic risk. We measured platelet and erythrocyte MPs (PMPs and ErMPs) in 60 young β-TM patients compared with 40 age- and sex-matched healthy controls and assessed their relation to clinicopathological characteristics and aortic elastic properties. Patients were studied stressing on transfusion history, splenectomy, thrombotic events, chelation therapy, hematological and coagulation profiles, flow cytometric measurement of PMPs (CD41b(+) ) and ErMPs (glycophorin A(+) ) as well as echocardiographic assessment of aortic elastic properties. Aortic stiffness index and pulmonary artery pressure were significantly higher, whereas aortic strain and distensibility were lower in TM patients than controls (P 2500 μg/L (P < 0.001). Compliant patients on chelation therapy had lower MPs levels than non-compliant patients (P < 0.001). PMPs and ErMPs were positively correlated to markers of hemolysis, serum ferritin, D-dimer, vWF Ag, and aortic stiffness, whereas negatively correlated to hemoglobin level and aortic distensibility (P < 0.05). We suggest that increased MPs may be implicated in vascular dysfunction, pulmonary hypertension risk, and aortic wall stiffness observed in thalassemia patients. Their quantification could provide utility for early detection of cardiovascular abnormalities and monitoring the biological efficacy of chelation therapy.

  18. A Simple Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity Enables Characterization of Affinity Maturation to Influenza A Virus

    Science.gov (United States)

    Frank, Gregory M.; Ince, William L.; Gibbs, James S.; Khurana, Surender; Wheatley, Adam K.; Max, Edward E.; McDermott, Adrian B.; Golding, Hana; Stevens, James; Bennink, Jack R.

    2015-01-01

    ABSTRACT Antibody (Ab) affinity maturation enables an individual to maintain immunity to an increasing number of pathogens within the limits of a total Ig production threshold. A better understanding of this process is critical for designing vaccines that generate optimal Ab responses to pathogens. Our study describes a simple flow-cytometric method that enumerates virus-specific germinal center (GC) B cells as well as their AC50, a measure of Ab avidity, defined as the antigen concentration required to detect 50% of specific B cells. Using a model of mouse Ab responses to the influenza A virus hemagglutinin (IAV HA), we obtained data indicating that AC50 decreases with time postinfection in an affinity maturation-dependent process. As proof of principle of the utility of the method, our data clearly show that relative to intranasal IAV infection, intramuscular immunization against inactivated IAV in adjuvant results in a diminished GC HA B cell response, with increased AC50 correlating with an increased serum Ab off-rate. Enabling simultaneous interrogation of both GC HA B cell quantity and quality, this technique should facilitate study of affinity maturation and rational vaccine design. PMID:26242629

  19. Rapid detection of BCR-ABL fusion genes using a novel combined LUX primer, in-cell RT-PCR and flow cytometric method.

    Science.gov (United States)

    Shi, Yan; Li, Li-Zhen; Sun, Jian-Zhi; Zhang, Ti; Peng, Jun; Xu, Cong-Gao

    2008-01-01

    Currently, quantitative and semiquantitative assays for minimal residual disease detection include fluorescence in situ hybridisation, multiparameter flow cytometric immunophenotyping and real-time quantitative polymerase chain reaction (RQ-PCR). We have developed a new approach to detect hybrid breakpoint cluster region and Abelson proto-oncogene (BCR-ABL) transcripts inside suspension cells using in situ RT-PCR and light upon extension (LUX) primer, followed by rapid quantitative analysis with flow cytometry. After cellular permeabilization and fixation of single cell suspension, the neoplastic mRNA was reverse transcribed and amplified by PCR with LUX primer. The results demonstrated that a strong positive yellow-green signal was observed in 99-100% cells of K562 cell line, only the red nucleus was detected in NB4 cell line and normal controls. The technique has been utilised to study 12 patients with chronic myeloid leukemia, and the results were compared with those of BCR-ABL fusion mRNA by RT-PCR and BCR-ABL fusion gene of the interphase cells by fluorescence in situ hybridization (FISH). In the five diagnosed patients, 90-98% cells were strongly positive. Four patients, including three patients treated with interferon-alpha and hydroxyurea and one patient treated with imatinib mesylate, had 26-82.5% positive cells. Three patients treated with imatinib mesylate were negative. The in situ RT-PCR results demonstrated complete concordance with the results of I-FISH and RT-PCR. A fluorescence signal was detectable at 1/10(4) cells and became negative below this threshold with flow cytometry. The results of the present study suggest that (1) LUX primers can be used to efficiently detect BCR-ABL fusion mRNA by in-cell RT-PCR; (2) the novel technique is a specific and sensitive way of detecting fusion gene with potential clinical usefulness.

  20. Flow cytometric detection of neutrophil-associated immunoglobulin in patients with or without neutropenia and establishment of the reference interval.

    Science.gov (United States)

    Hwang, Keumrock; Park, Chan-Jeoung; Huh, Hee Jin; Han, Sang Hee; Jang, Seongsoo; Chi, Hyun-Sook

    2011-01-01

    We measured neutrophil-associated immunoglobulin (NAIg) levels using flow cytometry to establish the reference interval for NAIg and to estimate NAIg in patients with or without neutropenia. Peripheral blood from 152 individuals was analyzed for NAIg detection by flow cytometry. Using fluorescescent-conjugated anti-CD10 monoclonal antibody and anti-human immunoglobulins, proportions of NAIgG, NAIgA, and NAIgM bound to neutrophils were measured. Reference intervals for NAIg were set as NAIgG reference intervals defined herein, patients with neutropenia or adverse transfusion reactions may be evaluated in a clinically relevant manner.

  1. Development of a flow cytometric method to analyze subpopulations of bacteria in probiotic products and dairy starters

    NARCIS (Netherlands)

    Bunthof, C.J.; Abee, T.

    2002-01-01

    Flow cytometry (FCM) is a rapid and sensitive technique that can determine cell numbers and measure various physiological characteristics of individual cells by using appropriate fluorescent probes. Previously, we developed an FCM assay with the viability probes carboxyfluorescein diacetate (cFDA) a

  2. Development of a multiplex flow cytometric microsphere immunoassay for mycotoxins and evaluation of its application in feed

    NARCIS (Netherlands)

    Peters, J.; Ploum, M.E.; Rijk, de T.C.; Haasnoot, W.

    2011-01-01

    A multi-mycotoxin immunoassay—using the MultiAnalyte Profiling (xMAP) technology—is developed and evaluated. This technology combines a unique color-coded microsphere suspension array, with a dedicated flow cytometer. We aimed for the combined detection of aflatoxins, ochratoxin A, deoxynivalenol,

  3. Stereologic, histopathologic, flow cytometric, and clinical parameters in the prognostic evaluation of 74 patients with intraoral squamous cell carcinomas

    DEFF Research Database (Denmark)

    Bundgaard, T; Sørensen, Flemming Brandt; Gaihede, M

    1992-01-01

    BACKGROUND AND METHODS: A consecutive series of all 78 incident cases of intraoral squamous cell carcinoma occurring during a 2-year period in a population of 1.4 million inhabitants were evaluated by histologic score (the modified classification of Jacobsson et al.), flow cytometry, stereology, ...

  4. Quantitation of minimal disease levels in chronic lymphocytic leukemia using a sensitive flow cytometric assay improves the prediction of outcome and can be used to optimize therapy.

    Science.gov (United States)

    Rawstron, A C; Kennedy, B; Evans, P A; Davies, F E; Richards, S J; Haynes, A P; Russell, N H; Hale, G; Morgan, G J; Jack, A S; Hillmen, P

    2001-07-01

    Previous studies have suggested that the level of residual disease at the end of therapy predicts outcome in chronic lymphocytic leukemia (CLL). However, available methods for detecting CLL cells are either insensitive or not routinely applicable. A flow cytometric assay was developed that can differentiate CLL cells from normal B cells on the basis of their CD19/CD5/CD20/CD79b expression. The assay is rapid and can detect one CLL cell in 10(4) to 10(5) leukocytes in all patients. We have compared this assay to conventional assessment in 104 patients treated with CAMPATH-1H and/or autologous transplant. During CAMPATH-1H therapy, circulating CLL cells were rapidly depleted in responding patients, but remained detectable in nonresponders. Patients with more than 0.01 x 10(9)/L circulating CLL cells always had significant (> 5%) marrow disease, and blood monitoring could be used to time marrow assessments. In 25 out of 104 patients achieving complete remission by National Cancer Institute (NCI) criteria, the detection of residual bone marrow disease at more than 0.05% of leukocytes in 6 out of 25 patients predicted significantly poorer event-free (P =.0001) and overall survival (P =.007). CLL cells are detectable at a median of 15.8 months (range, 5.5-41.8) posttreatment in 9 out of 18 evaluable patients with less than 0.05% CLL cells at end of treatment. All patients with detectable disease have progressively increasing disease levels on follow-up. The use of sensitive techniques, such as the flow assay described here, allow accurate quantitation of disease levels and provide an accurate method for guiding therapy and predicting outcome. These results suggest that the eradication of detectable disease may lead to improved survival and should be tested in future studies.

  5. Development of a five-plex flow cytometric immunoassay for the simultaneous detection of six coccidiostats in feed and eggs

    OpenAIRE

    Bienenmann-Ploum, Monique E.; Huet, Anne-Catherine; Campbell, Katrina; Fodey, Terence L.; Vincent, Ursula; Haasnoot, Willem; Delahaut, Philippe; Elliott, Christopher T; Nielen, Michel W. F.

    2012-01-01

    Coccidiostats are the only veterinary drugs still permitted to be used as feed additives to treat poultry for coccidiosis. To protect consumers, maximum levels for their presence in food and feed have been set by the European Union (EU). To monitor these coccidiostats, a rapid and inexpensive screening method would be a useful tool. The development of such a screening method, using a flow cytometry-based immunoassay, is described. The assay uses five sets of colour-coded paramagnetic microsph...

  6. 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.

  7. Direct detection of red blood cell fragments: a new flow cytometric method to evaluate hemolysis in blood pumps.

    Science.gov (United States)

    Linneweber, J; Chow, T W; Takano, T; Maeda, T; Nonaka, K; Schulte-Eistrup, S; Kawahito, S; Elert, O; Moake, J L; Nosé, Y

    2001-01-01

    Pump induced hemolysis is presently evaluated by measuring plasma free hemoglobin (fHb). However, this method has disadvantages because quantification of fHb depends on hematocrit (HCT) and hemoglobin (Hb) levels. The aim of this work was to devise a hemoglobin independent method, capable of quantifying cell trauma directly by measuring the number of red blood cell (RBC) fragments. Whole blood flow cytometry was used to quantify circulating RBC fragments derived from a roller pump (Sarns, Inc. Model 2 M 6,002) and a centrifugal pump (Gyro C1E3, Kyocera Corp.). The pumps were tested in a mock circuit for 2 hr (5 L/min flow against 100 mm Hg pressure head). Red blood cell fragments were quantified by a phycoerythrin (PE) labeled glycophorin A antibody specific for erythrocytes. Red blood cell fragments were smaller than the intact RBC population and overlapped in size with the platelet population (based on forward- and side-light scattering measurements). For the roller pump, the values for RBC fragments increased from 1,090 +/- 260/microl at 0 min to 14,880 +/- 5,900/microl after 120 min. In contrast, using the centrifugal pump, there was little increase in RBC fragments (from 730 +/- 270/microl at 0 min to 1,400 +/- 840/microl after 120 min). Flow cytometry can be used for the rapid, sensitive, hemoglobin independent evaluation of pump induced RBC trauma.

  8. Improved flow cytometric identification of myelopoiesis by the simultaneous labelling with CD13, CD14 and CD66 monoclonal antibodies

    DEFF Research Database (Denmark)

    Bonde, J; Meyer, K; Broe, M K

    1996-01-01

    in the fast determination of remission state. In MDS, the immature myeloid component could be distinguished in patients defined according to the FAB classification with the possibility of identifying aberrant phenotypes, the assay should also be of interest in other myeloproliferative disorders. Moreover......The aim of the present study was to increase our knowledge of myelopoiesis evaluated by flow cytometry. We therefore designed a triple-marker assay employing monoclonal antibodies against the CD13 (immature), the CD14 (monocytic), and the CD66 (mature myeloid) antigens using three...

  9. Development of a five-plex flow cytometric immunoassay for the simultaneous detection of six coccidiostats in feed and eggs.

    Science.gov (United States)

    Bienenmann-Ploum, Monique E; Huet, Anne-Catherine; Campbell, Katrina; Fodey, Terence L; Vincent, Ursula; Haasnoot, Willem; Delahaut, Philippe; Elliott, Christopher T; Nielen, Michel W F

    2012-09-01

    Coccidiostats are the only veterinary drugs still permitted to be used as feed additives to treat poultry for coccidiosis. To protect consumers, maximum levels for their presence in food and feed have been set by the European Union (EU). To monitor these coccidiostats, a rapid and inexpensive screening method would be a useful tool. The development of such a screening method, using a flow cytometry-based immunoassay, is described. The assay uses five sets of colour-coded paramagnetic microspheres for the detection of six selected priority coccidiostats. Different coccidiostats, with and without carrier proteins, were covalently coupled onto different bead sets and tested in combination with polyclonal antisera and with a fluorescent-labelled secondary antibody. The five optimal combinations were selected for this multiplex and a simple-to-use sample extraction method was applied for screening blank and spiked eggs and feed samples. A very good correlation (r ranging from 0.995 to 0.999) was obtained with the responses obtained in two different flow cytometers (Luminex 100 and FLEXMAP 3D). The sensitivities obtained were in accordance with the levels set by the EU as the measured limits of detection for narasin/salinomycin, lasalocid, diclazuril, nicarbazin (4,4'-dinitrocarbanilide) and monensin in eggs were 0.01, 0.1, 0.5, 53 and 0.1 μg/kg and in feed 0.1, 0.2, 0.3, 9 and 1.5 μg/kg, respectively.

  10. Level 2 validation of a flow cytometric method for detection of Escherichia coli O157:H7 in raw spinach.

    Science.gov (United States)

    Williams, Anna J; Cooper, Willie M; Summage-West, Christine V; Sims, Lillie M; Woodruff, Robert; Christman, Jessica; Moskal, Ted J; Ramsaroop, Shawn; Sutherland, John B; Alusta, Pierre; Wilkes, Jon G; Buzatu, Dan A

    2015-12-23

    The Bacteriological Analytical Manual (BAM) method currently used by the United States Food and Drug Administration (FDA) to detect Escherichia coli O157:H7 in spinach was systematically compared to a new flow cytometry based method. This Food and Drug Administration (FDA) level 2 external laboratory validation study was designed to determine the latter method's sensitivity and speed for analysis of this pathogen in raw spinach. Detection of target cell inoculations with a low cell count is critical, since enterohemorrhagic strains of E. coli require an infective dose of as few as 10 cells (Schmid-Hempel and Frank, 2007). Although, according to the FDA, the infectious dose is unknown (Food and Drug Administration, 1993). Therefore, the inoculation level into the spinach, a total of 2.0±2.6 viable E. coli O157 cells, was specified to yield between 25% and 75% detection by the new method, out of 20 samples (10 positives and 10 negatives). This criterion was met in that the new method detected 60% of the nominally positive samples; the corresponding sensitivity of the reference method was 50%. For both methods the most likely explanation for false negatives was that no viable cells were actually introduced into the sample. In this validation study, the flow cytometry method was equal to the BAM in sensitivity and far superior in speed. Published by Elsevier B.V.

  11. Flow cytometric comparison of platelets from a whole blood and finger-prick sample: impact of 24 hours storage.

    Science.gov (United States)

    Swanepoel, Albe C; Stander, Andre; Pretorius, Etheresia

    2013-03-01

    In this study, we investigate the validity and laboratory utility of flow cytometry when analyzing platelet activation by studying CD41, CD42b, CD62P and CD63. We compare flow cytometry results from citrated whole-blood and finger-prick samples directly after collection and also after storing both a finger-prick and whole-blood sample for 24 hours. Citrated whole-blood and finger-prick samples were taken from three healthy individuals on two occasions, and a total of 60,000 cells were analyzed for each of the four phycoerythrin-labeled monoclonal antibodies. Half of each sample was analyzed immediately after sampling while the other half was kept in the fridge at 6 °C for 24 hours before analysis. No significant difference was found between the sampling methods or the period of time before analysis. Results therefore suggest that an appropriately prepared finger-prick sample can be used for platelet function analysis, and samples can be stored for 24 hours in the fridge at 6 °C before analysis.

  12. Flow Cytometric Determination of the Expression of gp 51 Protein of Bovine Leukaemia Virus in Experimentally Infected Sheep

    Directory of Open Access Journals (Sweden)

    Szczotka Maria

    2014-10-01

    Full Text Available The study was performed on lambs experimentally infected with bovine leukaemia virus (BLV. The presence of BLV antibodies in sera of infected animals was detected by agar gel immunodiffusion test and ELISA. Proviral DNA was detected by PCR and nested PCR. Dual-colour flow cytometry analysis was performed with the use of specific monoclonal antibodies against lymphocyte CD markers and gp51 viral envelope protein, followed by incubation with fluorescent-labelled secondary antibodies conjugated with FITC or PE. Gp51 viral envelope protein was detected in tumours caused by BLV infection. The BLV infection resulted in depletion of CD4+ lymphocytes, increase in CD8+ lymphocytes, and decrease in CD4+ to CD8+ ratio in infected sheep. Proliferation of IgM+ CD19+ cells was also detected. These cells had an immature character without tendency to differentiate, and their vitality was prolonged. Flow cytometry enabled detection of gp51 expression in sheep blood lymphocytes at the early stages of the infection, before detection of serum antibodies using ELISA.

  13. Flow cytometric minimal residual disease monitoring in children with acute lymphoblastic leukemia treated by regimens with reduced intensity

    Directory of Open Access Journals (Sweden)

    A. M. Popov

    2015-01-01

    Full Text Available 191 consecutive unselected children with acute lymphoblastic leukemia aged from 1 to 16 years were enrolled in the study. Bone marrow samples were obtained at the time of initial diagnostics as well as at days 15 (n = 188, 36 (n = 191, and 85 (n = 187 of remission induction. Minimal residual disease (MRD was assessed by 6–10-color flow cytometry. Flow cytometry data at day 15 allowed distinguishing three patients groups with significantly different outcome (p ˂ 0.0001: 35.64 % patients with MRD < 0.1 % represented 5-year event-free survival (EFS of 100 %; 48.40 % cases with 0.1 % ≤ MRD< 10 % had EFS 84.6 ± 4.2 %; 15.96 % patients with very high MRD (≥ 10 % belonged to group with poor outcome (EFS 56.7 ± 9.0 %. At the end of remission induction (day 36 36 children (18.85 % with MRD higher than 0.1 % had significantly worse outcome compared to remaining ones (EFS 49.4 ± 9.0 and 93.5 ± 2.1 % respectively; p ˂ 0.0001. From a clinical standpoint it is relevant to evaluate both low-risk and high-risk criteria. Multivariate analysis showed that day 15 MRD data is better for low-risk patients definition while end-induction MRD is the strongest unfavorable prognostic factor.

  14. Flow cytometric quantification of all phases of the cell cycle and apoptosis in a two-color fluorescence plot.

    Directory of Open Access Journals (Sweden)

    Christine Vignon

    Full Text Available An optimal technology for cell cycle analysis would allow the concomitant measurement of apoptosis, G0, G1, S, G2 and M phases in combination with cell surface phenotyping. We have developed an easy method in flow cytometry allowing this discrimination in an only two-color fluorescent plot. It is based on the concomitant use of 7-amino-actinomycin D and the antibodies anti-Ki67 and anti-phospho(Ser10-histone H3, both conjugated to Alexa Fluor®488 to discriminate G0 and M phases, respectively. The method is particularly valuable in a clinical setting as verified in our laboratory by analyzing human leukemic cells from marrow samples or after exposure to cell cycle modifiers.

  15. Flow cytometric assessment of chicken T cell-mediated immune responses after Newcastle disease virus vaccination and challenge

    DEFF Research Database (Denmark)

    Dalgaard, T. S.; Norup, L. R.; Pedersen, A.R.

    2010-01-01

    . Despite a delayed NDV-specific antibody response to vaccination, L133 appeared to be better protected than L130 in the subsequent infection challenge as determined by the presence of viral genomes. Peripheral blood was analyzed by flow cytometry and responses in vaccinated/challenged birds were studied....... Furthermore, peripheral lymphocytes from L133 exhibited a significantly higher expression of CD44 and CD45 throughout the experiment. Interestingly, also vaccine-induced differences were observed in L133 as immune chickens had a significantly higher CD45 expression on their lymphocytes than the naïve controls....... Immune chickens from both lines had a significantly higher frequency of circulating γδ T cells than the naïve controls both after vaccination and challenge. Finally, the proliferative capacity of peripheral CD4+ and CD8+ cells specific for NDV was addressed 3 weeks after vaccination and 1 week after...

  16. Genetic stock assessment of spawning arctic cisco (Coregonus autumnalis) populations by flow cytometric determination of DNA content.

    Science.gov (United States)

    Lockwood, S F; Bickham, J W

    1991-01-01

    Intraspecific variation in cellular DNA content was measured in five Coregonus autumnalis spawning populations from the Mackenzie River drainage, Canada, using flow cytometry. The rivers assayed were the Peel, Arctic Red, Mountain, Carcajou, and Liard rivers. DNA content was determined from whole blood preparations of fish from all rivers except the Carcajou, for which kidney tissue was used. DNA content measurements of kidney and blood preparations of the same fish from the Mountain River revealed statistically indistinguishable results. Mosaicism was found in blood preparations from the Peel, Arctic Red, Mountain, and Liard rivers, but was not observed in kidney tissue preparations from the Mountain or Carcajou rivers. The Liard River sample had significantly elevated mean DNA content relative to the other four samples; all other samples were statistically indistinguishable. Significant differences in mean DNA content among spawning stocks of a single species reinforces the need for adequate sample sizes of both individuals and populations when reporting "C" values for a particular species.

  17. Flow cytometric analysis of the graft-versus-Leukemia-effect after hematopoietic stem cell transplantation in mice.

    Science.gov (United States)

    Schmidt, Felix; Hilger, Nadja; Oelkrug, Christoper; Svanidze, Ellen; Ruschpler, Peter; Eichler, Wolfram; Boldt, Andreas; Emmrich, Frank; Fricke, Stephan

    2015-04-01

    Acute Graft-versus-Host-Disease (aGvHD) is one of the major complications following allogeneic hematopoietic stem cell transplantation (HSCT). Although rather helpful, the use of conventional immunosuppressive drugs leads to general immunosuppression and is toxic. The effects of CD4(+) T-cells, in respect to the development of aGvHD, can be altered by administration of antihuman CD4 monoclonal antibodies, here MAX.16H5 IgG1 . This approach must be tested for possible interference with the Graft-versus-Leukemia-Effect (GvL). Thus, in vitro experiments were conducted, exposing P815 leukemic cells to bone marrow and splenocytes from cd4(-/-) -C57Bl/6 mice transgenic for human CD4 and HLA-DR3 (triple transgenic mice, [TTG]) as well as previously irradiated splenocytes from Balb/c(wt) mice. Using flow cytometry, the vitality of the various malignant and graft cells was analyzed over the course of 4 days. The survival rate of P815 cells did not change significantly when exposed to MAX.16H5 IgG1 , neither did the viability of the graft cells. This provides evidence that MAX.16H5 IgG1 does not impair the GvL effect in vitro. Additionally, P815-Balb/c(wt) leukemic mice were transplanted with P815(GFP) cells, bone marrow, and splenocytes from TTG mice with and without MAX.16H5 IgG1 . Without transplantation, P815(GFP) leukemic cells could be detected by flow cytometry in the liver, the bone marrow, and the spleen of recipients. The antibodies prevented aGvHD while leaving the GvL effect intact. These findings indicate no negative effect of MAX.16H5 IgG1 on the GvL effect in vitro and in vivo after HSCT in a murine model.

  18. Flow cytometric determination of stem/progenitor content in epithelial tissues: an example from nonsmall lung cancer and normal lung.

    Science.gov (United States)

    Donnenberg, Vera S; Landreneau, Rodney J; Pfeifer, Melanie E; Donnenberg, Albert D

    2013-01-01

    Single cell analysis and cell sorting has enabled the study of development, growth, differentiation, repair and maintenance of "liquid" tissues and their cancers. The application of these methods to solid tissues is equally promising, but several unique technical challenges must be addressed. This report illustrates the application of multidimensional flow cytometry to the identification of candidate stem/progenitor populations in non-small cell lung cancer and paired normal lung tissue. Seventeen paired tumor/normal lung samples were collected at the time of surgical excision and processed immediately. Tissues were mechanically and enzymatically dissociated into single cell suspension and stained with a panel of antibodies used for negative gating (CD45, CD14, CD33, glycophorin A), identification of epithelial cells (intracellular cytokeratin), and detection of stem/progenitor markers (CD44, CD90, CD117, CD133). DAPI was added to measure DNA content. Formalin fixed paraffin embedded tissue samples were stained with key markers (cytokeratin, CD117, DAPI) for immunofluorescent tissue localization of populations detected by flow cytometry. Disaggregated tumor and lung preparations contained a high proportion of events that would interfere with analysis, were they not eliminated by logical gating. We demonstrate how inclusion of doublets, events with hypodiploid DNA, and cytokeratin+ events also staining for hematopoietic markers reduces the ability to quantify epithelial cells and their precursors. Using the lung cancer/normal lung data set, we present an approach to multidimensional data analysis that consists of artifact removal, identification of classes of cells to be studied further (classifiers) and the measurement of outcome variables on these cell classes. The results of bivariate analysis show a striking similarity between the expression of stem/progenitor markers on lung tumor and adjacent tumor-free lung.

  19. Immune toxicity of TiO₂ under hypoxia in the green-lipped mussel Perna viridis based on flow cytometric analysis of hemocyte parameters.

    Science.gov (United States)

    Wang, Youji; Hu, Menghong; Li, Qiongzhen; Li, Jiale; Lin, Daohui; Lu, Weiqun

    2014-02-01

    The combined effects of DO and TiO2 (mixed rutile/anatase phase, 7/3) on immune responses in Perna viridis were examined. Mussels were exposed to six combinations of oxygen levels (hypoxia: 1.5 mg O2l(-1), normoxia: 6.0 mg O2 l(-1)) and TiO2 concentrations (0, 2.5 mg l(-1) and 10 mg l(-1)) for 216 h. Mussels were sampled after 24h, 48h, 120 h and 216 h, and immune parameters of hemocytes, including mortality, phagocytosis, non-specific esterase, ROS production, lysosomal content and total hemocyte count were investigated using flow cytometric assay. Hemocyte mortality was higher under hypoxia than normoxia, and increased with TiO2 concentrations, but no interaction was found between DO and TiO2. Phagocytosis was reduced under hypoxia and decreased with TiO2 exposure, and the interactive effect between time and TiO2 was observed. The percentage of hemocytes showing non-specific esterase activity was lower under hypoxia, and decreased as TiO2 concentration increased with the significant interactive effect of DO and TiO2. ROS production and lysosomal content were lower under hypoxia and reduced as concentration of TiO2 increased, and interactive effect of DO and TiO2 on ROS was evident. THC was significantly affected by the interactive effect between TiO2 and DO, with higher values under normoxia in the presence of TiO2. The present study demonstrated that immune functions of P. viridis were influenced by both nano-TiO2 and hypoxia with some synergistic effects between the two stressors. This implies that DO has to be considered in the evaluation of the toxicity of nano-materials to bivalves. © 2013.

  20. Equal overall rejection rate in pre-transplant flow-cytometric cross-match negative and positive adult recipients in liver transplantation.

    Science.gov (United States)

    Matinlauri, Irma H; Höckerstedt, Krister A; Isoniemi, Helena M

    2005-10-01

    T cell IgG flow-cytometric cross-matches (FCXM) using 48 stored pre-transplant patient serum samples and 40 stored serum samples collected 3 wk after liver transplantation and frozen spleen cells of cadaveric donors in 48 consecutive liver transplantations were performed retrospectively. T cell IgG FCXM using pre-transplant serum samples was compared with 46 complement-dependent lymphocytotoxic cross-matches (CDCXM) performed at the time of transplantation. Clinical relevance of these tests was evaluated in relation to acute rejection, 1-, 3- and 5-yr graft and patient survival. The incidence of positive FCXM was 33% (16 of 48) and 13% (six of 46) by CDCXM. The median time of acute rejection was 29 d after transplantation in FCXM positive group (range 13-101 d) and 22 d in FCXM negative group (range 7-157 d, NS). Rejection rate was similar in 16 pre-transplant FCXM positive patients (eight of 16, 50%) compared with six pre-transplant CDCXM positive patients (three of six, 50%; NS). Recipients having graft rejection tended to be more often pre-transplant FCXM positive (eight of 21, 38%) than CDCXM positive (three of 21, 14%), but the difference was not significant (p > 0.1). No difference was found in the positive predictive value in relation to acute rejection between positive FCXM and CDCXM (69% vs. 50%; NS). Furthermore there was no correlation between post-transplant positive FCXM and acute rejection. No difference was found between pre-transplant T cell IgG FCXM positive and negative recipients in relation to graft or patient survival. Our findings are supportive for little risk associated with preformed donor-specific antibodies in liver transplantation.

  1. Flow cytometric applications of tumor biology: prospects and pitfalls. [Applications in study of spontaneous dog tumors and in drug and radiation effects on cultured V79 cells

    Energy Technology Data Exchange (ETDEWEB)

    Raju, M.R.; Johnson, T.S.; Tokita, N.; Gillette, E.L.

    1979-01-01

    A brief review of cytometry instrumentation and its potential applications in tumor biology is presented using our recent data. Age-distribution measurements of cells from spontaneous dog tumors and cultured cells after exposure to x rays, alpha particles, or adriamycin are shown. The data show that DNA fluorescence measurements have application in the study of cell kinetics after either radiation or drug treatment. Extensive and careful experimentation is needed to utilize the sophisticated developments in flow cytometry instrumentation.

  2. Multiparameter flow cytometric analysis of CD4 and CD8 T cell subsets in young and old people

    Directory of Open Access Journals (Sweden)

    Özcelik Dennis

    2008-07-01

    Full Text Available Abstract Background T cell-mediated immunity in elderly people is compromised in ways reflected in the composition of the peripheral T cell pool. The advent of polychromatic flow cytometry has made analysis of cell subsets feasible in unprecedented detail. Results Here we document shifts in subset distribution within naïve (N, central memory (CM and effector memory (EM cells defined by CD45RA and CCR7 expression in the elderly, additionally using the costimulatory receptors CD27 and CD28, as well as the coinhibitory receptors CD57 and KLRG-1, to further dissect these. Although differences between young and old were more marked in CD8 than in CD4 cells, a similar overall pattern prevailed in both. Thus, the use of all these markers together, and inclusion of assays of proliferation and cytokine secretion, may enable the construction of a differentiation scheme applicable to CD4 as well as CD8 cells, with the model (based on Romero et al. suggesting the progression N→CM→EM1→EM2→pE1→pE2→EM4→EM3→E end-stage non-proliferative effector cells. Conclusion Overall, the results suggest that both differences in subset distribution and differences between subsets are responsible for age-related changes in CD8 cells but that differences within rather than between subsets are more prominent for CD4 cells.

  3. Protease substrate profiling using bacterial display of self-blocking affinity proteins and flow-cytometric sorting.

    Science.gov (United States)

    Sandersjöö, Lisa; Jonsson, Andreas; Löfblom, John

    2017-01-01

    Proteases are involved in fundamental biological processes and are important tools in both biotechnological and biomedical research. An important property of proteases is to discriminate among potential substrates. Here, a new method for substrate profiling of proteases is presented. The substrates are displayed between two anti-idiotypic affinity domains on the Gram-positive bacterium Staphylococcus carnosus. The first domain functions as a reporter tag and has affinity for a labeled reporter protein, whereas the second domain blocks the reporter tag from interacting with the reporter protein. Site-specific proteolysis of the substrate results in release of the blocking domain, enabling the reporter tag to bind the labeled reporter protein. Proteolysis is therefore reflected in reporter binding, which is quantified by flow cytometry. First, the method with tobacco etch virus protease (TEVp) is evaluated and then the substrate preference of matrix metalloprotease-1 (MMP-1) is determined using two libraries of around three million substrates each. Identified substrate peptides contained the previously reported motif (PXXXHy ) and on-cell determination of apparent kcat /KM revealed that the enriched substrate peptides are hydrolyzed six to eight-fold more efficiently than a previously reported substrate peptide. The method thus works as intended and the authors believe it has potential as an efficient tool for substrate profiling.

  4. Picoplankton Community Composition by CARD-FISH and Flow Cytometric Techniques: A Preliminary Study in Central Adriatic Sea Water

    Directory of Open Access Journals (Sweden)

    Anita Manti

    2012-01-01

    Full Text Available Data concerning picoplanktonic community composition and abundance in the Central Adriatic Sea are presented in an effort to improve the knowledge of bacterioplankton and autotrophic picoplankton and their seasonal changes. Flow cytometry analyses revealed the presence of two distinct bacteria populations: HNA and LNA cells. HNA cells showed an explicit correlation with viable and actively respiring cells. The study of viability and activity may increase our knowledge of the part that contributes really to the remineralization and bacterial biomass production. Authotrophic picoplankton abundance, especially picocyanobacteria, was strongly influenced by seasonality, indicating that light availability and water temperature are very important regulating factors. In terms of total carbon biomass, the main contribution came from heterotrophic bacteria with a lower contribution from autotrophic picoplankton. CARD-FISH evidenced, within the Eubacteria domain, the dominance of members of the phyla Alphaproteobacteria, with a strong contribution from SAR11clade, followed by Cytophaga-Flavobacterium and Gammaproteobacteria. The bacterial groups detected contributed differently depending when the sample was taken, suggesting possible seasonal patterns. This study documents for the first time picoplankton community composition in the Central Adriatic Sea using two different approaches, FCM and CARD-FISH, and could provide preliminary data for future studies.

  5. Bivariate flow cytometric analysis of DNA content versus immunopositivity for ribonucleotide reductase M1 subunit in the cell cycle.

    Science.gov (United States)

    Mangiarotti, R; Bottone, M G; Danova, M; Pellicciari, C

    1998-06-01

    Ribonucleotide reductase (RR) is a cytoplasmatic enzyme catalyzing the reduction of all four ribonucleotides to their corresponding deoxyribonucleotides. Its activity strongly correlates to the rate of DNA synthesis. By using a specific monoclonal antibody against the large M1 subunit of RR, we assessed the expression of M1-RR versus DNA content by dual-parameter flow cytometry. The aim of this paper was to compare the variations in the immunopositivity for M1-RR during the cell cycle to the positivity for other cell cycle markers identifying either proliferating cells (Ki-67 and PCNA) or quiescent cells (statin). To do this, normal human embryonic fibroblasts in different growth conditions as well as several other mammalian cell lines (rat C6 glioma cells; mouse 3T3 fibroblasts and B16 melanoma cells; human epithelial EUE cells and mammary carcinoma MCF-7 cells) were used. The expression of M1-RR antigen was found to correlate positively with the expression of Ki-67 and PCNA, and negatively with the expression of statin. During early G1 phase, M1-RR becomes detectable by specific antibodies relatively later compared to PCNA and Ki-67; therefore, the lack of immunopositivity for M1-RR cannot be taken as an absolute indication of cell quiescence in G0.

  6. Detection of DNA damage in individual cells by flow cytometric analysis using anti-DNA monoclonal antibody

    Energy Technology Data Exchange (ETDEWEB)

    Frankfurt, O.S. (Roswell Park Memorial Institute, Buffalo, NY (USA))

    1987-06-01

    A new method for the measurement of DNA damage in individual cells treated with alkylating agents is described. The method is based on the binding of anti-DNA monoclonal antibody to DNA in situ. Binding of antibody was evaluated by flow cytometry with indirect immunofluorescence. No binding of antibody to DNA in non-treated HeLa S3 cells was detected. Treatment of cells with HN2 or L-phenylalanine mustard induced binding of antibody to DNA in situ. Binding of antibody was observed after treating cells with doses of drugs which reduced the surviving fraction below 20%. Intensity of binding increased in proportion to the drug dose. In HN2-treated cells a cell subset with the lowest antibody binding was observed among cells in G1 phase. Binding of antibody to DNA in HN2-treated cells was eliminated by single-strand (ss) specific S1 nuclease. In competition assay, antibody was inhibited by thermally denatured DNA, but not by native double-stranded (ds) DNA, RNA, nucleosides and deoxyribohomopolymers. Immunoreactivity of cells with the monoclonal antibody F7-26 may be a useful probe for the assessment of cell damage induced by alkylating agents, especially in heterogeneous cell populations.

  7. FLOW CYTOMETRIC DETECTION OF SUBHAPLOID NUCLEI IN HUMAN SPERM AS A MEASURE OF DNA FRAGMENTATION AND APOPTOSIS.

    Science.gov (United States)

    Gröbner, S; Franz, M; Hoberg, U; Wetzka, B; Schweizer, T

    2015-01-01

    The use of assisted reproductive technologies (ARTs) is increasing worldwide. In order to predict the rate of pregnancy after ART the DNA fragmentation index (DFI) of ejaculated spermatocytes may be a better marker than conventional semen quality parameters. Spermatocytes with fragmented DNA are associated with apoptotic stages and are characterized by a low DNA content. The subhaploid nuclei of DNA-damaged spermatocytes can be easily detected by flow cytometry. We here analyzed the percentage of subhaploid nuclei of semen samples from 163 patients aged 26 to 74 years who consulted one of the ten centres for reproductive medicine which routinely send sperm samples to our laboratory in order to determine special sperm parameters. The percentage of subhaploid nuclei indicating the DFI of spermatocytes did not correlate with age and sperm volume, but inversely correlated with sperm concentration and the percentage of motile spermatocytes. This is in concordance with previous studies which demonstrated that DNA damage of spermatozoa correlates with conventional semen quality parameters. Since DNA-damaged spermatocytes are associated with an impaired outcome of assisted conception technologies, this method could help to monitor sperm quality of subfertile men after measures to increase sperm quality and to improve selection criteria of cryopreserved sperm samples in assisted reproduction medicine.

  8. Immunophenotypic Characterization of Human Bone Marrow Mast Cells. A Flow Cytometric Study of Normal and Pathological Bone Marrow Samples

    Directory of Open Access Journals (Sweden)

    Luis Escribano

    1998-01-01

    Full Text Available The goal of the present paper was to define the immunophenotype of bone marrow mast cells (BMMC from healthy controls and patients with hematologic malignancies (HM based on the use of multiple stainings with monoclonal antibodies analyzed by flow cytometry. Our results show that BMMC from both groups of individuals display a similar but heterogenous immunophenotype. The overall numbers of BMMC are higher in the HM group of individuals (p = 0.08. Three patterns of antigen expression were detected: (1 markers constantly positive in all cases analyzed (CD9, CD29, CD33, CD43, CD44, CD49d, CD49e, CD51, CD71, CD117, and FcεRI, (2 antigens that were constantly negative (CD1a, CD2, CD3, CD5, CD6, CD11a, CD14, CD15, CD16, CD19, CD20, CD21, CD23, CD25, CD30, CD34, CD38, CD41a, CD42b, CD65, CD66b, HLA-DR, and CD138, and (3 markers that were positive in a variable proportion of cases – CD11b (50%, CD11c (77%, CD13 (40%, CD18 (20%, CD22 (68%, CD35 (27%, CD40 (67%, CD54 (88% and CD61 (40%. In addition, BMMC from all cases explored were CD45+, and this antigen was expressed at an intensity similar to that of mature granulocytes.

  9. Flow cytometric assessment of reactive oxygen species generations that are directly related to cellular ZnO nanoparticle uptake.

    Science.gov (United States)

    Yoo, Hyun Ju; Yoon, Tae Hyun

    2014-07-01

    In this study, a simple flow cytometry protocol to evaluate nanoparticle associated biological response was proposed. Particularly, we have evaluated the effect of surface charge on the cellular nanoparticle associations and nanoparticle-induced apoptosis. Significant enhancement in side scattering intensity was observed for the HeLa cells treated with positively charged (PLL)ZnO nanoparticles, suggesting that the (PLL)ZnO nanoparticles may induce cell death via adsorption and endocytosis of the nanoparticles. On the other hand, the negatively charged (PAA)ZnO nanoparticle seems to cause cell death process indirectly via the released Zn ions, with less contribution from cellular association of nanoparticles. Time- and dose-dependent studies on cellular association of ZnO nanoparticles, and ZnO associated reactive oxygen species generation were also performed for the HeLa cells exposed to the (PLL)ZnO nanoparticle. For those cells associated with (PLL)ZnO nanoparticle, a significant enhancement in reactive oxygen species generation was observed even at a lower concentration (10 ppm), which was not observable for the results with the whole cell population. By using this approach, we are able to distinguish biological responses (e.g., reactive oxygen species (ROS) generation) directly related to the cellular associations of NPs from those indirectly related to the cellular associations of NPs, such as the cytotoxicity caused by the NP released metal ions.

  10. Ultrastructural and flow cytometric analyses of lipid accumulation in microalgae: Annual report, Solar Energy Research Institute, Aquatic Species Program

    Energy Technology Data Exchange (ETDEWEB)

    Solomon, J.A.; Hand, R.E. Jr.; Mann, R.C.

    1986-01-01

    Lipid accumulation in three species of microalgae was investigated with flow cytometry (FCM) and transmission electron microscopy (TEM). Previous studies using batch cultures of algae have led to the assumption that lipid accumulation in microalgae is a gradual process requiring at least several days for completion. However, FCM reveals, through changes in the chlorophyll:lipid ratio, that the time span required for individual cells to change metabolic state is short. Simultaneous FCM measurements of chlorophyll and nile red (neutral lipid) fluorescence in individual cells of nitrogen-deficient Isochrysis populations revealed a bimodal population distribution as one stage in the lipid accumulation process. The fact that two discrete populations exist, with few cells in an intermediate stage, suggests rapid response to a lipid trigger. Interpretations of light and electron microscopic observations are consistent with this hypothesis. The time required for an entire population to achieve maximum lipid content is considerably longer than that required for a single cell, due to the variation in response time among cells. In this study high lipid cultures were sometimes obtained by using FCM to separate high lipid cells from the remainder of the population. FCM holds much promise for strain enhancement but considerable developmental work, directed at providing more consistent results, remains to be done. 8 refs., 33 figs.

  11. Flow cytometric analysis of the H2O2-induced increase in intracellular Ca2+ concentration of rat thymocytes.

    Science.gov (United States)

    Okazaki, E; Chikahisa, L; Kanemaru, K; Oyama, Y

    1996-08-01

    The effect of hydrogen peroxide (H2O2) on the intracellular Ca2+ concentration ([Ca2+]i) of rat thymocytes was examined by a flow cytometer and two fluorescent dyes, fluo-3-AM and ethidium bromide, a dye impermeant to intact membranes, to characterize the H2O2-induced increase in [Ca2+]i. H2O2 at concentrations greater than 30 microM dose-dependently increased the [Ca2+]i of thymocytes which were not stained with ethidium. Removal of external Ca2+ greatly reduced the degree of H2O2-induced increase in [Ca2+]i. However, H2O2 still increased the [Ca2+]i under the external Ca(2+)-free condition. Diethylmaleate, which is known to produce a chemical depletion of cellular nonprotein thiol, significantly increased the [Ca2+]i. Dithiothreitol, which is used to protect cellular nonprotein thiol, slightly decreased the [Ca2+]i, but greatly reduced the H2O2-induced increase in [Ca2+]i. Therefore, it is considered that H2O2 may increase the [Ca2+]i through a mechanism related to the effects of H2O2 on the cellular nonprotein thiol.

  12. Quality control of flow cytometric immunophenotyping%流式细胞术表型分析的质量控制

    Institute of Scientific and Technical Information of China (English)

    吴丽娟; 许东升

    2011-01-01

    FCM表型分析已经从外周血淋巴细胞的双参数定量测定发展到当今对血液细胞病理学包括骨髓、淋巴结分析的5个或更多参数的定性测定,其中白血病和淋巴瘤的免疫表型分析在血液淋巴系统恶性疾病诊断、分类及病情监测上都是对形态学检验的一种重要补充.FCM 5个或更多参数分析的复杂性以及数据的解释依赖于仪器、试剂、程序的标准化及校准.此外,在美国的流式细胞学实验室还必须保留有关水平测试、样品制备、方法学的准确性、特异性、灵敏度和精密度的相关资料.CLSI和UCCC建议每个流式细胞学实验室需要对定性和定量检测程序进行验证.本文对美国临床流式细胞学实验室FCM检验的相关验证、室内与室间质量控制措施进行介绍.%Flow cytometric immunophenotyping has evolved from two-parameter quantitative measurement of peripheral blood lymphocytes to five-or more parameter qualitative evaluation of bone marrow and lymph node in hematopathology.Leukemia/lymphoma immunophenotyping represents an important addition to histomorphology in the diagnosis,classification and monitoring of hematolymphoid neoplasms.The complexity of five-or more parameter analyses and the interpretation of the data rely on standardization and validation of the instrument,the reagent and the procedure.In addition,clinical flow cytometry laboratories in U.S.A are required to document proficiency testing,sample preparation as well as accuracy,specificity,sensitivity and precision of methodology.CLSI and UCCC recommend that each laboratory should validate its own qualitative and quantitative procedures.This paper introduces the procedures for validation and quality control in a clinical flow cytometry laboratory in the United States.

  13. Flow cytometric analysis of the inhibition of human basophil activation by histamine high dilutions – a replication study

    Directory of Open Access Journals (Sweden)

    Chantal Wälchli

    2012-09-01

    perform in order to reduce the possibility of artifacts but was omitted in the former study. Conclusions: Laboratory independent replication of homeopathic basic research experiments is still a challenge. Assuming that the results formerly obtained with this model were not due to systematic errors, the quest identifying the crucial factors for successful reproducibility is open for future research. Keywords: Human basophils; histamine; high dilutions; flow cytometry Reference: [1] Sainte-Laudy J, Belon P. Improvement of flow cytometric analysis of basophil activation inhibition by high histamine dilutions. A novel basophil specific marker: CD 203c. Homeopathy. 2006;95:3-8.

  14. Evaluation of zinc oxide nanoparticles toxicity on marine algae chlorella vulgaris through flow cytometric, cytotoxicity and oxidative stress analysis.

    Science.gov (United States)

    Suman, T Y; Radhika Rajasree, S R; Kirubagaran, R

    2015-03-01

    The increasing industrial use of nanomaterials during the last decades poses a potential threat to the environment and in particular to organisms living in the aquatic environment. In the present study, the toxicity of zinc oxide nanoparticles (ZnO NPs) was investigated in Marine algae Chlorella vulgaris (C. vulgaris). High zinc dissociation from ZnONPs, releasing ionic zinc in seawater, is a potential route for zinc assimilation and ZnONPs toxicity. To examine the mechanism of toxicity, C. vulgaris were treated with 50mg/L, 100mg/L, 200mg/L and 300 mg/L ZnO NPs for 24h and 72h. The detailed cytotoxicity assay showed a substantial reduction in the viability dependent on dose and exposure. Further, flow cytometry revealed the significant reduction in C. vulgaris viable cells to higher ZnO NPs. Significant reductions in LDH level were noted for ZnO NPs at 300 mg/L concentration. The activity of antioxidant enzyme superoxide dismutase (SOD) significantly increased in the C. vulgaris exposed to 200mg/L and 300 mg/L ZnO NPs. The content of non-enzymatic antioxidant glutathione (GSH) significantly decreased in the groups with a ZnO NPs concentration of higher than 100mg/L. The level of lipid peroxidation (LPO) was found to increase as the ZnO NPs dose increased. The FT-IR analyses suggested surface chemical interaction between nanoparticles and algal cells. The substantial morphological changes and cell wall damage were confirmed through microscopic analyses (FESEM and CM).

  15. Applications of immuno-magnetic bead and immunofluorescent flow cytometric techniques for the quantitative detection of HAB microalgae

    Institute of Scientific and Technical Information of China (English)

    HUANG Jian; WEN Ruobing; BAO Zhenmin; SUI Zhenghong; SUN Ningbo; KANG Kyoungho

    2012-01-01

    Over the last severaldecades,harmful algal blooms (HABs) havebecome a serious environmental problem in many parts of the world.A rapid and accurate detection process for HAB algae has yet to be developed.Heterosigma akashiwo is one of the most important HABs species in China.The objective of this study was to develop an immunologic technique that can rapidly and sensitively count H.akashiwo cells.Five HABs species (Alexandrium catenella,Thalassiosira sp.,Cryptomonas sp.,Alexandrium tamarense and Symbiodinium sp.,) were used in this study to evaluate the analysis process we developed.A polyclonal antibody with high titers against H.akashiwo was obtained by injecting H.akashiwo cells into rabbits.Immuno-magnetic beads (IMB) were produced via conjugated polyclonal antibodies with magnetic beads and applied to isolate and count H.akashiwo cells from the culture.Results show that 66.7%-91.6% of the cells were captured from unialgal culture by IMBs,and only 5.3%-12.5% of the four other HAB microalgae species were captured,indicating that the constructed IMBs combined specifically with the H.akashiwo cells.At the same time,flow cytometry (FCM) sorting was exploited to screen H.akashiwo cells after labeling with FITC conjugated polyclonal antibodies.Using the FCM technique,91.7% of the targeted cells were sorted out from mixed microalgae samples in just a few minutes.These results indicate that both antibody-involved IMB and antibody-based FCM techniques are highly effective at detecting and quantifying HAB species.These techniques,especially immuno-magnetic separation,have low associated cost,and are fast and simple processes compared with other techniques currently in use.

  16. Flow-cytometric analysis of T-lymphocyte subsets after different treatment methods in patients with pericoronitis.

    Science.gov (United States)

    Orbak, Recep; Dayi, Ertunç

    2003-02-01

    The aim of this study was to determine whether there was any change in T-lymphocyte subsets in patients with periocoronitis after the application of different treatment methods. Twenty-six patients with acute pericoronitis were included in the study. In every phase of the treatment (pretreatment, postcurettage, and postextraction), the biopsy samples were taken from the gingival tissues at sites of pericoronitis. Then, CD4(+) and CD8(+) lymphocyte and CD4(+)/CD8(+) ratio values were determined using flow cytometry in the biopsy samples. At the same time, gingival index (Löe-Silness) and plaque index (Silness-Löe) scores were recorded to assess the periodontal status in patients. To determine the correlation between the clinical measurements and the laboratory results obtained before the treatment, after curettage, and after extraction, we conducted an analysis using a paired t-test. The normal values in peripheral blood of CD4(+) and CD8(+) lymphocytes are 25% to 29% and 19% to 48%, respectively. However, the CD4(+) and CD8(+) lymphocyte values in the patients with acute pericoronitis were found to be 22.12% +/- 6.15% and 7.69% +/- 4.12%, respectively. These values are lower than the normal values. The CD4(+) lymphocyte value increased to 31.06% +/- 7.09% postcurettage and to 32.24% +/- 3.11% postextraction. The CD8(+) lymphocyte value increased to 16.21% +/- 5.27% postcurettage and to 18.25% +/- 3.13% postextraction. The CD4/CD8 ratio increased postcurettage and postextraction. This increase was statistically significant (P pericoronitis pathobiology.

  17. Modeling of inter-sample variation in flow cytometric data with the joint clustering and matching procedure.

    Science.gov (United States)

    Lee, Sharon X; McLachlan, Geoffrey J; Pyne, Saumyadipta

    2016-01-01

    We present an algorithm for modeling flow cytometry data in the presence of large inter-sample variation. Large-scale cytometry datasets often exhibit some within-class variation due to technical effects such as instrumental differences and variations in data acquisition, as well as subtle biological heterogeneity within the class of samples. Failure to account for such variations in the model may lead to inaccurate matching of populations across a batch of samples and poor performance in classification of unlabeled samples. In this paper, we describe the Joint Clustering and Matching (JCM) procedure for simultaneous segmentation and alignment of cell populations across multiple samples. Under the JCM framework, a multivariate mixture distribution is used to model the distribution of the expressions of a fixed set of markers for each cell in a sample such that the components in the mixture model may correspond to the various populations of cells, which have similar expressions of markers (that is, clusters), in the composition of the sample. For each class of samples, an overall class template is formed by the adoption of random-effects terms to model the inter-sample variation within a class. The construction of a parametric template for each class allows for direct quantification of the differences between the template and each sample, and also between each pair of samples, both within or between classes. The classification of a new unclassified sample is then undertaken by assigning the unclassified sample to the class that minimizes the distance between its fitted mixture density and each class density as provided by the class templates. For illustration, we use a symmetric form of the Kullback-Leibler divergence as a distance measure between two densities, but other distance measures can also be applied. We show and demonstrate on four real datasets how the JCM procedure can be used to carry out the tasks of automated clustering and alignment of cell

  18. Improved flow cytometric method to enumerate residual cells: minimal linear detection limits for platelets, erythrocytes, and leukocytes.

    Science.gov (United States)

    Pichler, J; Printz, D; Scharner, D; Trbojevic, D; Siekmann, J; Fritsch, G

    2002-08-15

    Increasing demand for quality control of blood products requires more sensitive methods to enumerate residual cells. Presently, the reported threshold (in cells per microliter) is 400 for red blood cells, 30-500 for platelets, and 1 for leukocytes. To examine precision and linearity in enumerating residual platelets and red blood cells, EDTA-anticoagulated blood from healthy donors was serially diluted with serum, stained in TruCount tubes using a no-lyse/no-wash procedure and a monoclonal antibody cocktail against the CD42a (FL1) and glycophorin-A (FL2) epitopes, and analyzed by flow cytometry. Leukocyte counts were determined in separate tubes. Cell preparation and analysis were performed once for 20 blood samples each and 20 times using the same specimen. Acquisition from the same tube was performed separately for platelets (threshold on FL1) and red blood cells (threshold on FL2). Multiparameter analysis was used for data evaluation. Linear results were obtained for platelets per microliter between 3,410 and 5 and for red blood cells per microliter between 54,000 and 3. For the lower cell concentrations, the coefficient of variation was 16.7% for platelets and 10.9% for red blood cells. The presented method allows the distinction between physiologically intact and ghost red blood cells. The method represents a reliable, sensitive, and accurate approach to quantify platelets and red blood cells in diluted blood. It can be applied to enumerate residual cells in plasma products and meets the increasing demand for quality control in blood components.

  19. Applications of immuno-magnetic bead and immunofluorescent flow cytometric techniques for the quantitative detection of HAB microalgae

    Science.gov (United States)

    Huang, Jian; Wen, Ruobing; Bao, Zhenmin; Sui, Zhenghong; Sun, Ningbo; Kang, Kyoungho

    2012-05-01

    Over the last several decades, harmful algal blooms (HABs) have become a serious environmental problem in many parts of the world. A rapid and accurate detection process for HAB algae has yet to be developed. Heterosigma akashiwo is one of the most important HABs species in China. The objective of this study was to develop an immunologic technique that can rapidly and sensitively count H. akashiwo cells. Five HABs species ( Alexandrium catenella, Thalassiosira sp., Cryptomonas sp., Alexandrium tamarense and Symbiodinium sp.), were used in this study to evaluate the analysis process we developed. A polyclonal antibody with high titers against H. akashiwo was obtained by injecting H. akashiwo cells into rabbits. Immuno-magnetic beads (IMB) were produced via conjugated polyclonal antibodies with magnetic beads and applied to isolate and count H. akashiwo cells from the culture. Results show that 66.7%-91.6% of the cells were captured from unialgal culture by IMBs, and only 5.3%-12.5% of the four other HAB microalgae species were captured, indicating that the constructed IMBs combined specifically with the H. akashiwo cells. At the same time, flow cytometry (FCM) sorting was exploited to screen H. akashiwo cells after labeling with FITC conjugated polyclonal antibodies. Using the FCM technique, 91.7% of the targeted cells were sorted out from mixed microalgae samples in just a few minutes. These results indicate that both antibody-involved IMB and antibody-based FCM techniques are highly effective at detecting and quantifying HAB species. These techniques, especially immuno-magnetic separation, have low associated cost, and are fast and simple processes compared with other techniques currently in use.

  20. Flow cytometric analysis on tri-n-butyltin-induced increase in annexin V binding to membranes of rat thymocytes.

    Science.gov (United States)

    Nakata, M; Oyama, Y; Okada, Y; Yamazaki, Y; Chikahisa, L; Satoh, M

    1999-10-01

    Effects of tri-n-butyltin chloride (TBT) on rat thymocytes were examined by using a flow cytometer and three fluorescent dyes (annexin V-FITC, ethidium bromide and fluo-3-AM) to further characterize its cytotoxic action. TBT at concentrations of 100 nM or greater, time- and dose-dependently increased the population of annexin V-positive live cells in the cell suspension. Most of cells became to be annexin V-positive within 60 min after the start of application of 300 nM TBT. Some of annexin V-positive live cells were further stained with ethidium, indicating that some of the cells were killed, in continued presence of TBT at 300 nM or greater. When the cells were exposed to 300 nM TBT only for 15 min, the population of annexin V-positive live cells increased after removal of TBT from incubation medium. TBT-induced increase in the population of annexin V-positive live cells was partly attenuated under Ca(2+)-free condition, although that was not the case for the dead cells. TBT at 30 nM or greater increased [Ca(2+)]i in a dose-dependent manner. Triethyltin and trimethyltin even at 1 μM did not increase the [Ca(2+)]i and the population of annexin V-positive live cells. The population of annexin V-positive live cells increased as the [Ca(2+)]i was increased by ionomycin, a calcium ionophore. Results suggest an involvement of Ca(2+) in some of TBT-induced cytotoxicity.

  1. Radio-protective effect of vitamin E on spermatogenesis in mice exposed to γ-irradiation: a flow cytometric study

    Institute of Scientific and Technical Information of China (English)

    C.Songthaveesin; J.Saikhun; Y.Kitiyanant; K.Pavasuthipaisit

    2004-01-01

    Aim: To investigate the effect of vitamin E on the radioprotection of spermatogenesis and chromatin condensation of spermatozoa during passage through the epididymis in mice exposed to irradiation. Methods: Adult outbred male ICR mice were orally administered natural vitamin E (VE,D-a-tocopheryl acetate) at 400 IU/kg for 7 days before exposure to 1 Gy of y-irradiation. The animals were sacrificed at day 1,7,14,21, 28, 35 and 70 post-irradiation (IR) and the percentage of testicular germ cells and epididymal sperm chromatin condensation was analyzed using flow cytometry. Results: Serum D-a-tocopheryl acetate levels were 47.4±3.2μg/dL in the treatedgroup, yet it could not be detected in the control group. The testicular weight of irradiated mice pretreated with VE+IR was significantly (P<0.05) higher than that of those without VE treatment (IR) at day 14 and 21 post-irradiation. The percentage of primary spermatocytes (4C) in the VE+IR group was comparable to the controls but significantly (P<0.05) higher than those in the IR group from day 7 to 35 post-irradiation. The percentage of round spermatids (1C) in the VE+IR group was also significantly (P<0.05) higher than those in the IR group at day 28 postirradiation. The primary spermatocytes:spermatogonia ratio in the IR group was significantly (P<0.05) declined at day 7 to 35 post-irradiation when compared to the VE+IR and control groups. The round spermatid:spermatogonia ratio in the VE+IR group was significantly (P<0.05) higher than that of the IR group at day 14 and 28 post-irradiation.The chromatin condensation of epididymal spermatozoa measured by propidium iodide uptake was not affected by 1 Gy of γ-irradiation. Conclusion: The administration of VE prior to irradiation protects spermatogenic cells fromradiation. (Asian J Androl 2004 Dec; 6: 331-336)

  2. Breakdown parameter for kinetic modeling of multiscale gas flows.

    Science.gov (United States)

    Meng, Jianping; Dongari, Nishanth; Reese, Jason M; Zhang, Yonghao

    2014-06-01

    Multiscale methods built purely on the kinetic theory of gases provide information about the molecular velocity distribution function. It is therefore both important and feasible to establish new breakdown parameters for assessing the appropriateness of a fluid description at the continuum level by utilizing kinetic information rather than macroscopic flow quantities alone. We propose a new kinetic criterion to indirectly assess the errors introduced by a continuum-level description of the gas flow. The analysis, which includes numerical demonstrations, focuses on the validity of the Navier-Stokes-Fourier equations and corresponding kinetic models and reveals that the new criterion can consistently indicate the validity of continuum-level modeling in both low-speed and high-speed flows at different Knudsen numbers.

  3. Flow cytometric evaluation of the contribution of ionic silver to genotoxic potential of nanosilver in human liver HepG2 and colon Caco2 cells.

    Science.gov (United States)

    Sahu, Saura C; Njoroge, Joyce; Bryce, Steven M; Zheng, Jiwen; Ihrie, John

    2016-04-01

    Exposure to nanosilver found in food- and cosmetics-related consumer products is of public concern because of the lack of information about its safety. In this study, two widely used in vitro cell culture models, human liver HepG2 and colon Caco2 cells, and the flow cytometric micronucleus (FCMN) assay were evaluated as tools for rapid predictive screening of the potential genotoxicity of nanosilver. Recently, we reported the genotoxicity of 20 nm nanosilver using these systems. In the current study presented here, we tested the hypothesis that the nanoparticle size and cell types were critical determinants of its genotoxicity. To test this hypothesis, we used the FCMN assay to evaluate the genotoxic potential of 50 nm nanosilver of the same shape, composition, surface charge and obtained from the same commercial source using the same experimental conditions and in vitro models (HepG2 and Caco2) as previously tested for the 20 nm silver. Results of our study show that up to the concentrations tested in these cultured cell test systems, the smaller (20 nm) nanoparticle is genotoxic to both the cell types by inducing micronucleus (MN). However, the larger (50 nm) nanosilver induces MN only in HepG2 cells, but not in Caco2 cells. Also in this study, we evaluated the contribution of ionic silver to the genotoxic potential of nanosilver using silver acetate as the representative ionic silver. The MN frequencies in HepG2 and Caco2 cells exposed to the ionic silver in the concentration range tested are not statistically significant from the control values except at the top concentrations for both the cell types. Therefore, our results indicate that the ionic silver may not contribute to the MN-forming ability of nanosilver in HepG2 and Caco2 cells. Also our results suggest that the HepG2 and Caco2 cell cultures and the FCMN assay are useful tools for rapid predictive screening of a genotoxic potential of food- and cosmetics-related chemicals including nanosilver.

  4. Kinetic derivation of a Hamilton-Jacobi traffic flow model

    CERN Document Server

    Borsche, Raul; Kimathi, Mark

    2012-01-01

    Kinetic models for vehicular traffic are reviewed and considered from the point of view of deriving macroscopic equations. A derivation of the associated macroscopic traffic flow equations leads to different types of equations: in certain situations modified Aw-Rascle equations are obtained. On the other hand, for several choices of kinetic parameters new Hamilton-Jacobi type traffic equations are found. Associated microscopic models are discussed and numerical experiments are presented discussing several situations for highway traffic and comparing the different models.

  5. Rapid Multiplexed Flow Cytometric Assay for Botulinum Neurotoxin Detection Using an Automated Fluidic Microbead-Trapping Flow Cell for Enhanced Sensitivity

    Energy Technology Data Exchange (ETDEWEB)

    Ozanich, Richard M.; Bruckner-Lea, Cindy J.; Warner, Marvin G.; Miller, Keith D.; Antolick, Kathryn C.; Marks, James D.; Lou, Jianlong; Grate, Jay W.

    2009-07-15

    A bead-based sandwich immunoassay for botulinum neurotoxin serotype A (BoNT/A) has been developed and demonstrated using a recombinant 50 kDa fragment (BoNT/A-HC-fragment) of the BoNT/A heavy chain (BoNT/A-HC) as a structurally valid simulant. Three different anti-BoNT/A antibodies were attached to three different fluorescent dye encoded flow cytometry beads for multiplexing. The assay was conducted in two formats: a manual microcentrifuge tube format and an automated fluidic system format. Flow cytometry detection was used for both formats. The fluidic system used a novel microbead-trapping flow cell to capture antibody-coupled beads with subsequent sequential perfusion of sample, wash, dye-labeled reporter antibody, and final wash solutions. After the reaction period, the beads were collected for analysis by flow cytometry. Sandwich assays performed on the fluidic system gave median fluorescence intensity signals on the flow cytometer that were 2-4 times higher than assays performed manually in the same amount of time. Limits of detection were estimated at 1 pM (~50 pg/mL for BoNT/A-HC-fragment) for the 15 minute fluidic assay.

  6. Spectral kinetic energy transfer in turbulent premixed reacting flows.

    Science.gov (United States)

    Towery, C A Z; Poludnenko, A Y; Urzay, J; O'Brien, J; Ihme, M; Hamlington, P E

    2016-05-01

    Spectral kinetic energy transfer by advective processes in turbulent premixed reacting flows is examined using data from a direct numerical simulation of a statistically planar turbulent premixed flame. Two-dimensional turbulence kinetic-energy spectra conditioned on the planar-averaged reactant mass fraction are computed through the flame brush and variations in the spectra are connected to terms in the spectral kinetic energy transport equation. Conditional kinetic energy spectra show that turbulent small-scale motions are suppressed in the burnt combustion products, while the energy content of the mean flow increases. An analysis of spectral kinetic energy transfer further indicates that, contrary to the net down-scale transfer of energy found in the unburnt reactants, advective processes transfer energy from small to large scales in the flame brush close to the products. Triadic interactions calculated through the flame brush show that this net up-scale transfer of energy occurs primarily at spatial scales near the laminar flame thermal width. The present results thus indicate that advective processes in premixed reacting flows contribute to energy backscatter near the scale of the flame.

  7. A Kinetic Model for Vapor-liquid Flows

    Science.gov (United States)

    2005-07-13

    A Kinetic Model for Vapor-liquid Flows Aldo Frezzotti, Livio Gibelli and Silvia Lorenzani Dipartimento di Matematica del Politecnico di Milano Piazza...ES) Dipartimento di Matematica del Politecnico di Milano Piazza Leonardo da Vinci 32 - 20133 Milano - Italy 8. PERFORMING ORGANIZATION REPORT NUMBER

  8. Multiphase Flow and Fluidization Continuum and Kinetic Theory Descriptions

    CERN Document Server

    Gidaspow, Dimitri

    1994-01-01

    Useful as a reference for engineers in industry and as an advanced level text for graduate engineering students, Multiphase Flow and Fluidization takes the reader beyond the theoretical to demonstrate how multiphase flow equations can be used to provide applied, practical, predictive solutions to industrial fluidization problems. Written to help advance progress in the emerging science of multiphase flow, this book begins with the development of the conservation laws and moves on through kinetic theory, clarifying many physical concepts (such as particulate viscosity and solids pressure) and i

  9. Micellar kinetics of a fluorosurfactant through stopped-flow NMR.

    Science.gov (United States)

    Yushmanov, Pavel V; Furó, István; Stilbs, Peter

    2006-02-28

    19F NMR chemical shifts and transverse relaxation times T2 were measured as a function of time after quick stopped-flow dilution of aqueous solutions of sodium perfluorooctanoate (NaPFO) with water. Different initial concentrations of micellar solution and different proportions of mixing were tested. Previous stopped-flow studies by time-resolved small-angle X-ray scattering (TR-SAXS) detection indicated a slow (approximately 10 s) micellar relaxation kinetics in NaPFO solutions. In contrast, no evidence of any comparable slow (>100 ms) relaxation process was found in our NMR studies. Possible artifacts of stopped-flow experiments are discussed as well as differences between NMR and SAXS detection methods. Upper bounds on the relative weight of a slow relaxation process are given within existing kinetic theories of micellar dissolution.

  10. Cytometric analysis of mammalian sperm for induced morphologic and DNA content errors

    Energy Technology Data Exchange (ETDEWEB)

    Pinkel, D.

    1983-06-27

    Some flow-cytometric and image analysis procedures under development for quantitative analysis of sperm morphology are reviewed. The results of flow-cytometric DNA-content measurements on sperm from radiation exposed mice are also summarized, the results related to the available cytological information, and their potential dosimetric sensitivity discussed. (ACR)

  11. On the kinetic energy of the divergent and nondivergent flow in the atmosphere

    OpenAIRE

    Chen, Tsing-Chang; Wiin-Nielsen, Aksel C.

    2011-01-01

    The kinetic energy of horizontal flow in a hydrostatic atmosphere is divided into the kinetic energies of its divergent and nondivergent components. The law of conversion between these two energies for large-scale flows in the atmosphere is derived and discussed using balanced and unbalanced models of circulations in the atmosphere. It is shown that the total potential energy is converted into the kinetic energy of the divergent flow which, in turn, is converted into the kinetic energy of the...

  12. Chemistry Resolved Kinetic Flow Modeling of TATB Based Explosives

    Energy Technology Data Exchange (ETDEWEB)

    Vitello, P A; Fried, L E; Howard, W M; Levesque, G; Souers, P C

    2011-07-21

    Detonation waves in insensitive, TATB based explosives are believed to have multi-time scale regimes. The initial burn rate of such explosives has a sub-microsecond time scale. However, significant late-time slow release in energy is believed to occur due to diffusion limited growth of carbon. In the intermediate time scale concentrations of product species likely change from being in equilibrium to being kinetic rate controlled. They use the thermo-chemical code CHEETAH linked to an ALE hydrodynamics code to model detonations. They term their model chemistry resolved kinetic flow as CHEETAH tracks the time dependent concentrations of individual species in the detonation wave and calculates EOS values based on the concentrations. A HE-validation suite of model simulations compared to experiments at ambient, hot, and cold temperatures has been developed. They present here a new rate model and comparison with experimental data.

  13. On the drift kinetic equation driven by plasma flows

    Energy Technology Data Exchange (ETDEWEB)

    Shaing, K C [Plasma and Space Science Center and ISAPS, National Cheng Kung University, Tainan 70101, Taiwan (China); Department of Engineering Physics, University of Wisconsin, Madison, WI 53706 (United States)

    2010-07-15

    A drift kinetic equation that is driven by plasma flows has previously been derived by Shaing and Spong 1990 (Phys. Fluids B 2 1190). The terms that are driven by particle speed that is parallel to the magnetic field B have been neglected. Here, such terms are discussed to examine their importance to the equation and to show that these terms do not contribute to the calculations of plasma viscosity in large aspect ratio toroidal plasmas, e.g. tokamaks and stellarators. (brief communication)

  14. Reaction kinetics of fluorite in flow systems and surface chemistry

    Institute of Scientific and Technical Information of China (English)

    张荣华; 胡书敏

    1996-01-01

    The kinetic experiments of fluorite in water-HCl solution in an open-flow system at the temperatures ≤100℃ reveal that the variation of flow rate (U) can change the reaction rate orders from 0 to 2 or higher. In the far from equilibrium systems, the dissolution rates of fluorite in aqueous solutions have a zero order.The reaction rates are controlled by pH values of input solutions. In fact, the reaction rates are related to the concentrations of the active sites occupied by H+ on fluorite surface [SOH]. X-ray photospectroscopy observations on fluorite surface before and after reaction indicate that surface chemical processes control the reaction rates: Cl- cations attach on and enter into surface of fluorite besides H+ when fluorites react with HCl solutions, which affect the reaction rates.

  15. Kinetic assessment of measured mass flow rates and streamwise pressure distributions in microchannel gas flows

    Institute of Scientific and Technical Information of China (English)

    Jing Fan; Chong Xie; Jianzheng Jiang

    2007-01-01

    Measured mass flow rates and streamwise pressure distributions of gas flowing through microchannels were reported by many researchers. Assessment of these data is crucial before they are used in the examination of slip models and numerical schemes, and in the design of microchannel elements in various MEMS devices. On the basis of kinetic solutions of the mass flow rates and pressure distributions in microchannel gas flows, the measured data available are properly normalized and then are compared with each other. The 69 normalized data of measured pressure distributions are in excellent agreement, and 67 of them are within 1 ± 0.05. The normalized data of mass flow-rates ranging between 0.95 and 1 agree well with each other as the inlet Knudsen number Kni > 0.02, but they scat ter between 0.85 and 1.15 as Kni < 0.02 with, to some extent, a very interesting bifurcation trend.

  16. Fibrillization kinetics of insulin solution in an interfacial shearing flow

    Science.gov (United States)

    Balaraj, Vignesh; McBride, Samantha; Hirsa, Amir; Lopez, Juan

    2015-11-01

    Although the association of fibril plaques with neurodegenerative diseases like Alzheimer's and Parkinson's is well established, in-depth understanding of the roles played by various physical factors in seeding and growth of fibrils is far from well known. Of the numerous factors affecting this complex phenomenon, the effect of fluid flow and shear at interfaces is paramount as it is ubiquitous and the most varying factor in vivo. Many amyloidogenic proteins have been found to denature upon contact at hydrophobic interfaces due to the self-assembling nature of protein in its monomeric state. Here, fibrillization kinetics of insulin solution is studied in an interfacial shearing flow. The transient surface rheological response of the insulin solution to the flow and its effect on the bulk fibrillization process has been quantified. Minute differences in hydrophobic characteristics between two variants of insulin- Human recombinant and Bovine insulin are found to result in very different responses. Results presented will be in the form of fibrillization assays, images of fibril plaques formed, and changes in surface rheological properties of the insulin solution. The interfacial velocity field, measured from images (via Brewster Angle Microscopy), is compared with computations. Supported by NNX13AQ22G, National Aeronautics and Space Administration.

  17. High-Fidelity Kinetics and Radiation Transport for NLTE Hypersonic Flows Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The modeling of NLTE hypersonic flows combines several disciplines: chemistry, kinetics, radiation transport, fluid mechanics, and surface science. No single code or...

  18. Flow cytometric readout based on Mitotracker Red CMXRos staining of live asexual blood stage malarial parasites reliably assesses antibody dependent cellular inhibition

    DEFF Research Database (Denmark)

    Jogdand, Prajakta S; Singh, Susheel K; Christiansen, Michael;

    2012-01-01

    ABSTRACT: BACKGROUND: Functional in vitro assays could provide insights into the efficacy of malaria vaccine candidates. For estimating the anti-parasite effect induced by a vaccine candidate, an accurate determination of live parasite count is an essential component of most in vitro bioassays...... asynchronous and tightly synchronized asexual blood stage cultures of Plasmodium falciparum were stained with CMXRos and subjected to detection by flow cytometry and fluorescence microscopy. The parasite counts obtained by flow cytometry were compared to standard microscopic counts obtained through examination...

  19. Colour-encoded paramagnetic microbead-based direct inhibition triplex flow cytometric immunoassay for ochratoxin A, fumonisins and zearalenone in cereals and cereal-based feed

    NARCIS (Netherlands)

    Peters, J.; Thomas, D.; Boers, E.A.M.; Rijk, de T.C.; Berthiller, F.; Haasnoot, W.; Nielen, M.W.F.

    2013-01-01

    A combined (triplex) immunoassay for the simultaneous detection of three mycotoxins in grains was developed with superparamagnetic colour-encoded microbeads, in combination with two bead-dedicated flow cytometers. Monoclonal antibodies were coupled to the beads, and the amounts of bound mycotoxins

  20. In-depth validation of acridine orange staining for flow cytometric parasite and reticulocyte enumeration in an experimental model using Plasmodium berghei

    DEFF Research Database (Denmark)

    Hein-Kristensen, L; Wiese, L; Kurtzhals, J A L

    2009-01-01

    Flow cytometry is potentially an effective method for counting malaria parasites, but inconsistent results have hampered its routine use in rodent models. A published two-channel method using acridine orange offers clear discrimination between the infected and uninfected erythrocytes. However, pr...

  1. Colour-encoded paramagnetic microbead-based direct inhibition triplex flow cytometric immunoassay for ochratoxin A, fumonisins and zearalenone in cereals and cereal-based feed

    NARCIS (Netherlands)

    Peters, J.; Thomas, D.; Boers, E.A.M.; Rijk, de T.C.; Berthiller, F.; Haasnoot, W.; Nielen, M.W.F.

    2013-01-01

    A combined (triplex) immunoassay for the simultaneous detection of three mycotoxins in grains was developed with superparamagnetic colour-encoded microbeads, in combination with two bead-dedicated flow cytometers. Monoclonal antibodies were coupled to the beads, and the amounts of bound mycotoxins w

  2. Flow cytometric readout based on Mitotracker Red CMXRos staining of live asexual blood stage malarial parasites reliably assesses antibody dependent cellular inhibition

    Directory of Open Access Journals (Sweden)

    Jogdand Prajakta S

    2012-07-01

    Full Text Available Abstract Background Functional in vitro assays could provide insights into the efficacy of malaria vaccine candidates. For estimating the anti-parasite effect induced by a vaccine candidate, an accurate determination of live parasite count is an essential component of most in vitro bioassays. Although traditionally parasites are counted microscopically, a faster, more accurate and less subjective method for counting parasites is desirable. In this study mitochondrial dye (Mitotracker Red CMXRos was used for obtaining reliable live parasite counts through flow cytometry. Methods Both asynchronous and tightly synchronized asexual blood stage cultures of Plasmodium falciparum were stained with CMXRos and subjected to detection by flow cytometry and fluorescence microscopy. The parasite counts obtained by flow cytometry were compared to standard microscopic counts obtained through examination of Giemsa-stained thin smears. A comparison of the ability of CMXRos to stain live and compromised parasites (induced by either medium starvation or by anti-malarial drug treatment was carried out. Finally, parasite counts obtained by CMXRos staining through flow cytometry were used to determine specific growth inhibition index (SGI in an antibody-dependent cellular inhibition (ADCI assay. Results Mitotracker Red CMXRos can reliably detect live intra-erythrocytic stages of P. falciparum. Comparison between staining of live with compromised parasites shows that CMXRos predominantly stains live parasites with functional mitochondria. Parasite counts obtained by CMXRos staining and flow cytometry were highly reproducible and can reliably determine the ability of IgG from hyper-immune individuals to inhibit parasite growth in presence of monocytes in ADCI assay. Further, a dose-dependent parasite growth inhibitory effect could be detected for both total IgG purified from hyper-immune sera and affinity purified IgGs against the N-terminal non-repeat region of GLURP

  3. A Stopped-Flow Kinetics Experiment for the Physical Chemistry Laboratory Using Noncorrosive Reagents

    Science.gov (United States)

    Prigodich, Richard V.

    2014-01-01

    Stopped-flow kinetics techniques are important to the study of rapid chemical and biochemical reactions. Incorporation of a stopped-flow kinetics experiment into the physical chemistry laboratory curriculum would therefore be an instructive addition. However, the usual reactions studied in such exercises employ a corrosive reagent that can over…

  4. A Stopped-Flow Kinetics Experiment for the Physical Chemistry Laboratory Using Noncorrosive Reagents

    Science.gov (United States)

    Prigodich, Richard V.

    2014-01-01

    Stopped-flow kinetics techniques are important to the study of rapid chemical and biochemical reactions. Incorporation of a stopped-flow kinetics experiment into the physical chemistry laboratory curriculum would therefore be an instructive addition. However, the usual reactions studied in such exercises employ a corrosive reagent that can over…

  5. Unsteady mixed flows in non uniform closed water pipes: a Full Kinetic Appraoch

    CERN Document Server

    Bourdarias, Christian; Gerbi, Stéphane

    2011-01-01

    We recall the \\PFS} model constructed for the modeling of unsteady mixed flows in closed water pipes where transition points between the free surface and pressurized flow are treated as a free boundary associated to a discontinuity of the gradient of pressure. Then we present a numerical kinetic scheme for the computations of unsteady mixed flows in closed water pipes. This kinetic method that we call FKA for "Full Kinetic Approach" is an easy and mathematically elegant way to deal with multiple transition points when the changes of state between free surface and pressurized flow occur. We use two approaches namely the "ghost waves approach" and the "Full Kinetic Approach" to treat these transition points. We show that this kinetic numerical scheme has the following properties: it is wet area conservative, under a CFL condition it preserves the wet area positive, it treats "naturally" the drying and flooding area and most of all it preserves every stationary flow. Finally numerical experiment versus laborator...

  6. Flow Cytometric Panel-Reactive Antibody Results and the Ability to Find Transfusion-Compatible Platelets after Antibody-Desensitization for Allogeneic Bone Marrow Transplant.

    Science.gov (United States)

    Rosenbaum, Eric R; Pandey, Soumya; Harville, Terry O; Drobena, Gina A; Cottler-Fox, Michele

    2016-12-01

    Panel reactive antibody (PRA) reduction protocols are used to decrease anti-HLA antibodies with concomitant PRA monitoring as a measure of successful treatment prior to organ and haploidentical blood and marrow transplant (BMT). We hypothesized that the more sensitive flow cytometry (FC) based assays for PRA [FlowPRA(®) and Luminex(®) based Single Antigen Bead (SAB)] would also correlate with the ability to find compatible platelets for allosensitized recipients. A female patient with myelodysplastic syndrome and a high HLA class I PRA [>90% PRA and cPRA by complement-dependent cytotoxicity (CDC) assay and Flow PRA] required allogeneic BMT. Baseline HLA Class I and class II antigen typing was performed and a matched sibling donor was identified. Although baseline anti-HLA class I and class II antibodies measured by FC and CDC revealed no donor specific antibodies (DSA), the decision was made to attempt antibody desensitization to facilitate platelet transfusion during BMT. FC and CDC assays were performed to determine anti-HLA class I antibodies and cPRA/%PRA prior to starting desensitization and at the end of desensitization. Over the course of desensitization and BMT, a total of 194 apheresis platelet units underwent cross-match (XM) using Capture-P(®). We compared temporally-related PRA results with platelet XM results. High PRA by FC or CDC assays correlates with a high % of XM-positive (incompatible) platelet units. When the CDC PRA fell to 2% after desensitization, platelet XM incompatibility fell from 100% to 63% positive (incompatible). When the FC PRA fell to 5% the positive platelet XM fell to 5%. Antibody desensitization facilitated platelet transfusion. PRA determination by FC appeared better correlated than determination by CDC with the ability to find XM-compatible platelets. © 2016 by the Association of Clinical Scientists, Inc.

  7. Super-resolved calibration-free flow cytometric characterization of platelets and cell-derived microparticles in platelet-rich plasma.

    Science.gov (United States)

    Konokhova, Anastasiya I; Chernova, Darya N; Moskalensky, Alexander E; Strokotov, Dmitry I; Yurkin, Maxim A; Chernyshev, Andrei V; Maltsev, Valeri P

    2016-02-01

    Importance of microparticles (MPs), also regarded as extracellular vesicles, in many physiological processes and clinical conditions motivates one to use the most informative and precise methods for their characterization. Methods based on individual particle analysis provide statistically reliable distributions of MP population over characteristics. Although flow cytometry is one of the most powerful technologies of this type, the standard forward-versus-side-scattering plots of MPs and platelets (PLTs) overlap considerably because of similarity of their morphological characteristics. Moreover, ordinary flow cytometry is not capable of measurement of size and refractive index (RI) of MPs. In this study, we 1) employed the potential of the scanning flow cytometer (SFC) for identification and characterization of MPs from light scattering; 2) suggested the reference method to characterize MP morphology (size and RI) with high precision; and 3) determined the lowest size of a MP that can be characterized from light scattering with the SFC. We equipped the SFC with 405 and 488 nm lasers to measure the light-scattering profiles and side scattering from MPs, respectively. The developed two-stage method allowed accurate separation of PLTs and MPs in platelet-rich plasma. We used two optical models for MPs, a sphere and a bisphere, in the solution of the inverse light-scattering problem. This solution provides unprecedented precision in determination of size and RI of individual spherical MPs-median uncertainties (standard deviations) were 6 nm and 0.003, respectively. The developed method provides instrument-independent quantitative information on MPs, which can be used in studies of various factors affecting MP population.

  8. Changes of proliferation kinetics after X-irradiation of a human malignant melanoma grown in nude mice

    DEFF Research Database (Denmark)

    Spang-Thomsen, M; Vindeløv, L L

    1984-01-01

    A human malignant melanoma grown in nude mice was exposed to single-dose X-irradiation and the effect on the proliferation kinetics was investigated by two methods. Flow cytometric DNA analysis was performed on tumour tissue obtained by sequential fine-needle aspirations after the treatment to mo......-related increasing proportion of radiation-inactivated tumour cells....

  9. Fourier transform infra-red spectroscopy and flow cytometric assessment of the antibacterial mechanism of action of aqueous extract of garlic (Allium sativum) against selected probiotic Bifidobacterium strains.

    Science.gov (United States)

    Booyens, Jemma; Thantsha, Mapitsi Silvester

    2014-08-06

    It is generally reported that garlic (Allium sativum) harms pathogenic but not beneficial bacteria. Although numerous studies supporting the alleged garlic effects on pathogens are available, there are limited studies to prove this claim for beneficial bacteria. We have recently shown that garlic exhibits antibacterial activity against probiotic bifidobacteria. The aim of the current study was to elucidate the mechanism of action of garlic clove extract (GCE) on Bifidobacterium bifidum LMG 11041, B. longum LMG 13197 and B. lactis Bb12 using Fourier transform infrared (FT-IR) spectroscopy and flow cytometry. Cultures (1 × 108 CFU ml-1) were individually incubated for 6 h at 37°C in garlic clove extract containing allicin at a corresponding predetermined minimum bactericidal concentration for each strain. For FTIR, an aliquot of each culture was deposited on CaF2 slide and vacuum dried. The slides were immediately viewed using a Bruker Vertex 70 V FT-IR spectrometer equipped with a Hyperion microscope and data analyzed using OPUS software (version 6, Bruker). Spectra were smoothed with a Savitsky-Goly function algorithim, base-line corrected and normalized. Samples for flow cytometry were stained using the Live/Dead BacLight bacterial viability kit L7012. Data compensation and analysis was performed using a BD FACSAria and FlowJo (version 7.6.1). Fourier transform infrared spectroscopy showed changes in spectral features of lipids and fatty acids in cell membranes, proteins, polysaccharides and nucleic acids. Spectral data as per principle component analysis (PCA) revealed segregation of control and GCE-treated cells for all the tested bifidobacteria. Flow cytometry not only showed increase in numbers of membrane damaged and possibly lysed cells after GCE treatment, but also displayed diffuse light scatter patterns for GCE treated cells, which is evidence for changes to the size, granularity and molecular content of the cells. Garlic has multiple target sites in

  10. Modeling turbulence structure. Chemical kinetics interaction in turbulent reactive flows

    Energy Technology Data Exchange (ETDEWEB)

    Magnussen, B.F. [The Norwegian Univ. of Science and Technology, Trondheim (Norway)

    1997-12-31

    The challenge of the mathematical modelling is to transfer basic physical knowledge into a mathematical formulation such that this knowledge can be utilized in computational simulation of practical problems. The combustion phenomena can be subdivided into a large set of interconnected phenomena like flow, turbulence, thermodynamics, chemical kinetics, radiation, extinction, ignition etc. Combustion in one application differs from combustion in another area by the relative importance of the various phenomena. The difference in fuel, geometry and operational conditions often causes the differences. The computer offers the opportunity to treat the individual phenomena and their interactions by models with wide operational domains. The relative magnitude of the various phenomena therefore becomes the consequence of operational conditions and geometry and need not to be specified on the basis of experience for the given problem. In mathematical modelling of turbulent combustion, one of the big challenges is how to treat the interaction between the chemical reactions and the fluid flow i.e. the turbulence. Different scientists adhere to different concepts like the laminar flamelet approach, the pdf approach of the Eddy Dissipation Concept. Each of these approaches offers different opportunities and problems. All these models are based on a sound physical basis, however none of these have general validity in taking into consideration all detail of the physical chemical interaction. The merits of the models can only be judged by their ability to reproduce physical reality and consequences of operational and geometric conditions in a combustion system. The presentation demonstrates and discusses the development of a coherent combustion technology for energy conversion and safety based on the Eddy Dissipation Concept by Magnussen. (author) 30 refs.

  11. Flow Cytometric Immunobead Assay for Detection of BCR-ABL1 Fusion Proteins in Chronic Myleoid Leukemia: Comparison with FISH and PCR Techniques.

    Directory of Open Access Journals (Sweden)

    Anna Grazia Recchia

    Full Text Available Chronic Myeloid Leukemia (CML is characterized by a balanced translocation juxtaposing the Abelson (ABL and breakpoint cluster region (BCR genes. The resulting BCR-ABL1 oncogene leads to increased proliferation and survival of leukemic cells. Successful treatment of CML has been accompanied by steady improvements in our capacity to accurately and sensitively monitor therapy response. Currently, measurement of BCR-ABL1 mRNA transcript levels by real-time quantitative PCR (RQ-PCR defines critical response endpoints. An antibody-based technique for BCR-ABL1 protein recognition could be an attractive alternative to RQ-PCR. To date, there have been no studies evaluating whether flow-cytometry based assays could be of clinical utility in evaluating residual disease in CML patients. Here we describe a flow-cytometry assay that detects the presence of BCR-ABL1 fusion proteins in CML lysates to determine the applicability, reliability, and specificity of this method for both diagnosis and monitoring of CML patients for initial response to therapy. We show that: i CML can be properly diagnosed at onset, (ii follow-up assessments show detectable fusion protein (i.e. relative mean fluorescent intensity, rMFI%>1 when BCR-ABL1IS transcripts are between 1-10%, and (iii rMFI% levels predict CCyR as defined by FISH analysis. Overall, the FCBA assay is a rapid technique, fully translatable to the routine management of CML patients.

  12. Flow Cytometric Immunobead Assay for Detection of BCR-ABL1 Fusion Proteins in Chronic Myleoid Leukemia: Comparison with FISH and PCR Techniques.

    Science.gov (United States)

    Recchia, Anna Grazia; Caruso, Nadia; Bossio, Sabrina; Pellicanò, Mariavaleria; De Stefano, Laura; Franzese, Stefania; Palummo, Angela; Abbadessa, Vincenzo; Lucia, Eugenio; Gentile, Massimo; Vigna, Ernesto; Caracciolo, Clementina; Agostino, Antolino; Galimberti, Sara; Levato, Luciano; Stagno, Fabio; Molica, Stefano; Martino, Bruno; Vigneri, Paolo; Di Raimondo, Francesco; Morabito, Fortunato

    2015-01-01

    Chronic Myeloid Leukemia (CML) is characterized by a balanced translocation juxtaposing the Abelson (ABL) and breakpoint cluster region (BCR) genes. The resulting BCR-ABL1 oncogene leads to increased proliferation and survival of leukemic cells. Successful treatment of CML has been accompanied by steady improvements in our capacity to accurately and sensitively monitor therapy response. Currently, measurement of BCR-ABL1 mRNA transcript levels by real-time quantitative PCR (RQ-PCR) defines critical response endpoints. An antibody-based technique for BCR-ABL1 protein recognition could be an attractive alternative to RQ-PCR. To date, there have been no studies evaluating whether flow-cytometry based assays could be of clinical utility in evaluating residual disease in CML patients. Here we describe a flow-cytometry assay that detects the presence of BCR-ABL1 fusion proteins in CML lysates to determine the applicability, reliability, and specificity of this method for both diagnosis and monitoring of CML patients for initial response to therapy. We show that: i) CML can be properly diagnosed at onset, (ii) follow-up assessments show detectable fusion protein (i.e. relative mean fluorescent intensity, rMFI%>1) when BCR-ABL1IS transcripts are between 1-10%, and (iii) rMFI% levels predict CCyR as defined by FISH analysis. Overall, the FCBA assay is a rapid technique, fully translatable to the routine management of CML patients.

  13. Flow cytometric analysis of hemetopoietic progenitor cells in peripheral blood stem cell harvest from patients with CD34 positive acute leukemia.

    Science.gov (United States)

    Miyazaki, T; Matsuda, I; Oguri, M; Amaya, H; Kiyosaki, M; Hamada, A; Tamaki, S; Tashiro, E; Kudo, Y; Taniguchi, O; Nakamura, T; Tomoyasu, S

    2001-01-01

    We analyzed CD34 positive cells in peripheral blood stem cell harvest (PBSCH) using flow cytometry. PBSCH from CD34 positive acute myelogeous leukemia (AML-M2) patient contained 1.87% CD34 positive cells, of which 1.21% was represented by MRD.PBSCH from CD34 positive acute lymphoblast leukemia (ALL) patient contained 3.14% CD34 positive cells, of which 0.11% was accounted for by minimal residual disease (MRD). If PBSCH from CD34 positive acute leukemia patient is analyzed for CD34 monoclonal antibody alone, the presence of CD34 positive MRD may escape attention so that CD34 positive hematopoietic progenitor cells may be overestimated. To avoid this risk, it is necessary to analyze PBSCH using both CD34 monoclonal antibody and characteristic markers of leukemia cells that were found pre-treatment.

  14. Single-colour flow cytometric assay to determine NK cell-mediated cytotoxicity and viability against non-adherent human tumor cells.

    Science.gov (United States)

    Thakur, Ajit; Zaman, Abeyat; Hummel, Jeff; Jones, Kim; Hortelano, Gonzalo

    2012-03-01

    A flow cytometry-based cytotoxicity (FCC) assay was developed using a single fluorophore, calcein-acetoxymethyl diacetylester (calcein-AM), to measure NK cell-mediated cytotoxicity. Non-adherent human K562 and U937 target cells were individually labelled with calcein-AM and co-incubated with effector NK cells to measure calcein loss, and therefore calculate target cell cytotoxicity. This FCC assay also provided a measure of sample viability. Notably, cell viability measured by traditional calcein/7-amino-actinomycin D (7-AAD) double labelling and Trypan Blue methods were comparable to the viability calculated using calcein-loss FCC. This FCC assay may also be used with various effector and target cell types and as a multi-parameter tool to measure viability and immunophenotype cells for tissue engineering purposes.

  15. Flow cytometric detection and quantification of CD56 (neural cell adhesion molecule, NCAM) expression in diffuse large B cell lymphomas and review of the literature.

    Science.gov (United States)

    Stacchini, Alessandra; Barreca, Antonella; Demurtas, Anna; Aliberti, Sabrina; di Celle, Paola Francia; Novero, Domenico

    2012-02-01

    To report unusual CD56 (neural cell adhesion molecule, NCAM) expression on diffuse large B cell lymphoma (DLBCL). CD56 expression was first detected and quantified on tissues obtained from five cases of DLBCL by flow cytometry (FC), then confirmed by immunohistochemistry. The CD56 expression pattern was heterogeneous among the cases [the molecular equivalent of soluble fluorochrome (MESF) level ranged from 2214 to 133 466]. All were CD10 and Bcl-6 positive, suggesting their germinal centre origin; one was also CD5 positive. An extranodal presentation occurred in three of five cases. CD56 expression in B cell lymphoma is a rare occurrence. FC is able to identify aberrant immunophenotypes that can be useful in the identification and monitoring of B cell lymphoma subtypes. The presence of CD56 reported by the literature on certain DLBCL with extranodal presentation might be related to mechanisms involved in growth and expansion. © 2012 Blackwell Publishing Limited.

  16. Evaluation of a multi-endpoint assay in rats, combining the bone-marrow micronucleus test, the Comet assay and the flow-cytometric peripheral blood micronucleus test.

    Science.gov (United States)

    Bowen, Damian E; Whitwell, James H; Lillford, Lucinda; Henderson, Debbie; Kidd, Darren; Mc Garry, Sarah; Pearce, Gareth; Beevers, Carol; Kirkland, David J

    2011-05-18

    With the publication of revised draft ICH guidelines (Draft ICH S2), there is scope and potential to establish a combined multi-end point in vivo assay to alleviate the need for multiple in vivo assays, thereby reducing time, cost and use of animals. Presented here are the results of an evaluation trial in which the bone-marrow and peripheral blood (via MicroFlow(®) flow cytometry) micronucleus tests (looking at potential chromosome breakage and whole chromosome loss) in developing erythrocytes or young reticulocytes were combined with the Comet assay (measuring DNA strand-breakage), in stomach, liver and blood lymphocytes. This allowed a variety of potential target tissues (site of contact, site of metabolism and peripheral distribution) to be assessed for DNA damage. This combination approach was performed with minimal changes to the standard and regulatory recommended sampling times for the stand-alone assays. A series of eight in vivo genotoxins (2-acetylaminofluorene, benzo[a]pyrene, carbendazim, cyclophosphamide, dimethylnitrosamine, ethyl methanesulfonate, ethyl nitrosourea and mitomycin C), which are known to act via different modes of action (direct- and indirect-acting clastogens, alkylating agents, gene mutagens, cross-linking and aneugenic compounds) were tested. Male rats were dosed at 0, 24 and 45 h, and bone marrow and peripheral blood (micronucleus endpoint), liver, whole blood and stomach (Comet endpoint) were sampled at three hours after the last dose. Comet and micronucleus responses were as expected based on available data for conventional (acute) stand-alone assays. All compounds were detected as genotoxic in at least one of the endpoints. The importance of evaluating both endpoints was highlighted by the uniquely positive responses for certain chemicals (benzo[a]pyrene and 2-acetylaminofluorene) with the Comet endpoint and certain other chemicals (carbendazim and mitomycin C) with the micronucleus endpoint. The data generated from these

  17. Accurate detection of the tumor clone in peripheral T-cell lymphoma biopsies by flow cytometric analysis of TCR-Vβ repertoire.

    Science.gov (United States)

    Salameire, Dimitri; Solly, Françoise; Fabre, Blandine; Lefebvre, Christine; Chauvet, Martine; Gressin, Rémy; Corront, Bernadette; Ciapa, Agnès; Pernollet, Martine; Plumas, Joël; Macintyre, Elizabeth; Callanan, Mary B; Leroux, Dominique; Jacob, Marie-Christine

    2012-09-01

    Multiparametric flow cytometry has proven to be a powerful method for detection and immunophenotypic characterization of clonal subsets, particularly in lymphoproliferative disorders of the B-cell lineage. Although in theory promising, this approach has not been comparably fulfilled in mature T-cell malignancies. Specifically, the T-cell receptor-Vβ repertoire analysis in blood can provide strong evidence of clonality, particularly when a single expanded Vß family is detected. The purpose of this study was to determine the relevance of this approach when applied to biopsies, at the site of tumor involvement. To this end, 30 peripheral T-cell lymphoma and 94 control biopsies were prospectively studied. Vβ expansions were commonly detected within CD4+ or CD8+ T cells (97% of peripheral T-cell lymphoma and 54% of non-peripheral T-cell lymphoma cases); thus, not differentiating malignant from reactive processes. Interestingly, we demonstrated that using a standardized evaluation, the detection of a high Vβ expansion was closely associated with diagnosis of peripheral T-cell lymphoma, with remarkable specificity (98%) and sensitivity (90%). This approach also identified eight cases of peripheral T-cell lymphoma that were not detectable by other forms of immunophenotyping. Moreover, focusing Vβ expression analysis to T-cell subsets with aberrant immunophenotypes, we demonstrated that the T-cell clone might be heterogeneous with regard to surface CD7 or CD10 expression (4/11 cases), providing indication on 'phenotypic plasticity'. Finally, among the wide variety of Vβ families, the occurrence of a Vβ17 expansion in five cases was striking. To our knowledge, this is the first report demonstrating the power of T-cell receptor-Vβ repertoire analysis by flow cytometry in biopsies as a basis for peripheral T-cell lymphoma diagnosis and precise T-cell clone identification and characterization.

  18. Normal adult ramified microglia separated from other central nervous system macrophages by flow cytometric sorting: Phenotypic differences defined and direct ex vivo antigen presentation to myelin basic protein-reactive CD4{sup +} T cells compared

    Energy Technology Data Exchange (ETDEWEB)

    Ford, A.L.; Goodsall, A.L.; Sedgwick, J.D. [Centenary Institute of Cancer Medicine and Cell Biology, Sydney (Australia)] [and others

    1995-05-01

    Ramified microglia in the adult central nervous system (CNS) are the principal glial element up-regulating MHC class I and II expression in response to inflammatory events or neuronal damage. A proportion of these cells also express MHC class II constitutively in the normal CNS. The role of microglia as APCs for CD4{sup +} cells extravasating into the CNS remains undefined. In this study, using irradiation bone marrow chimeras in CD45-congenic rats, the phenotype CD45{sup low}CD11b/c{sup +} is shown to identify microglial cells specifically within the CNS. Highly purified populations of microglia and nonmicroglial but CNS-associated macrophages (CD45{sup high}CD11b/c{sup +}) have been obtained directly from the adult CNS, by using flow cytometric sorting. Morphologically, freshly isolated microglia vs other CNS macrophages are quite distinct. Of the two populations recovered from the normal CNS, it is the minority CD45{sup high}CD11 b/c{sup +} transitional macrophage population, and not microglia, that is the effective APC for experimental autoimmune encephalomyelitis-inducing CD4{sup +} myelin basic protein (MBP)-reactive T cells. CD45{sup high}CD11b/c{sup +} CNS macrophages also stimulate MBP-reactive T cells without addition of MBP to culture suggesting presentation of endogenous Ag. This is the first study in which microglia vs other CNS macrophages have been analyzed for APC ability directly from the CNS, with substantial cross-contamination between the two populations eliminated. The heterogeneity of these populations in terms of APC function is clearly demonstrated. Evidence is still lacking that adult CNS microglia have the capacity to interact with and stimulate CD4{sup +} T cells to proliferate or secrete IL-2. 60 refs., 6 figs., 1 tab.

  19. Innovations in diagnosis and post-therapeutic monitoring of Chagas disease: Simultaneous flow cytometric detection of IgG1 antibodies anti-live amastigote, anti-live trypomastigote, and anti-fixed epimastigote forms of Trypanosoma cruzi.

    Science.gov (United States)

    Alessio, Glaucia Diniz; Côrtes, Denise Fonseca; Machado de Assis, Girley Francisco; Júnior, Policarpo Ademar Sales; Ferro, Eloisa Amália Vieira; Antonelli, Lis Ribeiro do Valle; Teixeira-Carvalho, Andréa; Martins-Filho, Olindo Assis; de Lana, Marta

    2014-11-01

    This study developed a remarkable methodological innovation (FC-ATE) which enables simultaneous detection of antibodies specific to the three evolutive forms of Trypanosoma cruzi: live amastigote (AMA), live trypomastigote (TRYPO), and fixed epimastigote (EPI) using a differential fluorescence staining as low (AMA), intermediate (TRYPO), and high (EPI). An outstanding performance (100%) was observed in the discrimination of the chagasic (CH) and non-chagasic (NCH) patients. In the applicability of FC-ATE in the diagnosis of Chagas disease, 100% of the CH samples presented positivity in the percentage of positive fluorescent parasites (PPFP) for all the three forms of T. cruzi. Moreover, 94% of the samples of NCH presented negative values of PPFP with AMA and TRYPO, and 88% with EPI. Samples from the NCH group with false-positive results were those belonging to the leishmaniasis patients. Considering the applicability of this technique in post-therapeutic monitoring of Chagas disease, 100% of non-treated (NT) and treated non-cured (TNC) samples were positive with the three T. cruzi evolutive forms, while a percentage of 100% from samples of the treated cured (TC) patients were negative with AMA, 93% with TRYPO and 96% with EPI. The comparison between FC-ATE and two other flow cytometric tests using the same samples of patients NT, TNC and TC showed that the three techniques presented different reactivities, although categorical correlation between the methodologies was observed. Taken together, the results obtained with the novel FC-ATE method have shown an outstanding performance in the diagnosis and post-therapeutic monitoring of Chagas disease. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. In vitro holding and PLD-repair. Pt. 2. A flow cytometric and electron microscopic analysis of some mammalian cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Stevenson, A.F.G. (Institute for Basic Research in Developmental Disabilities, Staten Island, NY (USA)); Palackal, T. (State Univ. of New York, Brooklyn, NY (USA). Div. of Radiobiology); Lange, C.S. (Kiel Univ. (Germany, F.R.). Abt. Frauenheilkunde)

    1989-12-01

    The repair of potentially lethal damage (PLDR) in mammalian cells is expected to be better in quiescent cultures since PLD is supposedly fixed during cycle progression. Plateau phase cultures, therefore, serve as models because of assumed mitotic quiescence. Four established cell lines (V79, CHO, L5178Y and HELA) and one euploid cell strain IMR-90 have been analysed by flow cytometry and electron microscopy to address questions on quiescence in the plateau phase and the effect of holding (induction of quiescence by nutrient privation). In contrast to commonly held views, our results indicate that the quiescent fraction in cultures from transformed cells is exceedingly low (1% or less). Plateau phase cultures of transformed cells are constantly turning over. Euploid cells like the IMR-90 show true quiescence in the plateau phase. Holding causes typical cytopathological changes. These changes have been ultrastructurely characterised. Resistant sub-populations of cells can be selected out under holding-conditions. Such selected cells show completely different radiobiological characteristics, which raise questions on the interpretation of data on PLDR. (orig.).

  1. Final results of a multicenter trial addressing role of CSF flow cytometric analysis in NHL patients at high risk for CNS dissemination.

    Science.gov (United States)

    Benevolo, Giulia; Stacchini, Alessandra; Spina, Michele; Ferreri, Andrés J M; Arras, Marcella; Bellio, Laura; Botto, Barbara; Bulian, Pietro; Cantonetti, Maria; Depaoli, Lorella; Di Renzo, Nicola; Di Rocco, Alice; Evangelista, Andrea; Franceschetti, Silvia; Godio, Laura; Mannelli, Francesco; Pavone, Vincenzo; Pioltelli, Pietro; Vitolo, Umberto; Pogliani, Enrico M

    2012-10-18

    This prospective study compared diagnostic and prognostic value of conventional cytologic (CC) examination and flow cytometry (FCM) of baseline samples of cerebrospinal fluid (CSF) in 174 patients with newly diagnosed aggressive non-Hodgkin lymphoma (NHL). FCM detected a neoplastic population in the CSF of 18 of 174 patients (10%), CC only in 7 (4%; P < .001); 11 patients (14%) were discordant (FCM(+)/CC(-)). At a median follow-up of 46 months, there were 64 systemic progressions and 10 CNS relapses, including 2 patients with both systemic and CNS relapses. Two-year progression-free and overall survival were significantly higher in patients with FCM(-) CSF (62% and 72%) compared with those FCM(+) CSF (39% and 50%, respectively), with a 2-year CNS relapse cumulative incidence of 3% (95% confidence interval [CI], 0-7) versus 17% (95% CI, 0-34; P = .004), respectively. The risk of CNS progression was significantly higher in FMC(+)/CC(-) versus FCM(-)/CC(-) patients (hazard ratio = 8.16, 95% CI, 1.45-46). In conclusion, FCM positivity in the CSF of patients with high-risk NHL is associated with a significantly higher CNS relapse risk and poorer outcome. The combination of IV drugs with a higher CNS bioavailability and intrathecal chemotherapy is advisable to prevent CNS relapses in FCM(+) patients.

  2. Clinical performance of human papillomavirus E6, E7 mRNA flow cytometric assay compared to human papillomavirus DNA typing.

    Science.gov (United States)

    Kottaridi, Christine; Tsiodras, Sotirios; Spathis, Aris; Chranioti, Aikaterini; Pappas, Asimakis; Kassanos, Dimitrios; Panayiotides, Ioannis; Karakitsos, Petros

    2011-12-01

    To use flow cytometry to screen cervical samples for the overexpression of human papillomavirus (HPV) E6 and E7 mRNA and compare the performance of this assay with an HPV DNA array for the detection of high-grade cervical lesions. Cervical samples were analyzed for HPV DNA by clinical arrays, and the overexpression of E6 and E7 viral oncogenes was monitored using an HPV mRNA detection kit that quantifies the intracellular HPV E6 and E7 mRNA on a cell-by-cell basis. HPV positivity increased with severity of histologic lesions. On the basis of histology-confirmed CIN 2+ cases the specificity of HPV assay was 73.9% (95% CI 66.07, 80.88), whereas it was 39.3% (95% CI 31.85, 47.1) for the DNA assay. The HPV assay provides an early predictor of persistent HPV infection and may improve cervical cancer screening by increasing the specificity of detecting high-grade lesions.

  3. Genotoxicity of doxorubicin in F344 rats by combining the comet assay, flow-cytometric peripheral blood micronucleus test, and pathway-focused gene expression profiling.

    Science.gov (United States)

    Manjanatha, Mugimane G; Bishop, Michelle E; Pearce, Mason G; Kulkarni, Rohan; Lyn-Cook, Lascelles E; Ding, Wei

    2014-01-01

    Doxorubicin (DOX) is an antineoplastic drug effective against many human malignancies. DOX's clinical efficacy is greatly limited because of severe cardiotoxicity. To evaluate if DOX is genotoxic in the heart, ~7-week-old, male F344 rats were administered intravenously 1, 2, and 3 mg/kg bw DOX at 0, 24, 48, and 69 hr and the Comet assays in heart, liver, kidney, and testis and micronucleus (MN) assay in the peripheral blood (PB) erythrocytes using flow cytometry were conducted. Rats were euthanized at 72 hr and PB was removed for the MN assay and single cells were isolated from multiple tissues for the Comet assays. None of the doses of DOX induced a significant DNA damage in any of the tissues examined by the alkaline Comet assay. Contrastingly, the glycosylase enzymes-modified Comet assay showed a significant dose dependent increase in the oxidative DNA damage in the cardiac tissue (P ≤ 0.05). In the liver, only the top dose induced significant increase in the oxidative DNA damage (P ≤ 0.05). The histopathology showed no severe cardiotoxicity but non-neoplastic lesions were present in both untreated and treated samples. A severe toxicity likely occurred in the bone marrow because no viable reticulocytes could be screened for the MN assay. Gene expression profiling of the heart tissues showed a significant alteration in the expression of 11 DNA damage and repair genes. These results suggest that DOX is genotoxic in the heart and the DNA damage may be induced primarily via the production of reactive oxygen species.

  4. Dynamics of Zonal Flows: Failure of Wave-Kinetic Theory, and New Geometrical Optics Approximations

    CERN Document Server

    Parker, Jeffrey B

    2016-01-01

    The self-organization of turbulence into regular zonal flows can be fruitfully investigated with quasilinear methods and statistical descriptions. A wave kinetic equation that assumes asymptotically large-scale zonal flows is pathological. From an exact description of quasilinear dynamics emerges two better geometrical optics approximations. These involve not only the mean flow shear but also the second and third derivative of the mean flow. One approximation takes the form of a new wave kinetic equation, but is only valid when the zonal flow is quasi-static and wave action is conserved.

  5. EPR-Spin Trapping and Flow Cytometric Studies of Free Radicals Generated Using Cold Atmospheric Argon Plasma and X-Ray Irradiation in Aqueous Solutions and Intracellular Milieu.

    Directory of Open Access Journals (Sweden)

    Hidefumi Uchiyama

    Full Text Available Electron paramagnetic resonance (EPR-spin trapping and flow cytometry were used to identify free radicals generated using argon-cold atmospheric plasma (Ar-CAP in aqueous solutions and intracellularly in comparison with those generated by X-irradiation. Ar-CAP was generated using a high-voltage power supply unit with low-frequency excitation. The characteristics of Ar-CAP were estimated by vacuum UV absorption and emission spectra measurements. Hydroxyl (·OH radicals and hydrogen (H atoms in aqueous solutions were identified with the spin traps 5,5-dimethyl-1-pyrroline N-oxide (DMPO, 3,3,5,5-tetramethyl-1-pyrroline-N-oxide (M4PO, and phenyl N-t-butylnitrone (PBN. The occurrence of Ar-CAP-induced pyrolysis was evaluated using the spin trap 3,5-dibromo-4-nitrosobenzene sulfonate (DBNBS in aqueous solutions of DNA constituents, sodium acetate, and L-alanine. Human lymphoma U937 cells were used to study intracellular oxidative stress using five fluorescent probes with different affinities to a number of reactive species. The analysis and quantification of EPR spectra revealed the formation of enormous amounts of ·OH radicals using Ar-CAP compared with that by X-irradiation. Very small amounts of H atoms were detected whereas nitric oxide was not found. The formation of ·OH radicals depended on the type of rare gas used and the yield correlated inversely with ionization energy in the order of krypton > argon = neon > helium. No pyrolysis radicals were detected in aqueous solutions exposed to Ar-CAP. Intracellularly, ·OH, H2O2, which is the recombination product of ·OH, and OCl- were the most likely formed reactive oxygen species after exposure to Ar-CAP. Intracellularly, there was no practical evidence for the formation of NO whereas very small amounts of superoxides were formed. Despite the superiority of Ar-CAP in forming ·OH radicals, the exposure to X-rays proved more lethal. The mechanism of free radical formation in aqueous solutions and

  6. EPR-Spin Trapping and Flow Cytometric Studies of Free Radicals Generated Using Cold Atmospheric Argon Plasma and X-Ray Irradiation in Aqueous Solutions and Intracellular Milieu.

    Science.gov (United States)

    Uchiyama, Hidefumi; Zhao, Qing-Li; Hassan, Mariame Ali; Andocs, Gabor; Nojima, Nobuyuki; Takeda, Keigo; Ishikawa, Kenji; Hori, Masaru; Kondo, Takashi

    2015-01-01

    Electron paramagnetic resonance (EPR)-spin trapping and flow cytometry were used to identify free radicals generated using argon-cold atmospheric plasma (Ar-CAP) in aqueous solutions and intracellularly in comparison with those generated by X-irradiation. Ar-CAP was generated using a high-voltage power supply unit with low-frequency excitation. The characteristics of Ar-CAP were estimated by vacuum UV absorption and emission spectra measurements. Hydroxyl (·OH) radicals and hydrogen (H) atoms in aqueous solutions were identified with the spin traps 5,5-dimethyl-1-pyrroline N-oxide (DMPO), 3,3,5,5-tetramethyl-1-pyrroline-N-oxide (M4PO), and phenyl N-t-butylnitrone (PBN). The occurrence of Ar-CAP-induced pyrolysis was evaluated using the spin trap 3,5-dibromo-4-nitrosobenzene sulfonate (DBNBS) in aqueous solutions of DNA constituents, sodium acetate, and L-alanine. Human lymphoma U937 cells were used to study intracellular oxidative stress using five fluorescent probes with different affinities to a number of reactive species. The analysis and quantification of EPR spectra revealed the formation of enormous amounts of ·OH radicals using Ar-CAP compared with that by X-irradiation. Very small amounts of H atoms were detected whereas nitric oxide was not found. The formation of ·OH radicals depended on the type of rare gas used and the yield correlated inversely with ionization energy in the order of krypton > argon = neon > helium. No pyrolysis radicals were detected in aqueous solutions exposed to Ar-CAP. Intracellularly, ·OH, H2O2, which is the recombination product of ·OH, and OCl- were the most likely formed reactive oxygen species after exposure to Ar-CAP. Intracellularly, there was no practical evidence for the formation of NO whereas very small amounts of superoxides were formed. Despite the superiority of Ar-CAP in forming ·OH radicals, the exposure to X-rays proved more lethal. The mechanism of free radical formation in aqueous solutions and an

  7. Gas-kinetic unified algorithm for hypersonic flows covering various flow regimes solving Boltzmann model equation in nonequilibrium effect

    Science.gov (United States)

    Li, Zhihui; Wu, Junlin; Ma, Qiang; Jiang, Xinyu; Zhang, Hanxin

    2014-12-01

    Based on the Gas-Kinetic Unified Algorithm (GKUA) directly solving the Boltzmann model equation, the effect of rotational non-equilibrium is investigated recurring to the kinetic Rykov model with relaxation property of rotational degrees of freedom. The spin movement of diatomic molecule is described by moment of inertia, and the conservation of total angle momentum is taken as a new Boltzmann collision invariant. The molecular velocity distribution function is integrated by the weight factor on the internal energy, and the closed system of two kinetic controlling equations is obtained with inelastic and elastic collisions. The optimization selection technique of discrete velocity ordinate points and numerical quadrature rules for macroscopic flow variables with dynamic updating evolvement are developed to simulate hypersonic flows, and the gas-kinetic numerical scheme is constructed to capture the time evolution of the discretized velocity distribution functions. The gas-kinetic boundary conditions in thermodynamic non-equilibrium and numerical procedures are studied and implemented by directly acting on the velocity distribution function, and then the unified algorithm of Boltzmann model equation involving non-equilibrium effect is presented for the whole range of flow regimes. The hypersonic flows involving non-equilibrium effect are numerically simulated including the inner flows of shock wave structures in nitrogen with different Mach numbers of 1.5-Ma-25, the planar ramp flow with the whole range of Knudsen numbers of 0.0009-Kn-10 and the three-dimensional re-entering flows around tine double-cone body.

  8. Gas-kinetic unified algorithm for hypersonic flows covering various flow regimes solving Boltzmann model equation in nonequilibrium effect

    Energy Technology Data Exchange (ETDEWEB)

    Li, Zhihui; Ma, Qiang [Hypervelocity Aerodynamics Institute, China Aerodynamics Research and Development Center, P.O.Box 211, Mianyang 621000, China and National Laboratory for Computational Fluid Dynamics, No.37 Xueyuan Road, Beijing 100191 (China); Wu, Junlin; Jiang, Xinyu [Hypervelocity Aerodynamics Institute, China Aerodynamics Research and Development Center, P.O.Box 211, Mianyang 621000 (China); Zhang, Hanxin [National Laboratory for Computational Fluid Dynamics, No.37 Xueyuan Road, Beijing 100191 (China)

    2014-12-09

    Based on the Gas-Kinetic Unified Algorithm (GKUA) directly solving the Boltzmann model equation, the effect of rotational non-equilibrium is investigated recurring to the kinetic Rykov model with relaxation property of rotational degrees of freedom. The spin movement of diatomic molecule is described by moment of inertia, and the conservation of total angle momentum is taken as a new Boltzmann collision invariant. The molecular velocity distribution function is integrated by the weight factor on the internal energy, and the closed system of two kinetic controlling equations is obtained with inelastic and elastic collisions. The optimization selection technique of discrete velocity ordinate points and numerical quadrature rules for macroscopic flow variables with dynamic updating evolvement are developed to simulate hypersonic flows, and the gas-kinetic numerical scheme is constructed to capture the time evolution of the discretized velocity distribution functions. The gas-kinetic boundary conditions in thermodynamic non-equilibrium and numerical procedures are studied and implemented by directly acting on the velocity distribution function, and then the unified algorithm of Boltzmann model equation involving non-equilibrium effect is presented for the whole range of flow regimes. The hypersonic flows involving non-equilibrium effect are numerically simulated including the inner flows of shock wave structures in nitrogen with different Mach numbers of 1.5-Ma-25, the planar ramp flow with the whole range of Knudsen numbers of 0.0009-Kn-10 and the three-dimensional re-entering flows around tine double-cone body.

  9. Impact of the kinetic boundary condition on porous media flow in the lattice Boltzmann formulation

    Science.gov (United States)

    Singh, Shiwani; Jiang, Fei; Tsuji, Takeshi

    2017-07-01

    To emphasize the importance of the kinetic boundary condition for micro- to nanoscale flow, we present an ad hoc kinetic boundary condition suitable for torturous geological porous media. We found that the kinetic boundary condition is one of the essential features which should be supplemented to the standard lattice Boltzmann scheme in order to obtain accurate continuum observables. The claim is validated using a channel flow setup by showing the agreement of mass flux with analytical value. Further, using a homogeneous porous structure, the importance of the kinetic boundary condition is shown by comparing the permeability correction factor with the analytical value. Finally, the proposed alternate to the kinetic boundary condition is validated by showing its capability to capture the basic feature of the kinetic boundary condition.

  10. Simulation of horizontal slug-flow pneumatic conveying with kinetic theory

    Institute of Scientific and Technical Information of China (English)

    GU Zhengmeng; GUO Liejin

    2007-01-01

    Wavelike slug-flow is a representative flow type in horizontal pneumatic conveying.Kinetic theory was introduced to establish a 3D kinetic numerical model for wavelike slug gas-solid flow in this paper.Wavelike motion of particulate slugs in horizontal pipes was numerically investigated.The formation and motion process of slugs and settled layer were simulated.The characteristics of the flow,such as pressure drop,air velocity distribution,slug length and settled layer thickness,and the detailed changing characteristics of slug length and settled layer thickness with air velocity were obtained.The results indicate that kinetic theory can represent the physical characteristics of the non-suspension dense phase flow of wavelike slug pneumatic conveying.The experiment in this paper introduced a new idea for the numerical calculation of slug-flow pneumatic conveying.

  11. Novel swirl-flow reactor for kinetic studies of semiconductor photocatalysis

    NARCIS (Netherlands)

    Ray, A.K; Beenackers, A.A C M

    1997-01-01

    A new two-phase swirl-flow monolithic-type reactor was designed to study the kinetics of heterogeneous photocatalytic processes on immobilized semiconductor catalysts. True kinetic rate constants for destruction of a textile dye were measured as a function of wavelength of light intensity and angle

  12. Gas-Kinetic Computational Algorithms for Hypersonic Flows in Continuum and Transitional Regimes Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This SBIR Phase I project explores two gas-kinetic computational algorithms for simulation of hypersonic flows in both continuum and transitional regimes. One is the...

  13. Growth kinetics and in vivo radiosensitivity in nude mice of two subpopulations derived from a single human small cell carcinoma of the lung

    DEFF Research Database (Denmark)

    Spang-Thomsen, M; Clerici, M; Engelholm, S A;

    1986-01-01

    were described according to a transformed Gompertz function, and the cell kinetics were examined by flow cytometric DNA analysis (FCM) and by the technique of labelled mitoses. The effect of single-dose irradiation was estimated by the specific growth delay calculated from the growth curves...

  14. A Gas-Kinetic Scheme for Turbulent Flow

    Science.gov (United States)

    2014-09-19

    prediction of this area may lead to un- suitable geometries or high development costs. In this study a gas-kinetic scheme is used, follow- ing the RANS...reconstruction and Roe’s approximate Riemann solver. Remarkably, the gas-kinetic scheme provides a rather accurate predic- tion, whereas the conventional Navier...GKS on medium grid, ( ) GKS on coarsest grid, ( ) Navier-Stokes (Roe’s approximate Riemann solver) on finest grid, ( o ): experimental data from

  15. Unified Kinetic Approach for Simulation of Gas Flows in Rarefied and Continuum Regimes

    Science.gov (United States)

    2007-06-01

    a low-speed flow induced by temperature gradients. The nonuniform boundary temperature distribution can induce flows in reactor : a significant flow...Rotational Spectrum and Molecular Interaction Potential, ibid R. R. Arslanbekov and V. I. Kolobov, Simulation of Low Pressure Plasma Processing Reactors ... Microchannel flow in the slip regime: gas-kinetic BGK—Burnett solutions, J. Fluid Mech. 513, 87 (2004) 59 R.L.Bayut, PhD thesis, MIT 1999 60

  16. Identification of Streptococcus pneumoniae serotype 11E, serovariant 11Av and mixed populations by high-resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR spectroscopy and flow cytometric serotyping assay (FCSA.

    Directory of Open Access Journals (Sweden)

    Romina Camilli

    Full Text Available BACKGROUND: Recent studies have identified Streptococcus pneumoniae serotype 11E and serovariant 11Av among isolates previously typed as 11A by classical serotyping methods. Serotype 11E and serovariant 11Av differ from serotype 11A by having totally or partially inactive wcjE, a gene in cps locus coding for an O-acetyl transferase. Serotype 11E is rare among carriage isolates but common among invasive isolates suggesting that it survives better during invasion. Aim of this work was to investigate the epidemiology of serotype 11A in a pneumococcal collection using a new serotyping approach based on High-Resolution Magic Angle Spinning Nuclear Magnetic Resonance (HR-MAS NMR spectroscopy to distinguish serotypes 11A and 11E. METHODS: A collection of 48 (34 invasive and 14 carriage S. pneumoniae isolates from Italy, previously identified as serotype 11A by the Quellung reaction, were investigated by wcjE sequencing, HR-MAS NMR spectroscopy and the reference flow cytometric serotyping assay (FCSA based on monoclonal antibodies. RESULTS: HR-MAS NMR spectra from serotypes 11A and 11E showed different NMR peaks indicating that HR-MAS NMR could be used to distinguish these serotypes, although HR-MAS NMR could not distinguish serotype 11Av from serotype 11E unambiguously. Thirty-eight isolates were confirmed to be serotype 11A, 8 isolates with a mutated wcjE were serotype 11E, 1 isolate belonged to serovariant 11Av, and 1 isolate was a mixed population 11A/11Av. All 11E isolates were identified among invasive isolates. CONCLUSIONS: We proved that HR-MAS NMR can be of potential use for pneumococcal serotyping. The detection of serotype 11E among invasive isolates in our collection, supports previous epidemiological studies suggesting that mutations in wcjE can represent a mechanism promoting pneumococcal survival during invasion. The discovery of a spectrum of immunochemical diversity within established serotypes should stimulate efforts to develop new

  17. Assessment of the kinetic-frictional model for dense granular flow

    Institute of Scientific and Technical Information of China (English)

    Boon Ho Ng; Yulong Ding; Mojtaba Ghadiri

    2008-01-01

    This paper aims to quantitatively assess the application of kinetic-frictional model to simulate the motion of dry granular materials in dense condition, in particular, the annular shearing in Couette configuration. The weight of frictional stress was varied to study the contribution of the frictional stress in dense granular flows. The results show that the pure kinetic-theory-based computational fluid dynamics (CFD) model (without frictional stress) over-predicts the dominant solids motion of dense granular flow while adding frictional stress [Schaeffer, D. G. (1987). Instability in the evolution equations describing incompressible granular flow. Journal of Differential Equations, 66(1), 19-50] with the solids pressure of [Lun, C. KTK., Savage, S. B., Jeffrey, D. J., & Chepurniy, N. (1984). Kinetic theories for granular flow: Inelastic particles in Couette flow and slightly inelastic particles in a general flow field. Journal of Fluid Mechanics, 140, 223-256] in the CFD model improves the simulation to better conform available experimental results. The results also suggest that frictional stress transmission plays an important role in dense granular flow and should not be neglected in granular flow simulations. Compatible simulation results to the experimental data are seen by increasing the weight of frictional stress to a factor of 1.25-1.5. These improved simulation results suggest the current constitutive relations (kinetic-frictional model) need to be improved in order to better reflect the real dense granular flow.

  18. Flow-Based Systems for Rapid and High-Precision Enzyme Kinetics Studies

    Directory of Open Access Journals (Sweden)

    Supaporn Kradtap Hartwell

    2012-01-01

    Full Text Available Enzyme kinetics studies normally focus on the initial rate of enzymatic reaction. However, the manual operation of steps of the conventional enzyme kinetics method has some drawbacks. Errors can result from the imprecise time control and time necessary for manual changing the reaction cuvettes into and out of the detector. By using the automatic flow-based analytical systems, enzyme kinetics studies can be carried out at real-time initial rate avoiding the potential errors inherent in manual operation. Flow-based systems have been developed to provide rapid, low-volume, and high-precision analyses that effectively replace the many tedious and high volume requirements of conventional wet chemistry analyses. This article presents various arrangements of flow-based techniques and their potential use in future enzyme kinetics applications.

  19. Flow cytometric analysis of peripheral blood leukemic cells in relapse of acute leukemia%急性白血病复发外周血白血病细胞的流式细胞术检验分析

    Institute of Scientific and Technical Information of China (English)

    杨莉; 何浩明

    2015-01-01

    Objective To analyse status of peripheral blood leukemic cells detected by flow cytometry in patients with acute leu‐kemia(AL) ,and to provide references for evaluating clinical efficacy and prognosis of AL .Methods The peripheral blood specimens of 87 cases of patients with AL ,including 53 cases of patients with acute myelocytic leukemia and 34 cases of patients with acute lymphoblastic leukemia ,were detected by using flow cytometry ,morphological changes in bone marrow cells were detected ,as well . Results The sensitivity ,specificity and positive predictive value in determination of acute myelocytic leukemia was 95 .6% ,34 .5%and 81 .3% respectively ,and those in acute lymphoblastic leukemia was 87 .3% ,45 .6% and 68 .9% respectively ,statistically signif‐icant differences were found in sensitivity ,specificity and positive predictive value (P<0 .05) .A total of 19 cases with negative mini‐mal residual disease had recurrence(26 .31% ) after 24 months ,and 68 cases with positive minimal residual disease had recurrence (86 .76% ) after 24 months ,and the recurrence rate between the two groups was statistically significant (P<0 .05) .Among all pa‐tients with positive minimal residual disease ,the recurrence rate in patients with high expression level of minimal residual disease (88 .23% ) was higher than that in patients with low expression level of minimal residual disease (47 .09% ) ,and the difference was statistically significant (P<0 .05) .Conclusion Flow cytometric analysis of peripheral blood leukemic cells may has significance for diagnosing relapse of AL and guiding clinical medication .%目的:分析急性白血病(AL)患者外周血白血病细胞流式细胞术检测情况,为分析AL的临床疗效和预后提供参考。方法采用流式细胞术检测87例A L患者(急性髓细胞白血病53例、急性淋巴细胞白血病34例)外周血标本,同时检测骨髓细胞形态学变化。结果急性髓细胞白

  20. Kinetic theory and turbulent discontinuities. [shock tube flow

    Science.gov (United States)

    Johnson, J. A., III; I, L.; Li, Y.; Ramaian, R.; Santigo, J. P.

    1981-01-01

    Shock tube discontinuities were used to test and extend a kinetic theory of turbulence. In shock wave and contact surface fluctuations, coherent phenomena were found which provide new support for the microscopic nonempirical approach to turbulent systems, especially those with boundary layer-like instabilities.

  1. Gas kinetic algorithm for flows in Poiseuille-like microchannels using Boltzmann model equation

    Institute of Scientific and Technical Information of China (English)

    LI; Zhihui; ZHANG; Hanxin; FU; Song

    2005-01-01

    The gas-kinetic unified algorithm using Boltzmann model equation have been extended and developed to solve the micro-scale gas flows in Poiseuille-like micro-channels from Micro-Electro-Mechanical Systems (MEMS). The numerical modeling of the gas kinetic boundary conditions suitable for micro-scale gas flows is presented. To test the present method, the classical Couette flows with various Knudsen numbers, the gas flows from short microchannels like plane Poiseuille and the pressure-driven gas flows in two-dimensional short microchannels have been simulated and compared with the approximate solutions of the Boltzmann equation, the related DSMC results, the modified N-S solutions with slip-flow boundary theory, the gas-kinetic BGK-Burnett solutions and the experimental data. The comparisons show that the present gas-kinetic numerical algorithm using the mesoscopic Boltzmann simplified velocity distribution function equation can effectively simulate and reveal the gas flows in microchannels. The numerical experience indicates that this method may be a powerful tool in the numerical simulation of micro-scale gas flows from MEMS.

  2. Non-equilibrium reacting gas flows kinetic theory of transport and relaxation processes

    CERN Document Server

    Nagnibeda, Ekaterina; Nagnibeda, Ekaterina

    2009-01-01

    This volume develops the kinetic theory of transport phenomena and relaxation processes in the flows of reacting gas mixtures. The theory is applied to the modeling of non-equilibrium flows behind strong shock waves, in the boundary layer, and in nozzles.

  3. Cytometric Catheter for Neurosurgical Applications

    Energy Technology Data Exchange (ETDEWEB)

    Evans III, Boyd Mccutchen [ORNL; Allison, Stephen W [ORNL; Fillmore, Helen [ORNL; Broaddus, William C [ORNL; Dyer, Rachel L [ORNL; Gillies, George [ORNL

    2010-01-01

    Implantation of neural progenitor cells into the central nervous system has attracted strong interest for treatment of a variety of pathologies. For example, the replacement of dopamine-producing (DA) neural cells in the brain appears promising for the treatment of patients affected by Parkinson's disease. Previous studies of cell-replacement strategies have shown that less than 90% of implanted cells survive longer than 24 - 48 hours following the implantation procedure. However, it is unknown if these cells were viable upon delivery, or if they were affected by other factors such as brain pathology or an immune response. An instrumented cell-delivery catheter has been developed to assist in answering these questions by facilitating quantification and monitoring of the viability of the cells delivered. The catheter uses a fiber optic probe to perform flourescence-based cytometric measurments on cells exiting the port at the catheter tip. The current implementation of this design is on a 3.2 mm diameter catheter with 245 micrometer diameter optical fibers. Results of fluorescence testing data are presented and show that the device can characterize the quantity of cell densities ranging from 60,000 cells/ml to 600,000 cells/ml with a coefficient of determination of 0.93.

  4. Kinetic modelling of nitrogen and organics removal in vertical and horizontal flow wetlands.

    Science.gov (United States)

    Saeed, Tanveer; Sun, Guangzhi

    2011-05-01

    This paper provides a comparative evaluation of the kinetic models that were developed to describe the biodegradation of nitrogen and organics removal in wetland systems. Reaction kinetics that were considered in the model development included first order kinetics, Monod and multiple Monod kinetics; these kinetics were combined with continuous-stirred tank reactor (CSTR) or plug flow pattern to produce equations to link inlet and outlet concentrations of each key pollutants across a single wetland. Using three statistical parameters, a critical evaluation of five potential models was made for vertical and horizontal flow wetlands. The results recommended the models that were developed based on Monod models, for predicting the removal of nitrogen and organics in a vertical and horizontal flow wetland system. No clear correlation was observed between influent BOD/COD values and kinetic coefficients of BOD(5) in VF and HF wetlands, illustrating that the removal of biodegradable organics was insensitive to the nature of organic matter. Higher effluent COD/TN values coincided with greater denitrification kinetic coefficients, signifying the dependency of denitrification on the availability of COD in VF wetland systems. In contrast, the trend was opposite in HF wetlands, indicating that availability of NO(3)-N was the main limiting step for nitrogen removal. Overall, the results suggested the possible application of the developed alternative predictive models, for understanding the complex biodegradation routes of nitrogen and organics removal in VF and HF wetland systems.

  5. An atmospheric pressure flow reactor: Gas phase kinetics and mechanism in tropospheric conditions without wall effects

    Science.gov (United States)

    Koontz, Steven L.; Davis, Dennis D.; Hansen, Merrill

    1988-01-01

    A new type of gas phase flow reactor, designed to permit the study of gas phase reactions near 1 atm of pressure, is described. A general solution to the flow/diffusion/reaction equations describing reactor performance under pseudo-first-order kinetic conditions is presented along with a discussion of critical reactor parameters and reactor limitations. The results of numerical simulations of the reactions of ozone with monomethylhydrazine and hydrazine are discussed, and performance data from a prototype flow reactor are presented.

  6. A Gas-Kinetic Scheme for Multimaterial Flows and Its Application in Chemical Reaction

    Science.gov (United States)

    Lian, Yongsheng; Xu, Kun

    1999-01-01

    This paper concerns the extension of the multicomponent gas-kinetic BGK-type scheme to multidimensional chemical reactive flow calculations. In the kinetic model, each component satisfies its individual gas-kinetic BGK equation and the equilibrium states of both components are coupled in space and time due to the momentum and energy exchange in the course of particle collisions. At the same time, according to the chemical reaction rule one component can be changed into another component with the release of energy, where the reactant and product could have different gamma. Many numerical test cases are included in this paper, which show the robustness and accuracy of kinetic approach in the description of multicomponent reactive flows.

  7. Self-powered water splitting using flowing kinetic energy.

    Science.gov (United States)

    Tang, Wei; Han, Yu; Han, Chang Bao; Gao, Cai Zhen; Cao, Xia; Wang, Zhong Lin

    2015-01-14

    By utilizing a water-flow-driven triboelectric nanogenerator, a fully self-powered water-splitting process is demonstrated using the electricity converted from a water flow without additional energy costs. Considering the extremely low costs, the demonstrated approach is universally applicable and practically usable for future water electrolysis, which may initiate a research direction in the field of triboelectrolysis and possibly impacts energy science in general. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. On kinetic Boltzmann equations and related hydrodynamic flows with dry viscosity

    Directory of Open Access Journals (Sweden)

    Nikolai N. Bogoliubov (Jr.

    2007-01-01

    Full Text Available A two-component particle model of Boltzmann-Vlasov type kinetic equations in the form of special nonlinear integro-differential hydrodynamic systems on an infinite-dimensional functional manifold is discussed. We show that such systems are naturally connected with the nonlinear kinetic Boltzmann-Vlasov equations for some one-dimensional particle flows with pointwise interaction potential between particles. A new type of hydrodynamic two-component Benney equations is constructed and their Hamiltonian structure is analyzed.

  9. Kinetic description of bottleneck effects in traffic flow

    Institute of Scientific and Technical Information of China (English)

    Peng ZHANG; Dong-yan WU; S. C. WONG; Yi-zhou TAO

    2009-01-01

    This paper deals with the effects of traffic bottlenecks using an extended Lighthill-Whitham-Richards (LWR) model. The solution structure is analytically indi-cated by the study of the Riemann problem characterized by a discontinuous flux. This leads to a typical solution describing a queue upstream of the bottleneck and its width and height, and informs the design of a 5-mapping algorithm. More significantly, it is found that the kinetic model is able to reproduce stop-and-go waves for a triangular fun-damental diagram. Some simulation examples, which are in agreement with the analytical solutions, are given to support these conclusions.

  10. Analysis of atmospheric flow over a surface protrusion using the turbulence kinetic energy equation

    Science.gov (United States)

    Frost, W.; Harper, W. L.; Fichtl, G. H.

    1975-01-01

    Atmospheric flow fields resulting from a semi-elliptical surface obstruction in an otherwise horizontally homogeneous statistically stationary flow are modelled with the boundary-layer/Boussinesq-approximation of the governing equation of fluid mechanics. The turbulence kinetic energy equation is used to determine the dissipative effects of turbulent shear on the mean flow. Mean-flow results are compared with those given in a previous paper where the same problem was attacked using a Prandtl mixing-length hypothesis. Iso-lines of turbulence kinetic energy and turbulence intensity are plotted in the plane of the flow. They highlight regions of high turbulence intensity in the stagnation zone and sharp gradients in intensity along the transition from adverse to favourable pressure gradient.

  11. Flow cytometric analysis of mitotic cycle perturbation by chemical carcinogens in cultured epithelial cells. [Effects of benzo(a)pyrene-diol-epoxide on mitotic cycle of cultural mouse liver epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Pearlman, A.L.

    1978-08-01

    A system for kinetic analysis of mitotic cycle perturbation by various agents was developed and applied to the study of the mitotic cycle effects and dependency of the chemical carcinogen benzo(a)pyrene-diolepoxide, DE, upon a mouse lever epithelial cell line, NMuLi. The study suggests that the targets of DE action are not confined to DNA alone but may include cytoplasmic structures as well. DE was found to affect cells located in virtually every phase of the mitotic cycle, with cells that were actively synthesizing DNA showing the strongest response. However, the resulting perturbations were not confined to S-phase alone. DE slowed traversal through S-phase by about 40% regardless of the cycle phase of the cells exposed to it, and slowed traversal through G/sub 2/M by about 50%. When added to G/sub 1/ cells, DE delayed recruitment of apparently quiescent (G/sub 0/) cells by 2 hours, and reduced the synchrony of the cohort of cells recruited into active proliferation. The kinetic analysis system consists of four elements: tissue culture methods for propagating and harvesting cell populations; an elutriation centrifugation system for bulk synchronization of cells in various phases of the mitotic cycle; a flow cytometer (FCM), coupled with appropriate staining protocols, to enable rapid analysis of the DNA distribution of any given cell population; and data reduction and analysis methods for extracting information from the DNA histograms produced by the FCM. The elements of the system are discussed. A mathematical analysis of DNA histograms obtained by FCM is presented. The analysis leads to the detailed implementation of a new modeling approach. The new modeling approach is applied to the estimation of cell cycle kinetic parameters from time series of DNA histograms, and methods for the reduction and interpretation of such series are suggested.

  12. Immune complex stimulation of human neutrophils involves a novel Ca2+/H+ exchanger that participates in the regulation of cytoplasmic pH: flow cytometric analysis of Ca2+/pH responses by subpopulations.

    Science.gov (United States)

    Bernardo, John; Hartlaub, Hilary; Yu, Xin; Long, Heidi; Simons, Elizabeth R

    2002-12-01

    The activation of human phagocytic leukocytes by immune complexes (IC) or opsonized microbes via their Fc and complement receptors has been well-described. The mechanisms involved in this process are complex and depend on the receptors involved. The biochemical events that lead to the destruction of invading organisms in turn display varying degrees of interdependence, but the controlling elements that lead to the ultimate killing of ingested organisms within phagosomes by lysosomal enzymes and reactive oxygen intermediates are still not completely understood. We have addressed these mechanisms by following and correlating the kinetics of responses by individual cells, using multiparameter flow cytometry. Using nonopsonized IC as stimuli, we document here the presence of a novel Ca(2)(+)/H(+) voltage-independent channel in human neutrophils, which helps to control their cytoplasmic pH.

  13. Kinetic lattice Boltzmann method for microscale gas flows: issues on boundary condition, relaxation time, and regularization.

    Science.gov (United States)

    Niu, Xiao-Dong; Hyodo, Shi-Aki; Munekata, Toshihisa; Suga, Kazuhiko

    2007-09-01

    It is well known that the Navier-Stokes equations cannot adequately describe gas flows in the transition and free-molecular regimes. In these regimes, the Boltzmann equation (BE) of kinetic theory is invoked to govern the flows. However, this equation cannot be solved easily, either by analytical techniques or by numerical methods. Hence, in order to efficiently maneuver around this equation for modeling microscale gas flows, a kinetic lattice Boltzmann method (LBM) has been introduced in recent years. This method is regarded as a numerical approach for solving the BE in discrete velocity space with Gauss-Hermite quadrature. In this paper, a systematic description of the kinetic LBM, including the lattice Boltzmann equation, the diffuse-scattering boundary condition for gas-surface interactions, and definition of the relaxation time, is provided. To capture the nonlinear effects due to the high-order moments and wall boundaries, an effective relaxation time and a modified regularization procedure of the nonequilibrium part of the distribution function are further presented based on previous work [Guo et al., J. Appl. Phys. 99, 074903 (2006); Shan et al., J. Fluid Mech. 550, 413 (2006)]. The capability of the kinetic LBM of simulating microscale gas flows is illustrated based on the numerical investigations of micro Couette and force-driven Poiseuille flows.

  14. Kinetic modeling of temperature driven flows in short microchannels

    Energy Technology Data Exchange (ETDEWEB)

    Alexeenko, Alina A.; Gimelshein, Sergey F.; Muntz, E. Phillip [Department of Aerospace and Mechanical Engineering, University of Southern California, Los Angeles, CA 90089 (United States); Ketsdever, Andrew D. [Propulsion Directorate, Air Force Research Laboratory, Edwards Air Force Base, CA 93524 (United States)

    2006-11-15

    The temperature driven gas flows in both a two-dimensional finite length microchannel and a cylindrical tube have been studied numerically, with a goal of investigating performance optimization for a nano-membrane-based Knudsen Compressor. The numerical solutions were obtained using the direct simulation Monte Carlo (DSMC) method and a discrete ordinate method for the ellipsoidal statistical and Bhatnagar-Gross-Krook models. The Knudsen number was 0.2 and the length-to-height ratio 5. Three different wall temperature distributions were considered: linear, step-wise, and a non-monotonic profile typical for a radiantly heated Knudsen Compressor's membrane. The short channel end effects are characterized, and the sensitivity of the mass flow to a non-monotonic temperature distribution is shown. (author)

  15. Reaction kinetics of solid fuels during entrained flow gasification

    Energy Technology Data Exchange (ETDEWEB)

    Tremel, Alexander

    2012-10-24

    Despite the application of entrained flow gasification on larger scales, the reaction rates in the hot conversion zone are almost unknown. But the knowledge of the rates of the gasification reactions at high temperature and high pressure is crucial for the detailed design and optimisation of these gasifiers. This dissertation provides measurements of fuel conversion under operation conditions relevant to industrial gasifiers and aims at the transfer of the data to larger scale applications. A novel pilot-scale research reactor is developed that enables the study of gasification reactions at high temperature, high pressure and under entrained flow conditions. The Pressurised High Temperature Entrained Flow Reactor (PiTER) is operated at up to 1600 C and 4.0 MPa in pyrolysis and gasification experiments. The data set is extended by measurements in an atmospheric entrained flow reactor and a pressurised wire mesh reactor. Devolatilisation and gasification behaviour of a wide range of fuels is analysed including anthracite, bituminous coal, lignite, biocoal (from hydrothermal carbonisation), and biomass; however, Rhenish lignite is used in most of the experiments. The pyrolysis data enable the validation of a simple first order reaction model that describes the influence of pressure, temperature, and residence time on volatile yield. Char samples collected from the three reactors are analysed using laboratory procedures and bench-scale setups. Specific char surface area is measured by CO{sub 2} adsorption at 273 K, and is found to be significantly influenced by char conversion, reaction temperature, and devolatilisation pressure. The surface data are described by an extension of the Random Pore Model. Intrinsic char reactivity is measured in a pressurised thermogravimetric analyser and the influence of reactant partial pressure and temperature on the char-CO{sub 2} and char-H{sub 2}O reaction is studied. The intrinsic reaction rate is described by nth order and

  16. Effect on growth and cell cycle kinetics of estradiol and tamoxifen on MCF-7 human breast cancer cells grown in vitro and in nude mice

    DEFF Research Database (Denmark)

    Brünner, N; Bronzert, D; Vindeløv, L L

    1989-01-01

    determined by repeated flow cytometric DNA analyses in vitro and in vivo and by the technique of labeled mitosis in nude mouse-grown tumors. Under in vitro conditions, estradiol induced a pronounced increase in S-phase fraction and cell number. TAM inhibited growth of MCF-7 cells with a concomitant increase...... cytometric DNA analysis and percentage of labeled mitosis investigations revealed no significant differences in the proliferation kinetics of TAM-treated and control tumors. Calculating the cell loss factor demonstrated an increase from 69% in control tumors to 107% in TAM-treated tumors. These experiments...

  17. Determination of rapid chlorination rate constants by a stopped-flow spectrophotometric competition kinetics method.

    Science.gov (United States)

    Song, Dean; Liu, Huijuan; Qiang, Zhimin; Qu, Jiuhui

    2014-05-15

    Free chlorine is extensively used for water and wastewater disinfection nowadays. However, it still remains a big challenge to determine the rate constants of rapid chlorination reactions although competition kinetics and stopped-flow spectrophotometric (SFS) methods have been employed individually to investigate fast reaction kinetics. In this work, we proposed an SFS competition kinetics method to determine the rapid chlorination rate constants by using a common colorimetric reagent, N,N-diethyl-p-phenylenediamine (DPD), as a reference probe. A kinetic equation was first derived to estimate the reaction rate constant of DPD towards chlorine under a given pH and temperature condition. Then, on that basis, an SFS competition kinetics method was proposed to determine directly the chlorination rate constants of several representative compounds including tetracycline, ammonia, and four α-amino acids. Although Cl2O is more reactive than HOCl, its contribution to the overall chlorination kinetics of the test compounds could be neglected in this study. Finally, the developed method was validated through comparing the experimentally measured chlorination rate constants of the selected compounds with those obtained or calculated from literature and analyzing with Taft's correlation as well. This study demonstrates that the SFS competition kinetics method can measure the chlorination rate constants of a test compound rapidly and accurately.

  18. The propagation of kinetic energy across scales in turbulent flows

    CERN Document Server

    Cardesa, José I; Dong, Siwei; Jiménez, Javier

    2015-01-01

    A temporal study of energy transfer across length scales is performed in 3D numerical simulations of homogeneous shear flow and isotropic turbulence, at Reynolds numbers in the range $Re_{\\lambda}=107-384$. The average time taken by perturbations in the energy flux to travel between scales is measured and shown to be additive, as inferred from the agreement between the total travel time from a given scale to the smallest dissipative motions, and the time estimated from successive jumps through intermediate scales. Our data suggests that the propagation of disturbances in the energy flux is independent of the forcing and that it defines a `velocity' that determines the energy flux itself. These results support that the cascade is, on average, a scale-local process where energy is continuously transmitted from one scale to the next in order of decreasing size.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. Cytometric patterns reveal growth states of Shewanella putrefaciens.

    Science.gov (United States)

    Melzer, Susanne; Winter, Gudrun; Jäger, Kathrin; Hübschmann, Thomas; Hause, Gerd; Syrowatka, Frank; Harms, Hauke; Tárnok, Attila; Müller, Susann

    2015-05-01

    Bacterial growth is often difficult to estimate beyond classical cultivation approaches. Low cell numbers, particles or coloured and dense media may disturb reliable growth assessment. Further difficulties appear when cells are attached to surfaces and detachment is incomplete. Therefore, flow cytometry was tested and used for analysis of bacterial growth on the single-cell level. Shewanella putrefaciens was cultivated as a model organism in planktonic or biofilm culture. Materials of smooth and rough surfaces were used for biofilm cultivation. Both aerobic and anaerobic as well as feast and famine conditions were applied. Visualization of growth was also done using Environmental Scanning and Phase Contrast Microscopy. Bioinformatic tools were applied for data interpretation. Cytometric proliferation patterns based on distributions of DNA contents per cell corresponded distinctly to the various lifestyles, electron acceptors and substrates tested. Therefore, cell cycling profiles of S. putrefaciens were found to mirror growth conditions. The cytometric patterns were consistently detectable with exception of some biofilm types whose resolution remained challenging. Corresponding heat maps proved to be useful for clear visualization of growth behaviour under all tested conditions. Therefore, flow cytometry in combination with bioinformatic tools proved to be powerful means to determine various growth states of S. putrefaciens, even in constrained environments. The approach is universal and will also be applicable for other bacterial species. © 2014 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  1. Second Law Analysis for a Variable Viscosity Reactive Couette Flow under Arrhenius Kinetics

    Directory of Open Access Journals (Sweden)

    N. S. Kobo

    2010-01-01

    Full Text Available This study investigates the inherent irreversibility associated with the Couette flow of a reacting variable viscosity combustible material under Arrhenius kinetics. The nonlinear equations of momentum and energy governing the flow system are solved both analytically using a perturbation method and numerically using the standard Newton Raphson shooting method along with a fourth-order Runge Kutta integration algorithm to obtain the velocity and temperature distributions which essentially expedite to obtain expressions for volumetric entropy generation numbers, irreversibility distribution ratio, and the Bejan number in the flow field.

  2. Copper and cobalt complexes of octadentate azamacrocycles: spectrophotometric titration, stopped-flow kinetics and crystallographic study.

    Science.gov (United States)

    Ozay, Hava; Baran, Yakup; Ishii, Youichi

    2011-12-01

    Details of complex formation kinetics are reported for tetrakis(2-hydroxyethyl) substituted cyclen (L(1)) and cyclam (L(2)) with Cu(II) and Co(II). Stopped-flow kinetics and spectroscopic titration methods were employed for the activation parameters and stability constants, respectively. X-ray studies revealed that the pendant 2-hydroxyethyl groups are not equivalent: two are folded over the macrocycle and maintained by intramolecular hydrogen bonds while the others are extended and pointed away from the macrocyclic cavity. Complex formation kinetics and spectroscopic titration were performed in aqueous acidic buffer solutions. Thermodynamic and kinetic parameters revealed that the ring size of the macrocycles plays an extremely important role for each metal ion studied. Stopped-flow kinetic measurements explained the mechanism of the complex formation process of both Cu(II) and Co(II) which proceed in outer-sphere interactions with ligands. There are two steps in the complex formation of the system studied. The initial step is a second order reaction between the metal ion and macrocycle with a second order rate constant.

  3. Properties of the kinetic energy budgets in wall-bounded turbulent flows

    Science.gov (United States)

    Zhou, Ang; Klewicki, Joseph

    2016-08-01

    Available high-quality numerical simulation data are used to investigate and characterize the kinetic energy budgets for fully developed turbulent flow in pipes and channels, and in the zero-pressure gradient turbulent boundary layer. The mean kinetic energy equation in these flows is empirically and analytically shown to respectively exhibit the same four-layer leading-order balance structure as the mean momentum equation. This property of the mean kinetic energy budget provides guidance on how to group terms in the more complicated turbulence and total kinetic energy budgets. Under the suggested grouping, the turbulence budget shows either a two- or three-layer structure (depending on channel or pipe versus boundary layer flow), while the total kinetic energy budget exhibits a clear four-layer structure. These layers, however, differ in position and size and exhibit variations with friction Reynolds number (δ+) that are distinct from the layer structure associated with the mean dynamics. The present analyses indicate that each of the four layers is characterized by a predominance of a reduced set of the grouped terms in the governing equation. The width of the third layer is mathematically reasoned to scale like δ+-√{δ+} at finite Reynolds numbers. In the boundary layer the upper bounds of both the second and third layers convincingly merge under this normalization, as does the width of the third layer. This normalization also seems to be valid for the width of the third layer in pipes and channels, but only for δ+>1000 . The leading-order balances in the total kinetic energy budget are shown to arise from a nontrivial interweaving of the mean and turbulence budget contributions with distance from the wall.

  4. Measurement of the Turbulence Kinetic Energy Budget of a Turbulent Planar Wake Flow in Pressure Gradients

    Science.gov (United States)

    Liu, Xiao-Feng; Thomas, Flint O.; Nelson, Robert C.

    2001-01-01

    Turbulence kinetic energy (TKE) is a very important quantity for turbulence modeling and the budget of this quantity in its transport equation can provide insight into the flow physics. Turbulence kinetic energy budget measurements were conducted for a symmetric turbulent wake flow subjected to constant zero, favorable and adverse pressure gradients in year-three of research effort. The purpose of this study is to clarify the flow physics issues underlying the demonstrated influence of pressure gradient on wake development and provide experimental support for turbulence modeling. To ensure the reliability of these notoriously difficult measurements, the experimental procedure was carefully designed on the basis of an uncertainty analysis. Four different approaches, based on an isotropic turbulence assumption, a locally axisymmetric homogeneous turbulence assumption, a semi-isotropy assumption and a forced balance of the TKE equation, were applied for the estimate of the dissipation term. The pressure transport term is obtained from a forced balance of the turbulence kinetic energy equation. This report will present the results of the turbulence kinetic energy budget measurement and discuss their implication on the development of strained turbulent wakes.

  5. A kinetic theory treatment of heat transfer in plane Poiseuille flow with uniform pressure

    Science.gov (United States)

    Bahrami, Parviz A.

    1992-01-01

    Plane compressible Poiseuille flow with uniform pressure (Couette flow with stationary boundaries) is revisited where the Lees two-steam method with the Enskog equation of change is applied. Single particle velocity distribution functions are chosen, which preserve the essential physical features of this flow with arbitrary but uniform plate temperatures and gas pressure. Lower moments are shown to lead to expressions for the parameter functions, molecular number densities, and temperatures which are entirely in agreement with those obtained in the analysis of Lees for compressible plane Couette flow in the limit of low Mach number and vanishing mean gas velocity. Important simplifications result, which are helpful in gaining insight into the power of kinetic theory in fluid mechanics. The temperature distribution, heat flux, as well as density, are completely determined for the whole range of Knudson numbers from free molecular flow to the continuum regime, when the pressure level is specified.

  6. Large Eddy Simulation of SGS Turbulent Kinetic Energy and SGS Turbulent Dissipation in a Backward-Facing Step Turbulent Flow

    Institute of Scientific and Technical Information of China (English)

    王兵; 张会强; 王希麟

    2004-01-01

    The instantaneous and time-averaged statistic characteristics of the sub-grid scale (SGS) turbulent kinetic energy and SGS dissipation in a backward-facing step turbulent flow have been studied bylarge eddy simulation. The SGS turbulent kinetic energy and SGS turbulent dissipation vary in different flow regions and decrease with the flow developing spatially. The fluid molecular dissipation shares about 14% to 28% of the whole dissipation.

  7. Flowing afterglow spectroscopy: an ultrasensitive probe into solid-phase decomposition kinetics. [TATB and NQ

    Energy Technology Data Exchange (ETDEWEB)

    Taylor, G.W.; Andrews, G.H. Jr.

    1979-01-01

    The thermal-decomposition kinetics of high explosives are important to manufacturers and these dangerous materials from the standpoint of processing and storage. Low-temperature kinetics (20 to 200/sup 0/C) are difficult to obtain. Either extremely sensitive analytical techniques must be employed or very large amounts of material must be tested by less sensitive methods. The latter technique has the disadvantage that when sensitive explosives or propellants are so tested, catastrophic reaction can result in the destruction of expensive equipment, and the time involved in testing may be extremely long. The flowing-afterglow method, proposed here, utilizes small explosive samples and an ultra-sensitive analytical approach. The paper describes the technique in detail and summarizes our recent efforts to elucidate the low-temperature kinetics of trinitrotriaminobenzene (TATB) and nitroguanidine (NQ).

  8. Improved Soft Abrasive Flow Finishing Method Based on Turbulent Kinetic Energy Enhancing

    Science.gov (United States)

    LI, Jun; JI, Shiming; TAN, Dapeng

    2017-03-01

    Soft abrasive flow(SAF) finishing can process the irregular geometric surfaces, but with the matter of low processing efficiency. To address the issue, an improved SAF finishing method based on turbulent kinetic energy enhancing is proposed. A constrained flow passage with serration cross-section is constructed to increase the turbulence intensity. Taking the constrained flow passage as the objective, a two-phase fluid dynamic model is set up by using particle trajectory model and standard k-ɛ turbulence model, and the flow field characteristics of the flow passage are acquired. The numerical results show that the serration flow passage can enhance the turbulence intensity, uniform the particles distribution, and increase the particle concentration near the bottom wall. The observation results by particle image velocimetry(PIV) show that the internal vortex structures are formed in flow passage, and the abrasive flow takes on turbulence concentrating phenomenon in near-wall region. The finishing experiments prove that the proposed method can obtain better surface uniformity, and the processing efficiency can be improved more 35%. This research provides an abrasive flow modeling method to reveal the particle motion regulars, and can offer references to the technical optimization of fluid-based precision processing.

  9. Discrete unified gas kinetic scheme with force term for incompressible fluid flows

    CERN Document Server

    Wu, Chen; Chai, Zhenhua; Wang, Peng

    2014-01-01

    The discrete unified gas kinetic scheme (DUGKS) is a finite-volume scheme with discretization of particle velocity space, which combines the advantages of both lattice Boltzmann equation (LBE) method and unified gas kinetic scheme (UGKS) method, such as the simplified flux evaluation scheme, flexible mesh adaption and the asymptotic preserving properties. However, DUGKS is proposed for near incompressible fluid flows, the existing compressible effect may cause some serious errors in simulating incompressible problems. To diminish the compressible effect, in this paper a novel DUGKS model with external force is developed for incompressible fluid flows by modifying the approximation of Maxwellian distribution. Meanwhile, due to the pressure boundary scheme, which is wildly used in many applications, has not been constructed for DUGKS, the non-equilibrium extrapolation (NEQ) scheme for both velocity and pressure boundary conditions is introduced. To illustrate the potential of the proposed model, numerical simul...

  10. On Riemann Solvers and Kinetic Relations for Isothermal Two-Phase Flows with Surface Tension

    CERN Document Server

    Rohde, Christian

    2016-01-01

    We consider a sharp-interface approach for the inviscid isothermal dynamics of compressible two-phase flow, that accounts for phase transition and surface tension effects. To fix the mass exchange and entropy dissipation rate across the interface kinetic relations are frequently used. The complete uni-directional dynamics can then be understood by solving generalized two-phase Riemann problems. We present new well-posedness theorems for the Riemann problem and corresponding computable Riemann solvers, that cover quite general equations of state, metastable input data and curvature effects. The new Riemann solver is used to validate different kinetic relations on physically relevant problems including a comparison with experimental data. Riemann solvers are building blocks for many numerical schemes that are used to track interfaces in two-phase flow. It is shown that the new Riemann solver enables reliable and efficient computations for physical situations that could not be treated before.

  11. BRIEF COMMUNICATION: On the drift kinetic equation driven by plasma flows

    Science.gov (United States)

    Shaing, K. C.

    2010-07-01

    A drift kinetic equation that is driven by plasma flows has previously been derived by Shaing and Spong 1990 (Phys. Fluids B 2 1190). The terms that are driven by particle speed that is parallel to the magnetic field B have been neglected. Here, such terms are discussed to examine their importance to the equation and to show that these terms do not contribute to the calculations of plasma viscosity in large aspect ratio toroidal plasmas, e.g. tokamaks and stellarators.

  12. Interplay between density profile and zonal flows in drift kinetic simulations of slab ITG turbulence

    Energy Technology Data Exchange (ETDEWEB)

    Sarazin, Y.; Garbet, X.; Grandgirard, V.; Ghendrih, Ph. [Association Euratom-CEA Cadarache (DSM/DRFC), 13 - Saint-Paul-lez-Durance (France). Dept. de Recherches sur la Fusion Controlee; Bertrand, P. [Universite Henri Poincare, Laboratoire de Physique des Milieux Ionises et Applications (LPMIA), 54 - Vandoeuvre les Nancy (France); Besse, N.; Sonnendruecker, E. [Universite Louis Pasteur, CNRS IRMA, 67 - Strasbourg (France)

    2004-07-01

    This paper reports on 4-dimensional drift kinetic simulations of the slab branch of the Ion Temperature Gradient driven turbulence in a cylinder. In the non-linear regime, the system is found to relax preferentially either via heat transport or via mean sheared flows, depending on the density profile. A strong density gradient appears to be stabilizing both linearly, by increasing the instability threshold, and non linearly, by activating sheared flows. This impedes the relaxation of the profiles and sustains a pressure transport barrier. (authors)

  13. Abnormal early diastolic intraventricular flow 'kinetic energy index' assessed by vector flow mapping in patients with elevated filling pressure.

    Science.gov (United States)

    Nogami, Yoshie; Ishizu, Tomoko; Atsumi, Akiko; Yamamoto, Masayoshi; Kawamura, Ryo; Seo, Yoshihiro; Aonuma, Kazutaka

    2013-03-01

    Recently developed vector flow mapping (VFM) enables evaluation of local flow dynamics without angle dependency. This study used VFM to evaluate quantitatively the index of intraventricular haemodynamic kinetic energy in patients with left ventricular (LV) diastolic dysfunction and to compare those with normal subjects. We studied 25 patients with estimated high left atrial (LA) pressure (pseudonormal: PN group) and 36 normal subjects (control group). Left ventricle was divided into basal, mid, and apical segments. Intraventricular haemodynamic energy was evaluated in the dimension of speed, and it was defined as the kinetic energy index. We calculated this index and created time-energy index curves. The time interval from electrocardiogram (ECG) R wave to peak index was measured, and time differences of the peak index between basal and other segments were defined as ΔT-mid and ΔT-apex. In both groups, early diastolic peak kinetic energy index in mid and apical segments was significantly lower than that in the basal segment. Time to peak index did not differ in apex, mid, and basal segments in the control group but was significantly longer in the apex than that in the basal segment in the PN group. ΔT-mid and ΔT-apex were significantly larger in the PN group than the control group. Multiple regression analysis showed sphericity index, E/E' to be significant independent variables determining ΔT apex. Retarded apical kinetic energy fluid dynamics were detected using VFM and were closely associated with LV spherical remodelling in patients with high LA pressure.

  14. Kinetic characteristics of continuous flow polymerase chain reaction chip: A numerical investigation

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Continuous flow PCR (polymerase chain reaction) chip holds impressive advantages compared to micro chamber PCR chip. In order to have better understanding of kinetic characteristics of continuous flow PCR chip, a comprehensive mathematical model is presented in this paper, including melting, annealing and extension phases of a typical PCR process which has the essence of a convection-diffusion-reaction system. Using this model, we can simulate the PCR process in series of reaction cycles. Numerical results show that the average sample velocity plays a significant role in affecting the amplification efficiency. Also, appropriate combination of the PCR mixture is important for high-quality DNA amplification. Giving a large initial DNA concentration range, the continuous flow PCR scheme holds excellent real-time detection ability theoretically. The present numerical model bridges the temperature distribution to the real DNA amplification, and thereby is able to successfully predict continuous flow PCR properties which are important for the chip design.

  15. A Gas-Kinetic Scheme For The Simulation Of Compressible Turbulent Flows

    CERN Document Server

    Righi, Marcello

    2013-01-01

    A gas-kinetic scheme for the continuum regime is applied to the simulation of turbu- lent compressible flow, by replacing the molecular relaxation time with a turbulent relaxation time in the BGK model. The turbulence dynamics is modelled on the basis of a standard, linear two-equation turbulence model. The hydrodynamic limit of the resulting turbulence model is linear in smooth flow and non-linear in the presence of stronger flow gradients. The non-linear correction terms in the numerical flux are weighed as a function of "rarefaction" - referred to turbulence dynamics and not to molecular dynamics, i.e. measured by the ratio of turbulence to mean flow scales of motion. Even though no assumptions on the nature of the turbulence have been made and a linear two-equation turbulence model is used, the turbulence gas-kinetic scheme seems able to correct the turbulent stress tensor in an effective way; on the basis of a number of turbulence modelling benchmark flow cases, characterized by strong shock - boundary l...

  16. Kinetics-based phase change approach for VOF method applied to boiling flow

    Science.gov (United States)

    Cifani, Paolo; Geurts, Bernard; Kuerten, Hans

    2014-11-01

    Direct numerical simulations of boiling flows are performed to better understand the interaction of boiling phenomena with turbulence. The multiphase flow is simulated by solving a single set of equations for the whole flow field according to the one-fluid formulation, using a VOF interface capturing method. Interface terms, related to surface tension, interphase mass transfer and latent heat, are added at the phase boundary. The mass transfer rate across the interface is derived from kinetic theory and subsequently coupled with the continuum representation of the flow field. The numerical model was implemented in OpenFOAM and validated against 3 cases: evaporation of a spherical uniformly heated droplet, growth of a spherical bubble in a superheated liquid and two dimensional film boiling. The computational model will be used to investigate the change in turbulence intensity in a fully developed channel flow due to interaction with boiling heat and mass transfer. In particular, we will focus on the influence of the vapor bubble volume fraction on enhancing heat and mass transfer. Furthermore, we will investigate kinetic energy spectra in order to identify the dynamics associated with the wakes of vapor bubbles. Department of Applied Mathematics, 7500 AE Enschede, NL.

  17. A gas kinetic scheme for hybrid simulation of partially rarefied flows

    Science.gov (United States)

    Colonia, S.; Steijl, R.; Barakos, G.

    2017-06-01

    Approaches to predict flow fields that display rarefaction effects incur a cost in computational time and memory considerably higher than methods commonly employed for continuum flows. For this reason, to simulate flow fields where continuum and rarefied regimes coexist, hybrid techniques have been introduced. In the present work, analytically defined gas-kinetic schemes based on the Shakhov and Rykov models for monoatomic and diatomic gas flows, respectively, are proposed and evaluated with the aim to be used in the context of hybrid simulations. This should reduce the region where more expensive methods are needed by extending the validity of the continuum formulation. Moreover, since for high-speed rare¦ed gas flows it is necessary to take into account the nonequilibrium among the internal degrees of freedom, the extension of the approach to employ diatomic gas models including rotational relaxation process is a mandatory first step towards realistic simulations. Compared to previous works of Xu and coworkers, the presented scheme is de¦ned directly on the basis of kinetic models which involve a Prandtl number correction. Moreover, the methods are defined fully analytically instead of making use of Taylor expansion for the evaluation of the required derivatives. The scheme has been tested for various test cases and Mach numbers proving to produce reliable predictions in agreement with other approaches for near-continuum flows. Finally, the performance of the scheme, in terms of memory and computational time, compared to discrete velocity methods makes it a compelling alternative in place of more complex methods for hybrid simulations of weakly rarefied flows.

  18. Michaelis-Menten kinetics in shear flow: Similarity solutions for multi-step reactions.

    Science.gov (United States)

    Ristenpart, W D; Stone, H A

    2012-03-01

    Models for chemical reaction kinetics typically assume well-mixed conditions, in which chemical compositions change in time but are uniform in space. In contrast, many biological and microfluidic systems of interest involve non-uniform flows where gradients in flow velocity dynamically alter the effective reaction volume. Here, we present a theoretical framework for characterizing multi-step reactions that occur when an enzyme or enzymatic substrate is released from a flat solid surface into a linear shear flow. Similarity solutions are developed for situations where the reactions are sufficiently slow compared to a convective time scale, allowing a regular perturbation approach to be employed. For the specific case of Michaelis-Menten reactions, we establish that the transversally averaged concentration of product scales with the distance x downstream as x(5/3). We generalize the analysis to n-step reactions, and we discuss the implications for designing new microfluidic kinetic assays to probe the effect of flow on biochemical processes.

  19. Gas-Kinetic Navier-Stokes Solver for Hypersonic Flows in Thermal and Chemical Non-Equilibrium Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This SBIR project proposes to develop a gas-kinetic Navier-Stokes solver for simulation of hypersonic flows in thermal and chemical non-equilibrium. The...

  20. Development of discrete gas kinetic scheme for simulation of 3D viscous incompressible and compressible flows

    Science.gov (United States)

    Yang, L. M.; Shu, C.; Wang, Y.; Sun, Y.

    2016-08-01

    The sphere function-based gas kinetic scheme (GKS), which was presented by Shu and his coworkers [23] for simulation of inviscid compressible flows, is extended to simulate 3D viscous incompressible and compressible flows in this work. Firstly, we use certain discrete points to represent the spherical surface in the phase velocity space. Then, integrals along the spherical surface for conservation forms of moments, which are needed to recover 3D Navier-Stokes equations, are approximated by integral quadrature. The basic requirement is that these conservation forms of moments can be exactly satisfied by weighted summation of distribution functions at discrete points. It was found that the integral quadrature by eight discrete points on the spherical surface, which forms the D3Q8 discrete velocity model, can exactly match the integral. In this way, the conservative variables and numerical fluxes can be computed by weighted summation of distribution functions at eight discrete points. That is, the application of complicated formulations resultant from integrals can be replaced by a simple solution process. Several numerical examples including laminar flat plate boundary layer, 3D lid-driven cavity flow, steady flow through a 90° bending square duct, transonic flow around DPW-W1 wing and supersonic flow around NACA0012 airfoil are chosen to validate the proposed scheme. Numerical results demonstrate that the present scheme can provide reasonable numerical results for 3D viscous flows.

  1. Oral squamous cell carcinoma. Cytometric parameters of prognostic interest.

    Science.gov (United States)

    Saiz-Bustillo, Ramón; Corchero-Martín, Guadalupe; García-Montesinos-Perea, Belén; Gonzalez-Terán, Tomás; Sánchez-Santolino, Sergio

    2005-01-01

    The present study was made in order to find possible prognostic factors in oral squamous cell carcinoma, given that it is a frequent disease (3-4% of all malignant tumors) and is the cause of a high morbidity and mortality which justifies any attempt to contribute something towards the understanding of this pathology. 81 oral squamous cell carcinomas, treated with the same procedure, and retrieved from the archive of the Hospital Universitario Marqués de Valdecilla (Santander) were studied. Flow cytometry was carried out on 67 of the samples. No statistically significant differences were found between the cellular proliferative index and the mitotic index, ploidy and the S-phase factor. Likewise, none of the cytometric variables studied presented any association with the appearance of local relapse, distant metastases or survival. These variables cannot be used as a prognostic factors in squamous cell carcinomas of the oral cavity.

  2. Discrepancy between femoral and capillary blood flow kinetics during knee extension exercise.

    Science.gov (United States)

    Schlup, S J; Ade, C J; Broxterman, R M; Barstow, T J

    2015-12-01

    Capillary blood flow (QCAP) kinetics have previously been shown to be significantly slower than femoral artery (QFA) kinetics following the onset of dynamic knee extension exercise. If the increase in QCAP does not follow a similar time course to QFA, then a substantial proportion of the available blood flow is not distributed to the working muscle. One possible explanation for this discrepancy is that blood flow also increases to the nonworking lower leg muscles. Therefore, the present study aimed to determine if a reduction in lower limb blood flow, via arterial occlusion below the knee, alters the kinetics of QFA and QCAP during knee extension exercise, and thus provide insight into the potential mechanisms controlling the rapid increase in QFA. Subjects performed a ramp max test to determine the work rate at which gas exchange threshold (GET) occurred. At least four constant work rate trials with and without below-knee occlusion were conducted at work rates eliciting ∼ 80% GET. Pulmonary gas exchange, near-infrared spectroscopy and QFA measurements were taken continuously during each exercise bout. Muscle oxygen uptake (VO2m) and deoxy[hemoglobin+myoglobin] were used to estimate QCAP. There was no significant difference between the uncuffed and cuffed conditions in any response (P>0.05). The mean response times (MRT) of QFA were 18.7 ± 14.2s (uncuffed) and 24.6 ± 14.9s (cuffed). QCAP MRTs were 51.8 ± 23.4s (uncuffed) and 56.7 ± 23.2s (cuffed), which were not significantly different from the time constants (τ) of VO2m (39.7 ± 23.2s (uncuffed) and 46.3 ± 24.1s (cuffed). However, the MRT of QFA was significantly faster (P<0.05) than the MRT of QCAP and τVO2m. τVO2m and MRT QCAP were significantly correlated and estimated QCAP kinetics tracked VO2m following exercise onset. Cuffing below the knee did not significantly change the kinetics of QFA, QCAP or VO2m, although an effect size of 1.02 suggested that a significant effect on QFA may have been hidden

  3. Elliptic flow at SPS and RHIC from kinetic transport to hydrodynamics

    CERN Document Server

    Kolb, P F; Heinz, Ulrich W; Heiselberg, H

    2001-01-01

    Anisotropic transverse flow is studied in Pb+Pb and Au+Au collisions at SPS and RHIC energies. The centrality and transverse momentum dependence at midrapidity of the elliptic flow coefficient v_2 is calculated in the hydrodynamic and low density limits. Hydrodynamics is found to agree well with the RHIC data for semicentral collisions up to transverse momenta of 1-1.5 GeV/c, but it considerably overestimates the measured elliptic flow at SPS energies. The low density limit LDL is inconsistent with the measured magnitude of v_2 at RHIC energies and with the shape of its p_t-dependence at both RHIC and SPS energies. The success of the hydrodynamic model points to very rapid thermalization in Au+Au collisions at RHIC and provides a serious challenge for kinetic approaches based on classical scattering of on-shell particles.

  4. Gas-kinetic numerical schemes for one- and two-dimensional inner flows

    Institute of Scientific and Technical Information of China (English)

    Zhi-hui LI; Lin BI; Zhi-gong TANG

    2009-01-01

    Several kinds of explicit and implicit finite-difference schemes directly solving the discretized velocity distribution functions are designed with precision of different orders by analyzing the inner characteristics of the gas-kinetic numerical algorithm for Boltzmann model equation.The peculiar flow phenomena and mechanism from various flow regimes are revealed in the numerical simulations of the unsteady Sod shock-tube problems and the two-dimensional channel flows with different Knudsen numbers.The numerical remainder-effects of the difference schemes are investigated and analyzed based on the computed results.The ways of improving the computational efficiency of the gaskinetic numerical method and the computing principles of difference discretization are discussed.

  5. Mitigating the Hook Effect in Lateral Flow Sandwich Immunoassays Using Real-Time Reaction Kinetics.

    Science.gov (United States)

    Rey, Elizabeth G; O'Dell, Dakota; Mehta, Saurabh; Erickson, David

    2017-05-02

    The quantification of analyte concentrations using lateral flow assays is a low-cost and user-friendly alternative to traditional lab-based assays. However, sandwich-type immunoassays are often limited by the high-dose hook effect, which causes falsely low results when analytes are present at very high concentrations. In this paper, we present a reaction kinetics-based technique that solves this problem, significantly increasing the dynamic range of these devices. With the use of a traditional sandwich lateral flow immunoassay, a portable imaging device, and a mobile interface, we demonstrate the technique by quantifying C-reactive protein concentrations in human serum over a large portion of the physiological range. The technique could be applied to any hook effect-limited sandwich lateral flow assay and has a high level of accuracy even in the hook effect range.

  6. DENSE MULTIPHASE FLOW SIMULATION: CONTINUUM MODEL FOR POLY-DISPERSED SYSTEMS USING KINETIC THEORY

    Energy Technology Data Exchange (ETDEWEB)

    Moses Bogere

    2011-08-31

    The overall objective of the project was to verify the applicability of the FCMOM approach to the kinetic equations describing the particle flow dynamics. For monodispersed systems the fundamental equation governing the particle flow dynamics is the Boltzmann equation. During the project, the FCMOM was successfully applied to several homogeneous and in-homogeneous problems in different flow regimes, demonstrating that the FCMOM has the potential to be used to solve efficiently the Boltzmann equation. However, some relevant issues still need to be resolved, i.e. the homogeneous cooling problem (inelastic particles cases) and the transition between different regimes. In this report, the results obtained in homogeneous conditions are discussed first. Then a discussion of the validation results for in-homogeneous conditions is provided. And finally, a discussion will be provided about the transition between different regimes. Alongside the work on development of FCMOM approach studies were undertaken in order to provide insights into anisotropy or particles kinetics in riser hydrodynamics. This report includes results of studies of multiphase flow with unequal granular temperatures and analysis of momentum re-distribution in risers due to particle-particle and fluid-particle interactions. The study of multiphase flow with unequal granular temperatures entailed both simulation and experimental studies of two particles sizes in a riser and, a brief discussion of what was accomplished will be provided. And finally, a discussion of the analysis done on momentum re-distribution of gas-particles flow in risers will be provided. In particular a discussion of the remaining work needed in order to improve accuracy and predictability of riser hydrodynamics based on two-fluid models and how they can be used to model segregation in risers.

  7. Skeletal blood flow, iliac histomorphometry, and strontium kinetics in osteoporosis: a relationship between blood flow and corrected apposition rate

    Energy Technology Data Exchange (ETDEWEB)

    Reeve, J.; Arlot, M.; Wootton, R.; Edouard, C.; Tellez, M.; Hesp, R.; Green, J.R.; Meunier, P.J.

    1988-06-01

    In 20 untreated patients with idiopathic or postmenopausal osteoporosis, kinetic studies of skeletal blood flow (using /sup 18/F) and bone turnover (using /sup 85/Sr) were combined with dynamic histomorphometry performed on transiliac biopsies taken within 6 weeks of each other. In 8 patients the combined studies were repeated after treatment. A further 5 patients were studied only while receiving treatment. As expected, skeletal blood flow measured by /sup 18/F correlated with an index of /sup 85/Sr uptake into the exchangeable pools of bone. Additionally and independently, skeletal blood flow correlated with an index of the work rate of the osteoblasts in each multicellular unit of bone (the corrected apposition rate of Parfitt). These correlations were statistically significant in both the untreated patients (P less than 0.05) and the whole group (P less than 0.001). Further indices related to bone turnover at the level of the skeleton as a whole were significantly associated with skeletal blood flow only in the combined group.

  8. Glutathione transferases immobilized on nanoporous alumina: flow system kinetics, screening, and stability.

    Science.gov (United States)

    Kjellander, Marcus; Mazari, Aslam M A; Boman, Mats; Mannervik, Bengt; Johansson, Gunnar

    2014-02-01

    The previously uncharacterized Drosophila melanogaster Epsilon-class glutathione transferases E6 and E7 were immobilized on nanoporous alumina. The nanoporous anodized alumina membranes were derivatized with 3-aminopropyl-triethoxysilane, and the amino groups were activated with carbonyldiimidazole to allow coupling of the enzymes via ε-amino groups. Kinetic analyses of the immobilized enzymes were carried out in a circulating flow system using CDNB (1-chloro-2,4-dinitrobenzene) as substrate, followed by specificity screening with alternative substrates. A good correlation was observed between the substrate screening data for immobilized enzyme and corresponding data for the enzyme in solution. A limited kinetic study was also carried out on immobilized human GST S1-1 (also known as hematopoietic prostaglandin D synthase). The stability of the immobilized enzymes was virtually identical to that of enzymes in solution, and no leakage of enzyme from the matrix could be observed.

  9. Chemical kinetic modeling of a methane opposed flow diffusion flame and comparison to experiments

    Energy Technology Data Exchange (ETDEWEB)

    Marinov, N.M., Pitz, W.J.; Westbrook, C.K. [Lawrence Livermore National Lab., CA (United States); Vincitore, A.M.; Senka, S.M. [Univ. of California, Los Angeles, CA (United States); Lutz, A.E. [Sandia National Labs., Livermore, CA (United States)

    1998-01-01

    The chemical structure of an opposed flow, methane diffusion flame is studied using a chemical kinetic model and the results are compared to experimental measurements. The chemical kinetic paths leading to aromatics and polycyclic aromatics hydrocarbons (PAHs) in the diffusion flame are identified. These paths all involve resonantly stabilized radicals which include propargyl, allyl, cyclopentadienyl, and benzyl radicals. The modeling results show reasonable agreement with the experimental measurements for the large hydrocarbon aliphatic compounds, aromatics, and PAHs. the benzene was predicted to be formed primarily by the reaction sequence of Allyl plus Propargyl equals Fulvene plus H plus H followed by fulvene isomerization to benzene. Naphthalene was modeled using the reaction of benzyl with propargyl, while the combination of cyclopentadienyl radicals were shown to be a minor contributor in the diffusion flame. The agreement between the model and experiment for the four-ring PAHs was poor.

  10. Estimating kinetic mass transfer by resting-period measurements in flow-interruption tracer tests.

    Science.gov (United States)

    Gong, R; Lu, C; Wu, W-M; Cheng, H; Gu, B; Watson, D B; Criddle, C S; Kitanidis, P K; Brooks, S C; Jardine, P M; Luo, J

    2010-09-20

    Flow-interruption tracer test is an effective approach to identify kinetic mass transfer processes for solute transport in subsurface media. By switching well pumping and resting, one may alter the dominant transport mechanism and generate special concentration patterns for identifying kinetic mass transfer processes. In the present research, we conducted three-phase (i.e., pumping, resting, and pumping) field-scale flow-interruption tracer tests using a conservative tracer bromide in a multiple-well system installed at the US Department of Energy Site, Oak Ridge, TN. A novel modeling approach based on the resting-period measurements was developed to estimate the mass transfer parameters. This approach completely relied on the measured breakthrough curves without requiring detailed aquifer characterization and solving transport equations in nonuniform, transient flow fields. Additional measurements, including hydraulic heads and tracer concentrations in large pumping wells, were taken to justify the assumption that mass transfer processes dominated concentration change during resting periods. The developed approach can be conveniently applied to any linear mass transfer model. Both first-order and multirate mass transfer models were applied to analyze the breakthrough curves at various monitoring wells. The multirate mass transfer model was capable of jointly fitting breakthrough curve behavior, showing the effectiveness and flexibility for incorporating aquifer heterogeneity and scale effects in upscaling effective mass transfer models.

  11. Discrete unified gas kinetic scheme for all Knudsen number flows: II. Compressible case

    CERN Document Server

    Guo, Zhaoli; Xu, Kun

    2014-01-01

    This paper is a continuation of our earlier work [Z.L. Guo {\\it et al.}, Phys. Rev. E {\\bf 88}, 033305 (2013)] where a multiscale numerical scheme based on kinetic model was developed for low speed isothermal flows with arbitrary Knudsen numbers. In this work, a discrete unified gas-kinetic scheme (DUGKS) for compressible flows with the consideration of heat transfer and shock discontinuity is developed based on the Shakhov model with an adjustable Prandtl number. The method is an explicit finite-volume scheme where the transport and collision processes are coupled in the evaluation of the fluxes at cell interfaces, so that the nice asymptotic preserving (AP) property is retained, such that the time step is limited only by the CFL number, the distribution function at cell interface recovers to the Chapman-Enskog one in the continuum limit while reduces to that of free-transport for free-molecular flow, and the time and spatial accuracy is of second-order accuracy in smooth region. These features make the DUGK...

  12. Kinetic performance limits of constant pressure versus constant flow rate gradient elution separations. Part I: theory.

    Science.gov (United States)

    Broeckhoven, K; Verstraeten, M; Choikhet, K; Dittmann, M; Witt, K; Desmet, G

    2011-02-25

    We report on a general theoretical assessment of the potential kinetic advantages of running LC gradient elution separations in the constant-pressure mode instead of in the customarily used constant-flow rate mode. Analytical calculations as well as numerical simulation results are presented. It is shown that, provided both modes are run with the same volume-based gradient program, the constant-pressure mode can potentially offer an identical separation selectivity (except from some small differences induced by the difference in pressure and viscous heating trajectory), but in a significantly shorter time. For a gradient running between 5 and 95% of organic modifier, the decrease in analysis time can be expected to be of the order of some 20% for both water-methanol and water-acetonitrile gradients, and only weakly depending on the value of V(G)/V₀ (or equivalently t(G)/t₀). Obviously, the gain will be smaller when the start and end composition lie closer to the viscosity maximum of the considered water-organic modifier system. The assumptions underlying the obtained results (no effects of pressure and temperature on the viscosity or retention coefficient) are critically reviewed, and can be inferred to only have a small effect on the general conclusions. It is also shown that, under the adopted assumptions, the kinetic plot theory also holds for operations where the flow rate varies with the time, as is the case for constant-pressure operation. Comparing both operation modes in a kinetic plot representing the maximal peak capacity versus time, it is theoretically predicted here that both modes can be expected to perform equally well in the fully C-term dominated regime (where H varies linearly with the flow rate), while the constant pressure mode is advantageous for all lower flow rates. Near the optimal flow rate, and for linear gradients running from 5 to 95% organic modifier, time gains of the order of some 20% can be expected (or 25-30% when accounting for

  13. Implicit unified gas-kinetic scheme for steady state solutions in all flow regimes

    Science.gov (United States)

    Zhu, Yajun; Zhong, Chengwen; Xu, Kun

    2016-06-01

    This paper presents an implicit unified gas-kinetic scheme (UGKS) for non-equilibrium steady state flow computation. The UGKS is a direct modeling method for flow simulation in all regimes with the updates of both macroscopic flow variables and microscopic gas distribution function. By solving the macroscopic equations implicitly, a predicted equilibrium state can be obtained first through iterations. With the newly predicted equilibrium state, the evolution equation of the gas distribution function and the corresponding collision term can be discretized in a fully implicit way for fast convergence through iterations as well. The lower-upper symmetric Gauss-Seidel (LU-SGS) factorization method is implemented to solve both macroscopic and microscopic equations, which improves the efficiency of the scheme. Since the UGKS is a direct modeling method and its physical solution depends on the mesh resolution and the local time step, a physical time step needs to be fixed before using an implicit iterative technique with a pseudo-time marching step. Therefore, the physical time step in the current implicit scheme is determined by the same way as that in the explicit UGKS for capturing the physical solution in all flow regimes, but the convergence to a steady state speeds up through the adoption of a numerical time step with large CFL number. Many numerical test cases in different flow regimes from low speed to hypersonic ones, such as the Couette flow, cavity flow, and the flow passing over a cylinder, are computed to validate the current implicit method. The overall efficiency of the implicit UGKS can be improved by one or two orders of magnitude in comparison with the explicit one.

  14. Dynamic evolution of a flow to localized, kinetics-driven ablation or coagulation

    Science.gov (United States)

    Hagan, Daniel; Crocker, Ryan; Dubief, Yves

    2012-11-01

    This research focuses on the numerical simulation of the ablative creation of a cavity or a coagulative formation at a wall in a flow. The fluid-solid interface is defined by a level set (LS) variable, whose transport equation is driven by the mass-loss or growth process. The boundary conditions at the fluid-solid interface are enforced by a mass and energy-conserving immersed boundary method (IBM) using the ghost-fluid node approach for the latter and for the transport of chemical species. The first application of the LS/IBM algorithm is a channel flow in which both walls are cavity-free, but one wall contains a section made of ablatable material, which could correspond to a hole or gap in a spacecraft thermal protection shield. The second application is a pipe flow in which the wall is capable of accumulating material, which could describe the coagulation of blood at a vessel wall. The solid mass loss or growth is driven by one step kinetics. For both flows, the dynamical interplay between the ablative or coagulative patch is investigated through statistics and flow topology. We gratefully acknowledge the financial support of NASA, grant No. NNX11AM07A, and NIH, grant No. P01HL46703, and the computational support of the Vermont Advanced Computing Core.

  15. NONINVASIVE MEASUREMENT OF INTRARENAL BLOOD-FLOW DISTRIBUTION - KINETIC-MODEL OF RENAL I-123 HIPPURAN HANDLING

    NARCIS (Netherlands)

    JANSSEN, WMT; BEEKHUIS, H; DEBRUIN, R; DEJONG, PE; DEZEEUW, D

    1995-01-01

    A new technique for noninvasive measurement of intrarenal blood flow distribution over cortex and medulla is proposed. The tech nique involves analysis of I-123-labeled hippuran renography, according to a kinetic model that describes the flow of I-123- hippuran from the heart (input) through the ren

  16. Discrete unified gas kinetic scheme for all Knudsen number flows: low-speed isothermal case.

    Science.gov (United States)

    Guo, Zhaoli; Xu, Kun; Wang, Ruijie

    2013-09-01

    Based on the Boltzmann-BGK (Bhatnagar-Gross-Krook) equation, in this paper a discrete unified gas kinetic scheme (DUGKS) is developed for low-speed isothermal flows. The DUGKS is a finite-volume scheme with the discretization of particle velocity space. After the introduction of two auxiliary distribution functions with the inclusion of collision effect, the DUGKS becomes a fully explicit scheme for the update of distribution function. Furthermore, the scheme is an asymptotic preserving method, where the time step is only determined by the Courant-Friedricks-Lewy condition in the continuum limit. Numerical results demonstrate that accurate solutions in both continuum and rarefied flow regimes can be obtained from the current DUGKS. The comparison between the DUGKS and the well-defined lattice Boltzmann equation method (D2Q9) is presented as well.

  17. A cutoff phenomenon in accelerated stochastic simulations of chemical kinetics via flow averaging (FLAVOR-SSA)

    Science.gov (United States)

    Bayati, Basil; Owhadi, Houman; Koumoutsakos, Petros

    2010-12-01

    We present a simple algorithm for the simulation of stiff, discrete-space, continuous-time Markov processes. The algorithm is based on the concept of flow averaging for the integration of stiff ordinary and stochastic differential equations and ultimately leads to a straightforward variation of the the well-known stochastic simulation algorithm (SSA). The speedup that can be achieved by the present algorithm [flow averaging integrator SSA (FLAVOR-SSA)] over the classical SSA comes naturally at the expense of its accuracy. The error of the proposed method exhibits a cutoff phenomenon as a function of its speed-up, allowing for optimal tuning. Two numerical examples from chemical kinetics are provided to illustrate the efficiency of the method.

  18. Kinetic theory for flows of nonhomogeneous rodlike liquid crystalline polymers with a nonlocal intermolecular potential.

    Science.gov (United States)

    Wang, Qi; E, Weinan; Liu, Chun; Zhang, Pingwen

    2002-05-01

    The Doi kinetic theory for flows of homogeneous, rodlike liquid crystalline polymers (LCPs) is extended to model flows of nonhomogeneous, rodlike LCPs through a nonlocal (long-range) intermolecular potential. The theory features (i) a nonlocal, anisotropic, effective intermolecular potential in an integral form that is consistent with the chemical potential, (ii) short-range elasticity as well as long-range isotropic and anisotropic elasticity, (iii) a closed-form stress expression accounting for the nonlocal molecular interaction, and (iv) an extra elastic body force exclusively associated with the integral form of the intermolecular potential. With the effective intermolecular potential, the theory is proven to be well posed in that it warrants a positive entropy production and thereby the second law of thermodynamics. Approximate theories are obtained by gradient expansions of the number density function in the free energy density.

  19. Cantorian Fractal Space-Time Fluctuations in Turbulent Fluid Flows and the Kinetic Theory of Gases

    CERN Document Server

    Selvam, A M

    1999-01-01

    Fluid flows such as gases or liquids exhibit space-time fluctuations on all scales extending down to molecular scales. Such broadband continuum fluctuations characterise all dynamical systems in nature and are identified as selfsimilar fractals in the newly emerging multidisciplinary science of nonlinear dynamics and chaos. A cell dynamical system model has been developed by the author to quantify the fractal space-time fluctuations of atmospheric flows. The earth's atmosphere consists of a mixture of gases and obeys the gas laws as formulated in the kinetic theory of gases developed on probabilistic assumptions in 1859 by the physicist James Clerk Maxwell. An alternative theory using the concept of fractals and chaos is applied in this paper to derive these fundamental gas laws.

  20. Accelerating moderately stiff chemical kinetics in reactive-flow simulations using GPUs

    CERN Document Server

    Niemeyer, Kyle E

    2014-01-01

    The chemical kinetics ODEs arising from operator-split reactive-flow simulations were solved on GPUs using explicit integration algorithms. Nonstiff chemical kinetics of a hydrogen oxidation mechanism (9 species and 38 irreversible reactions) were computed using the explicit fifth-order Runge-Kutta-Cash-Karp method, and the GPU-accelerated version performed faster than single- and six-core CPU versions by factors of 126 and 25, respectively, for 524,288 ODEs. Moderately stiff kinetics, represented with mechanisms for hydrogen/carbon-monoxide (13 species and 54 irreversible reactions) and methane (53 species and 634 irreversible reactions) oxidation, were computed using the stabilized explicit second-order Runge-Kutta-Chebyshev (RKC) algorithm. The GPU-based RKC implementation demonstrated an increase in performance of nearly 59 and 10 times, for problem sizes consisting of 262,144 ODEs and larger, than the single- and six-core CPU-based RKC algorithms using the hydrogen/carbon-monoxide mechanism. With the met...

  1. An upwind, kinetic flux-vector splitting method for flows in chemical and thermal non-equilibrium

    Science.gov (United States)

    Eppard, W. M.; Grossman, B.

    1993-01-01

    We have developed new upwind kinetic difference schemes for flows with non-equilibrium thermodynamics and chemistry. These schemes are derived from the Boltzmann equation with the resulting Euler schemes developed as moments of the discretized Boltzmann scheme with a locally Maxwellian velocity distribution. Splitting the velocity distribution at the Boltzmann level is seen to result in a flux-split Euler scheme and is called Kinetic Flux Vector Splitting (KFVS). Extensions to flows with finite-rate chemistry and vibrational relaxation is accomplished utilizing nonequilibrium kinetic theory. Computational examples are presented comparing KFVS with the schemes of Van Leer and Roe for a quasi-one-dimensional flow through a supersonic diffuser, inviscid flow through two-dimensional inlet, and viscous flow over a cone at zero angle-of-attack. Calculations are also shown for the transonic flow over a bump in a channel and the transonic flow over an NACA 0012 airfoil. The results show that even though the KFVS scheme is a Riemann solver at the kinetic level, its behavior at the Euler level is more similar to the existing flux-vector splitting algorithms than to the flux-difference splitting scheme of Roe.

  2. An upwind, kinetic flux-vector splitting method for flows in chemical and thermal non-equilibrium

    Science.gov (United States)

    Eppard, W. M.; Grossman, B.

    1993-01-01

    We have developed new upwind kinetic difference schemes for flows with non-equilibrium thermodynamics and chemistry. These schemes are derived from the Boltzmann equation with the resulting Euler schemes developed as moments of the discretized Boltzmann scheme with a locally Maxwellian velocity distribution. Splitting the velocity distribution at the Boltzmann level is seen to result in a flux-split Euler scheme and is called Kinetic Flux Vector Splitting (KFVS). Extensions to flows with finite-rate chemistry and vibrational relaxation is accomplished utilizing nonequilibrium kinetic theory. Computational examples are presented comparing KFVS with the schemes of Van Leer and Roe for a quasi-one-dimensional flow through a supersonic diffuser, inviscid flow through two-dimensional inlet, and viscous flow over a cone at zero angle-of-attack. Calculations are also shown for the transonic flow over a bump in a channel and the transonic flow over an NACA 0012 airfoil. The results show that even though the KFVS scheme is a Riemann solver at the kinetic level, its behavior at the Euler level is more similar to the existing flux-vector splitting algorithms than to the flux-difference splitting scheme of Roe.

  3. A semi-implicit gas-kinetic scheme for smooth flows

    Science.gov (United States)

    Wang, Peng; Guo, Zhaoli

    2016-08-01

    In this paper, a semi-implicit gas-kinetic scheme (SIGKS) is derived for smooth flows based on the Bhatnagar-Gross-Krook (BGK) equation. As a finite-volume scheme, the evolution of the average flow variables in a control volume is under the Eulerian framework, whereas the construction of the numerical flux across the cell interface comes from the Lagrangian perspective. The adoption of the Lagrangian aspect makes the collision and the transport mechanisms intrinsically coupled together in the flux evaluation. As a result, the time step size is independent of the particle collision time and solely determined by the Courant-Friedrichs-Lewy (CFL) condition. An analysis of the reconstructed distribution function at the cell interface shows that the SIGKS can be viewed as a modified Lax-Wendroff type scheme with an additional term. Furthermore, the addition term coming from the implicitness in the reconstruction is expected to be able to enhance the numerical stability of the scheme. A number of numerical tests of smooth flows with low and moderate Mach numbers are performed to benchmark the SIGKS. The results show that the method has second-order spatial accuracy, and can give accurate numerical solutions in comparison with benchmark results. It is also demonstrated that the numerical stability of the proposed scheme is better than the original GKS for smooth flows.

  4. A kinetic model for corrosion and precipitation in non-isothermal LBE flow loop

    Science.gov (United States)

    He, By Xiaoyi; Li, Ning; Mineev, Mark

    2001-08-01

    A kinetic model was developed to estimate the corrosion/precipitation rate in a non-isothermal liquid lead-bismuth eutectic (LBE) flow loop. The model was based on solving the mass transport equation with the assumptions that convective transport dominates in the longitudinal flow direction and diffusion dominates in the transverse direction. The species concentration at wall is assumed to be determined either by the solubility of species in LBE in the absence of oxygen or by the reduction reaction of the protective oxide film when active oxygen control is applied. Analyses show that the corrosion/precipitation rate depends on the flow velocity, the species diffusion rate, the oxygen concentration in LBE, as well as the temperature distribution along a loop. Active oxygen control can significantly reduce the corrosion/precipitation of the structural materials. It is shown that the highest corrosion/precipitation does not necessarily locate at places with the highest/lowest temperature. For a material testing loop being constructed at the Los Alamos National Laboratory (LANL), the highest corrosion occurs at the end of the heater zone, while the highest precipitation occurs in the return flow in the recuperator.

  5. In vitro evaluation of flow patterns and turbulent kinetic energy in trans-catheter aortic valve prostheses.

    Science.gov (United States)

    Giese, Daniel; Weiss, Kilian; Baeßler, Bettina; Madershahian, Navid; Choi, Yeong-Hoon; Maintz, David; Bunck, Alexander C

    2017-09-18

    The objective of the current work was to evaluate flow and turbulent kinetic energy in different transcatheter aortic valve implants using highly undersampled time-resolved multi-point 3-directional phase-contrast measurements (4D Flow MRI) in an in vitro setup. A pulsatile flow setup was used with a compliant tubing mimicking a stiff left ventricular outflow tract and ascending aorta. Five different implants were measured using a highly undersampled multi-point 4D Flow MRI sequence. Velocities and turbulent kinetic energy values were analysed and compared. Strong variations of turbulent kinetic energy distributions between the valves were observed. Maximum turbulent kinetic energy values ranged from 100 to over 500 J/m(3) while through-plane velocities were similar between all valves. Highly accelerated 4D Flow MRI for the measurement of velocities and turbulent kinetic energy values allowed for the assessment of hemodynamic parameters in five different implant models. The presented setup, measurement protocol and analysis methods provides an efficient approach to compare different valve implants and could aid future novel valve designs.

  6. Analysis of atmospheric flow over a surface protrusion using the turbulence kinetic energy equation with reference to aeronautical operating systems

    Science.gov (United States)

    Frost, W.; Harper, W. L.

    1975-01-01

    Flow over surface obstructions can produce significantly large wind shears such that adverse flying conditions can occur for aeronautical systems (helicopters, STOL vehicles, etc.). Atmospheric flow fields resulting from a semi-elliptical surface obstruction in an otherwise horizontally homogeneous statistically stationary flow are modelled with the boundary-layer/Boussinesq-approximation of the governing equation of fluid mechanics. The turbulence kinetic energy equation is used to determine the dissipative effects of turbulent shear on the mean flow. Iso-lines of turbulence kinetic energy and turbulence intensity are plotted in the plane of the flow and highlight regions of high turbulence intensity in the stagnation zone and sharp gradients in intensity along the transition from adverse to favourable pressure gradient. Discussion of the effects of the disturbed wind field in CTOL and STOL aircraft flight path and obstruction clearance standards is given. The results indicate that closer inspection of these presently recommended standards as influenced by wind over irregular terrains is required.

  7. A Triple-Stain Flow Cytometric Method to Assess Plasma- and Acrosome-Membrane Integrity of Cryopreserved Bovine Sperm Immediately after Thawing in Presence of Egg-Yolk Particles

    NARCIS (Netherlands)

    Nagy, S.; Jansen, J.; Topper, E.K.; Gadella, B.M.

    2003-01-01

    Simultaneously evaluating postthaw viability and acrosome integrity of spermatozoa by flow cytometry would provide a valuable testing tool in both research and routine work. In the present study, a new triple-stain combination was developed for the simultaneous evaluation of viability and acrosome i

  8. A semi-Lagrangian gas-kinetic scheme for smooth flows

    CERN Document Server

    Wang, Peng

    2014-01-01

    In this paper, a semi-Lagrangian gas-kinetic scheme is developed for smooth flows based on the Bhatnagar-Gross-Krook (BGK) equation. As a finite-volume scheme, the evolution of the average flow variables in a control volume is under the Eulerian framework, whereas the construction of the numerical flux across the cell interface comes from the Lagrangian perspective. The adoption of the Lagrangian aspect makes the collision and the transport mechanisms intrinsically coupled together in the flux evaluation. As a result, the time step is independent of the particle collision time and solely determined by the Courant-Friedrichs-Lewy (CFL) conditions. A set of simulations are carried out to validate the performance of the new scheme. The results show that with second-order spatial accuracy, the scheme exhibits low numerical dissipation, and can accurately capture the Navier-Stokers solutions for the smooth flows with viscous heat dissipation from the low-speed incompressible to hypersonic compressible regimes.

  9. Electro-Hydrodynamics and Kinetic Modeling of Dry and Humid Air Flows Activated by Corona Discharges

    Science.gov (United States)

    P. Sarrette, J.; Eichwald, O.; Marchal, F.; Ducasse, O.; Yousfi, M.

    2016-05-01

    The present work is devoted to the 2D simulation of a point-to-plane Atmospheric Corona Discharge Reactor (ACDR) powered by a DC high voltage supply. The corona reactor is periodically crossed by thin mono filamentary streamers with a natural repetition frequency of some tens of kHz. The study compares the results obtained in dry air and in air mixed with a small amount of water vapour (humid air). The simulation involves the electro-dynamics, chemical kinetics and neutral gas hydrodynamics phenomena that influence the kinetics of the chemical species transformation. Each discharge lasts about one hundred of a nanosecond while the post-discharge occurring between two successive discharges lasts one hundred of a microsecond. The ACDR is crossed by a lateral dry or humid air flow initially polluted with 400 ppm of NO. After 5 ms, the time corresponding to the occurrence of 50 successive discharge/post-discharge phases, a higher NO removal rate and a lower ozone production rate are found in humid air. This change is due to the presence of the HO2 species formed from the H primary radical in the discharge zone.

  10. Electro-Hydrodynamics and Kinetic Modeling of Dry and Humid Air Flows Activated by Corona Discharges

    Institute of Scientific and Technical Information of China (English)

    J.P.SARRETTE; O.EICHWALD; F.MARCHAL; O.DUCASSE; M.YOUSFI

    2016-01-01

    The present work is devoted to the 2D simulation of a point-to-plane Atmospheric Corona Discharge Reactor (ACDR) powered by a DC high voltage supply.The corona reactor is periodically crossed by thin mono filamentary streamers with a natural repetition frequency of some tens of kHz.The study compares the results obtained in dry air and in air mixed with a small amount of water vapour (humid air).The simulation involves the electro-dynamics,chemical kinetics and neutral gas hydrodynamics phenomena that influence the kinetics of the chemical species transformation.Each discharge lasts about one hundred of a nanosecond while the post-discharge occurring between two successive discharges lasts one hundred of a microsecond.The ACDR is crossed by a lateral dry or humid air flow initially polluted with 400 ppm of NO.After 5 ms,the time corresponding to the occurrence of 50 successive discharge/post-discharge phases,a higher NO removal rate and a lower ozone production rate are found in humid air.This change is due to the presence of the HO2 species formed from the H primary radical in the discharge zone.

  11. Kinetic freeze-out, particle spectra and harmonic flow coefficients from mode-by-mode hydrodynamics

    CERN Document Server

    Floerchinger, Stefan

    2014-01-01

    The kinetic freeze-out for the hydrodynamical description of relativistic heavy ion collisions is discussed using a background-fluctuation splitting of the hydrodynamical fields. For a single event, the particle spectrum, or its logarithm, can be written as the sum of background part that is symmetric with respect to azimuthal rotations and longitudinal boosts and a part containing the contribution of fluctuations or deviations from the background. Using a complete orthonormal basis to characterize the initial state allows one to write the double differential harmonic flow coefficients determined by the two-particle correlation method as matrix expressions involving the initial fluid correlations. We discuss the use of these expressions for a mode-by-mode analysis of fluctuating initial conditions in heavy ion collisions.

  12. A plant kinetic study of alcoholic fermentation using reversed-flow gas chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Economopoulos, N.; Athanassopoulos, N. (B.G. Spiliopoulos Distilleri S.A., Patras (Greece)); Katsanos, N.A.; Karaiskakis, G.; Agathonos, P.; Vassilakos, Ch. (Univ. of Patras (Greece))

    1992-12-01

    The reversed-flow gas chromatographic sampling technique is used to study the kinetics of alcoholic fermentation in a factory in conjunction with measurements of suspended particles in the fermenting medium. It was found that the overall process consists of four phases which have different first-order rate constants during ethanol formation. The second phase is the slowest with its rate constant being 4.3 and 13 times smaller than that of the first and third phases, respectively. There is also a decrease of suspended particles during the second phase. These results show that there is the possibility of increasing the rate constant during this phase, thereby increasing the overall production rate of ethanol significantly and thus lowering its cost of production.

  13. Influence of phosphorus on Cu sorption kinetics: Stirred flow chamber experiments

    Energy Technology Data Exchange (ETDEWEB)

    Perez-Novo, C. [Area de Edafoloxia e Quimica Agricola, Departamento de Bioloxia Vexetal e Ciencia do Solo, Universidade de Vigo, Facultade de Ciencias, 32004 Ourense (Spain); Fernandez-Calvino, D., E-mail: davidfc@uvigo.es [Area de Edafoloxia e Quimica Agricola, Departamento de Bioloxia Vexetal e Ciencia do Solo, Universidade de Vigo, Facultade de Ciencias, 32004 Ourense (Spain); Bermudez-Couso, A.; Lopez-Periago, J.E.; Arias-Estevez, M. [Area de Edafoloxia e Quimica Agricola, Departamento de Bioloxia Vexetal e Ciencia do Solo, Universidade de Vigo, Facultade de Ciencias, 32004 Ourense (Spain)

    2011-01-15

    A stirred flow reactor was used to study the influence of phosphorus on the adsorption and desorption kinetics of copper in two acid soils on granite and amphibolite. The presence of P was found to significantly increase Cu adsorption in both soils, albeit at different types of sites (mainly in slow adsorption sites in the soil on granite, and both in fast and slow adsorption sites in that on amphibolite). The increased Cu sorption at fast sites in the amphibolite soil was due to its high content in Fe oxyhydroxides, which bound P and released OH{sup -} as a result, thereby raising the pH and leading to a higher sorption capacity during fast reactions. On the other hand, the increased Cu sorption at slow adsorption sites was due to Cu{sup 2+} acting as a bridging element between P and organic matter.

  14. Multicentric study underlining the interest of adding CD5, CD7 and CD56 expression assessment to the flow cytometric Ogata score in myelodysplastic syndromes and myelodysplastic/myeloproliferative neoplasms.

    Science.gov (United States)

    Bardet, Valérie; Wagner-Ballon, Orianne; Guy, Julien; Morvan, Céline; Debord, Camille; Trimoreau, Franck; Benayoun, Emmanuel; Chapuis, Nicolas; Freynet, Nicolas; Rossi, Cédric; Mathis, Stéphanie; Gourin, Marie-Pierre; Toma, Andréa; Béné, Marie C; Feuillard, Jean; Guérin, Estelle

    2015-04-01

    Although numerous recent publications have demonstrated interest in multiparameter flow cytometry in the investigation of myelodysplastic disorders, it is perceived by many laboratory hematologists as difficult and expensive, requiring a high level of expertise. We report a multicentric open real-life study aimed at evaluating the added value of the technically simple flow cytometry score described by the Ogata group for the diagnosis of myelodysplastic syndromes. A total of 652 patients were recruited prospectively in four different centers: 346 myelodysplastic syndromes, 53 myelodysplastic/myeloproliferative neoplasms, and 253 controls. The Ogata score was assessed using CD45 and CD34 staining, with the addition of CD10 and CD19. Moreover, labeling of CD5, CD7 and CD56 for the evaluation of myeloid progenitors and monocytes was tested on a subset of 294 patients. On the whole series, the specificity of Ogata score reached 89%. Respective sensitivities were 54% for low-risk myelodysplastic syndromes, 68% and 84% for type 1 and type 2 refractory anemia with excess of blasts, and 72% for myelodysplastic/myeloproliferative neoplasms. CD5 expression was poorly informative. When adding CD56 or CD7 labeling to the Ogata score, sensitivity rose to 66% for low-risk myelodysplastic syndromes, to 89% for myelodysplastic/myeloproliferative neoplasms and to 97% for refractory anemia with excess of blasts. This large multicenter study confirms the feasibility of Ogata scoring in routine flow cytometry diagnosis but highlights its poor sensitivity in low-risk myelodysplastic syndromes. The addition of CD7 and CD56 in flow cytometry panels improves the sensitivity but more sophisticated panels would be more informative. Copyright© Ferrata Storti Foundation.

  15. Flow cytometric detection of micronuclei and cell cycle alterations in fish-derived cells after exposure to three model genotoxic agents: mitomycin C, vincristine sulfate and benzo(a)pyrene.

    Science.gov (United States)

    Sánchez, P; Llorente, M T; Castaño, A

    2000-02-16

    The measurement of cytogenetic alterations in vitro is considered an initial step in the risk assessment procedures for genotoxic agents. The concern about genotoxic pollutants in natural fish population makes the use of fish-derived cells an useful tool for these purposes. The technological improvements in well-established cytogenetic endpoints, such as micronuclei (MN) estimations by means of flow cytometry, have been proposed in the later years using mammalian cells. In this work, we test the capability of flow cytometry to evaluate MN induction and cell cycle alterations in an established fish cell line (RTG-2) using three agent-inductor models at different concentrations and exposure periods. For mitomycin C, an inverse relationship between length of exposure period and concentrations was observed. A dose-response relationship was observed after exposing RTG-2 cells to vincristine sulfate and benzo(a)pyrene. As this study shows, RTG-2 cells respond to clastogenic and aneugenic effects of the tested chemicals through the induction of MN at similar doses to mammalian cells and without the addition of exogenous metabolic activity. The possibility to check cell cycle alterations, in the same sample, gives the opportunity to evaluate early signals of cytotoxicity. The use of flow cytometry improves the assay by means of its speed and objectivity, which makes the assay very useful for genotoxicity assessment of aquatic chemicals.

  16. Kinetics of Fe(II)-catalyzed transformation of 6-line ferrihydrite under anaerobic flow conditions

    Energy Technology Data Exchange (ETDEWEB)

    Yang, L.; Steefel, C.I.; Marcus, M.A.; Bargar, J.R.

    2010-04-01

    The readsorption of ferrous ions produced by the abiotic and microbially-mediated reductive dissolution of iron oxy-hydroxides drives a series of transformations of the host minerals. To further understand the mechanisms by which these transformations occur and their kinetics within a microporous flow environment, flow-through experiments were conducted in which capillary tubes packed with ferrihydrite-coated glass spheres were injected with inorganic Fe(II) solutions under circumneutral pH conditions at 25 C. Synchrotron X-ray diffraction was used to identify the secondary phase(s) formed and to provide data for quantitative kinetic analysis. At concentrations at and above 1.8 mM Fe(II) in the injection solution, magnetite was the only secondary phase formed (no intermediates were detected), with complete transformation following a nonlinear rate law requiring 28 hours and 150 hours of reaction at 18 and 1.8 mM Fe(II), respectively. However, when the injection solution consisted of 0.36 mM Fe(II), goethite was the predominant reaction product and formed much more slowly according to a linear rate law, while only minor magnetite was formed. When the rates are normalized based on the time to react half of the ferrihydrite on a reduced time plot, it is apparent that the 1.8 mM and 18 mM input Fe(II) experiments can be described by the same reaction mechanism, while the 0.36 input Fe(II) experiment is distinct. The analysis of the transformation kinetics suggest that the transformations involved an electron transfer reaction between the aqueous as well as sorbed Fe(II) and ferrihydrite acting as a semiconductor, rather than a simple dissolution and recrystallization mechanism. A transformation mechanism involving sorbed inner sphere Fe(II) alone is not supported, since the essentially equal coverage of sorption sites in the 18 mM and 1.8 mM Fe(II) injections cannot explain the difference in the transformation rates observed.

  17. Kinetics of Fe(II)-catalyzed transformation of 6-line ferrihydrite under anaerobic flow conditions

    Energy Technology Data Exchange (ETDEWEB)

    Yang, L.; Steefel, C.I.; Marcus, M.A.; Bargar, J.R.

    2010-04-01

    The readsorption of ferrous ions produced by the abiotic and microbially-mediated reductive dissolution of iron oxy-hydroxides drives a series of transformations of the host minerals. To further understand the mechanisms by which these transformations occur and their kinetics within a microporous flow environment, flow-through experiments were conducted in which capillary tubes packed with ferrihydrite-coated glass spheres were injected with inorganic Fe(II) solutions under circumneutral pH conditions at 25 C. Synchrotron X-ray diffraction was used to identify the secondary phase(s) formed and to provide data for quantitative kinetic analysis. At concentrations at and above 1.8 mM Fe(II) in the injection solution, magnetite was the only secondary phase formed (no intermediates were detected), with complete transformation following a nonlinear rate law requiring 28 hours and 150 hours of reaction at 18 and 1.8 mM Fe(II), respectively. However, when the injection solution consisted of 0.36 mM Fe(II), goethite was the predominant reaction product and formed much more slowly according to a linear rate law, while only minor magnetite was formed. When the rates are normalized based on the time to react half of the ferrihydrite on a reduced time plot, it is apparent that the 1.8 mM and 18 mM input Fe(II) experiments can be described by the same reaction mechanism, while the 0.36 input Fe(II) experiment is distinct. The analysis of the transformation kinetics suggest that the transformations involved an electron transfer reaction between the aqueous as well as sorbed Fe(II) and ferrihydrite acting as a semiconductor, rather than a simple dissolution and recrystallization mechanism. A transformation mechanism involving sorbed inner sphere Fe(II) alone is not supported, since the essentially equal coverage of sorption sites in the 18 mM and 1.8 mM Fe(II) injections cannot explain the difference in the transformation rates observed.

  18. Optimization of a flow cytometric method for the simultaneous measurement of cell surface antigen, DNA content, and in vitro BrdUrd incorporation into normal and malignant hematopoietic cells.

    Science.gov (United States)

    Holm, M; Thomsen, M; Høyer, M; Hokland, P

    1998-05-01

    We have designed an assay for the simultaneous measurement of cell surface phenotype, S-phase fraction, and DNA content by single laser instrumentation for the purpose of determining the labeling index (LI), duration of S-phase (Ts), and the potential doubling time (Tpot) of leukocyte subpopulations. The procedure was optimized with regard to: mode of bromodeoxyuridine (BrdUrd) incorporation, selection of suitable leukocyte differentiation antigens (LDAs) as well as PE-conjugated monoclonal antibodies (MoAbs) against myeloid cells, overnight permeabilization and fixation (paraformaldehyde 1% and 0.05% Nonidet P40), DNase I treatment (250 Kunitz units), concentration of FITC-conjugated anti-BrdUrd MoAb (dilution 1:5), and DNA staining with 7-amino-actinomycin (7-AAD) (10 microg/ml). We validated this assay by measuring LI, Ts, and Tpot repeatedly in four leukemic cell lines and found these to be stable (coefficients of variation (CV): 0.06, 0.13, and 0.08, respectively). Finally, we employed the assay on different leukocyte preparations from normal donors (including purified CD34 + cells) and patients with malignant myeloid disorders, and we concluded that it will yield valuable data regarding the cell cycle kinetics of subsets of leukocytes in heterogeneous mixtures of hematopoietic cells.

  19. Analysis of fluid fuel flow to the neutron kinetics on molten salt reactor FUJI-12

    Energy Technology Data Exchange (ETDEWEB)

    Aji, Indarta Kuncoro, E-mail: indartaaji@s.itb.ac.id [Department of Physics, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesa 10 Bandung 40132 (Indonesia); Waris, Abdul, E-mail: awaris@fi.itb.ac.id; Permana, Sidik [Nuclear Physics & Biophysics Research Division, Department of Physics, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesa 10 Bandung 40132 (Indonesia)

    2015-09-30

    Molten Salt Reactor is a reactor are operating with molten salt fuel flowing. This condition interpret that the neutron kinetics of this reactor is affected by the flow rate of the fuel. This research analyze effect by the alteration velocity of the fuel by MSR type Fuji-12, with fuel composition LiF-BeF{sub 2}-ThF{sub 4}-{sup 233}UF{sub 4} respectively 71.78%-16%-11.86%-0.36%. Calculation process in this study is performed numerically by SOR and finite difference method use C programming language. Data of reactivity, neutron flux, and the macroscopic fission cross section for calculation process obtain from SRAC-CITATION (Standard thermal Reactor Analysis Code) and JENDL-4.0 data library. SRAC system designed and developed by JAEA (Japan Atomic Energy Agency). This study aims to observe the effect of the velocity of fuel salt to the power generated from neutron precursors at fourth year of reactor operate (last critical condition) with number of multiplication effective; 1.0155.

  20. Analysis of fluid fuel flow to the neutron kinetics on molten salt reactor FUJI-12

    Science.gov (United States)

    Aji, Indarta Kuncoro; Waris, Abdul; Permana, Sidik

    2015-09-01

    Molten Salt Reactor is a reactor are operating with molten salt fuel flowing. This condition interpret that the neutron kinetics of this reactor is affected by the flow rate of the fuel. This research analyze effect by the alteration velocity of the fuel by MSR type Fuji-12, with fuel composition LiF-BeF2-ThF4-233UF4 respectively 71.78%-16%-11.86%-0.36%. Calculation process in this study is performed numerically by SOR and finite difference method use C programming language. Data of reactivity, neutron flux, and the macroscopic fission cross section for calculation process obtain from SRAC-CITATION (Standard thermal Reactor Analysis Code) and JENDL-4.0 data library. SRAC system designed and developed by JAEA (Japan Atomic Energy Agency). This study aims to observe the effect of the velocity of fuel salt to the power generated from neutron precursors at fourth year of reactor operate (last critical condition) with number of multiplication effective; 1.0155.

  1. Studying depletion kinetics of circulating prostate cancer cells by in vivo flow cytometer

    Science.gov (United States)

    Liu, Guangda; Gu, Zhengqin; Guo, Jin; Li, Yan; Chen, Yun; Chen, Tong; Wang, Cheng; Wei, Xunbin

    2011-03-01

    Prostate cancer is the most common malignancy in American men and the second leading cause of deaths from cancer, after lung cancer. The tumor usually grows slowly and remains confined to the gland for many years. During this time, the tumor produces little or no symptoms or outward signs. As the cancer advances, however, it can metastasize throughout other areas of the body, such as the bones, lungs, and liver. Surgical resection, hormonal therapy, chemotherapy and radiation therapy are the foundation of current prostate cancer therapies. Treatments for prostate cause both short- and long-term side effects that may be difficult to accept. Molecular mechanisms of prostate cancer metastasis need to be understood better and new therapies must be developed to selectively target to unique characteristics of cancer cell growth and metastasis. We have developed the "in vivo microscopy" to study the mechanisms that govern prostate cancer cell spread through the microenvironment in vivo in real-time confocal near-infrared fluorescence imaging. A recently developed "in vivo flow cytometer" and optical imaging are used to assess prostate cancer cell spreading and the circulation kinetics of prostate cancer cells. A real- time quantitative monitoring of circulating prostate cancer cells by the in vivo flow cytometer will be useful to assess the effectiveness of the potential therapeutic interventions.

  2. Extracting kinetic freeze-out temperature and radial flow velocity from an improved Tsallis distribution

    CERN Document Server

    Lao, Hai-Ling; Lacey, Roy A

    2016-01-01

    We analyze the transverse momentum ($p_T$) spectra of identified particles ($\\pi^{\\pm}$, $K^{\\pm}$, $p$, and $\\bar p$) produced in gold-gold (Au-Au) and lead-lead (Pb-Pb) collisions over a $\\sqrt{s_{NN}}$ (center-of-mass energy per nucleon pair) range from 14.5 GeV [one of the Relativistic Heavy Ion Collider (RHIC) energies] to 2.76 TeV [one of the Large Hadron Collider (LHC) energies]. For the spectra with a narrow $p_T$ range, an improved Tsallis distribution which is in fact the Tsallis distribution with radial flow is used. For the spectra with a wide $p_T$ range, a superposition of the improved Tsallis distribution and an inverse power-law is used. Both the extracted kinetic freeze-out temperature ($T_0$) and radial flow velocity ($\\beta_T$) increase with the increase of $\\sqrt{s_{NN}}$, which indicate a higher excitation and larger expansion of the interesting system at the LHC. Both the values of $T_0$ and $\\beta_T$ in central collisions are slightly larger than those in peripheral collisions, and they...

  3. Biodrying of sewage sludge: kinetics of volatile solids degradation under different initial moisture contents and air-flow rates.

    Science.gov (United States)

    Villegas, Manuel; Huiliñir, Cesar

    2014-12-01

    This study focuses on the kinetics of the biodegradation of volatile solids (VS) of sewage sludge for biodrying under different initial moisture contents (Mc) and air-flow rates (AFR). For the study, a 3(2) factorial design, whose factors were AFR (1, 2 or 3L/minkgTS) and initial Mc (59%, 68% and 78% w.b.), was used. Using seven kinetic models and a nonlinear regression method, kinetic parameters were estimated and the models were analyzed with two statistical indicators. Initial Mc of around 68% increases the temperature matrix and VS consumption, with higher moisture removal at lower initial Mc values. Lower AFRs gave higher matrix temperatures and VS consumption, while higher AFRs increased water removal. The kinetic models proposed successfully simulate VS biodegradation, with root mean square error (RMSE) between 0.007929 and 0.02744, and they can be used as a tool for satisfactory prediction of VS in biodrying.

  4. Reduction Kinetics of Manganese Dioxide by Geobacter Sulfurreducens and Associated Biofilm Morphology in a Flow-Through Reactor

    Science.gov (United States)

    Berns, E.; Werth, C. J.; Valocchi, A. J.; Sanford, R. A.

    2015-12-01

    Biogeochemical interactions have been investigated extensively to characterize natural nutrient cycling and predict contaminant transport in surface and groundwater. Dissimilatory metal reducing bacteria, many of which form biofilms, play an important role in reducing a variety of metals in these systems. It has been shown that biofilm morphology is impacted by flow conditions, but there has been little work that explores how reduction kinetics change as a result of these different morphologies. Different flow rates may affect physical properties of the biofilm that influence the rate of substrate reduction. We introduce an approach to calculate changes in Monod kinetic parameters while simultaneously evaluating biofilm morphologies under different flow rates. A vertical, cylindrical flow cell with removable glass slide sections coated in manganese dioxide (electron acceptor) was used to grow a biofilm of Geobacter sulfurreducens with acetate as the electron donor under both high (50 mL/hr) and low (5 mL/h) flow rates. The removable sections allowed for visualization of the biofilm at different time points with a confocal microscope, and quantification of the biomass on the surface using a combination of a protein assay and image analysis. Data collected from the experiments was used to determine yield and specific growth rate at the different flow rates, and a simple numerical model was used to estimate the half saturation constant of manganese dioxide at both flow rates. A smaller half saturation constant was estimated at the higher flow rate, indicating that the biofilm was more efficient in the high flow system, but a strong correlation between morphology and the faster reduction rate was not observed. Monod kinetic parameters are important for the development of accurate nutrient cycling and contaminant transport models in natural environments, and understanding how they are impacted by flow will be important for the development of new, improved models.

  5. Techniques in Gas-Phase Thermolyses. Part 6. Pulse Pyrolysis: Gas Kinetic Studies in an Inductively Heated Flow Reactor

    DEFF Research Database (Denmark)

    Egsgaard, Helge; Bo, P.; Carlsen, Lars

    1985-01-01

    A prototype of an inductively heated flow reactor for gas kinetic studies is presented. The applicability of the system, which is based on a direct coupling between the reactor and the ion source of a mass spectrometer, is illustrated by investigations of a series of simple bond fission reactions...

  6. A well-balanced unified gas-kinetic scheme for multiscale flow transport under gravitational field

    Science.gov (United States)

    Xiao, Tianbai; Cai, Qingdong; Xu, Kun

    2017-03-01

    The gas dynamics under gravitational field is usually associated with multiple scale nature due to large density variation and a wide variation of local Knudsen number. It is challenging to construct a reliable numerical algorithm to accurately capture the non-equilibrium physical effect in different regimes. In this paper, a well-balanced unified gas-kinetic scheme (UGKS) for all flow regimes under gravitational field will be developed, which can be used for the study of non-equilibrium gravitational gas system. The well-balanced scheme here is defined as a method to evolve an isolated gravitational system under any initial condition to a hydrostatic equilibrium state and to keep such a solution. To preserve such a property is important for a numerical scheme, which can be used for the study of slowly evolving gravitational system, such as the formation of star and galaxy. Based on the Boltzmann model with external forcing term, the UGKS uses an analytic time-dependent (or scale-dependent) solution in the construction of the discretized fluid dynamic equations in the cell size and time step scales, i.e., the so-called direct modeling method. As a result, with the variation of the ratio between the numerical time step and local particle collision time, the UGKS is able to recover flow physics in different regimes and provides a continuous spectrum of gas dynamics. For the first time, the flow physics of a gravitational system in the transition regime can be studied using the UGKS, and the non-equilibrium phenomena in such a gravitational system can be clearly identified. Many numerical examples will be used to validate the scheme. New physical observation, such as the correlation between the gravitational field and the heat flux in the transition regime, will be presented. The current method provides an indispensable tool for the study of non-equilibrium gravitational system.

  7. Buoyant Turbulent Kinetic Energy Production in Steep-Slope Katabatic Flow

    Science.gov (United States)

    Oldroyd, Holly J.; Pardyjak, Eric R.; Higgins, Chad W.; Parlange, Marc B.

    2016-12-01

    We develop several critical concepts that should be considered when interpreting, modelling and designing future experiments for flows over sloping terrain. Vertical buoyancy fluxes in katabatic flows can be positive and a source of turbulent kinetic energy (TKE) despite the statically stable, thermal stratification that drives these flows. This phenomenon occurs when the ratio of along-slope to slope-normal kinematic heat fluxes is greater than the cotangent of the slope angle, suggesting a critical value of slope-angle steepness found in earlier studies. We provide field-data-based evidence that the along-slope heat flux may dominate the variables in this inequality, and therefore in generating buoyant TKE production or suppression over a steep slope. These data show the along-slope heat flux can be more variable and significantly larger in magnitude than the slope-normal component. The gradient Richardson number does not include the effects of the along-slope buoyancy; furthermore, none of the canonical stability parameters can properly reflect the TKE redistribution from turbulent transport divergence and the sink of TKE in cases of counter-gradient momentum fluxes, which we frequently observe near the peak of the katabatic jet. In such cases, canonical stability parameters inadequately represent the physical mechanisms associated with stability. These results have broad implications related to accurately modelling turbulence and surface exchanges over sloping terrain and illustrate the need to more thoroughly investigate the along-slope heat flux and its drivers, the meaning and definitions of stability, and the effects of non-local turbulent transport.

  8. Flow-cytometric method for observing the effects of pulsed electromagnetic fields on the growth of rat bone marrow mesenchymal stem cells%流式细胞仪观察脉冲电磁场干预骨髓间充质干细胞的生长

    Institute of Scientific and Technical Information of China (English)

    黄钊; 苏伟; 崔向荣; 覃万安

    2011-01-01

    BACKGROUND: Compared with the morphology, DNA electrophoresis and other methods, flow-cytometric method has more advantages on the detection of cell phenotype, proliferation rate and cell cycle of rat bone marrow mesenchymal stem cells (BMSCs). OBJECTIVE: To observe and discuss the effect of specific pulsed electromagnetic fields stimulation on the proliferation and cell cycle of rat BMSCs with the flow-cytometric method (FCM). METHODS: Rat BMSCs were exposed to pulsed electromagnetic fields (frequency for 1 kHz, magnetic flux density for 0.05 mT, power density for 5 mW/cm2). BMSCs without exposure to pulsed electromagnetic fields were used as controls. Expression of CD29, CD31, CD44 CD45, CD105 were detect in the control group. The proliferation rate and cell cycle of passage 3 cells were detected at 3, 6, 9, 12 days. RESULTS AND CONCLUSION: CD29, CD44, CD105 in the 3rd passage non-stimulated BMSCs was positively expressed and the CD31, CD45 was negatively expressed (P < 0.05). The survival rate of passage 3 BMSCs and percentage of S phase following intervention of pulsed electromagnetic fields were greater than those in the control group (P < 0.05). These results indicated that the specific pulsed electromagnetic fields can promote the growth and proliferation of BMSCs to some extent.%背景:应用流式细胞仪检测细胞表型、存活或凋亡细胞计数及细胞周期,较形态学观察、DNA电泳等检测方法更具优势.目的:采用流式细胞仪分选检测特定脉冲电磁场干预对大鼠骨髓间充质干细胞生长周期及其增殖效率的影响.方法:使用振荡频率1 kHz,磁感应强度0.05 mT、功率密度5 mW/cm2的脉冲电磁场照射大鼠骨髓间充质干细胞,以未经磁场干预的细胞为对照.采用流式细胞仪检测未经磁场干预细胞表面抗原CD29、CD31、CD44、CD45和CD105表达率;于传第3代后第3,6,9,12天测定不同干预细胞增殖率及生长周期.结果与结论:未经磁场干预的第3代

  9. A Fokker-Planck based kinetic model for diatomic rarefied gas flows

    Science.gov (United States)

    Gorji, M. Hossein; Jenny, Patrick

    2013-06-01

    A Fokker-Planck based kinetic model is presented here, which also accounts for internal energy modes characteristic for diatomic gas molecules. The model is based on a Fokker-Planck approximation of the Boltzmann equation for monatomic molecules, whereas phenomenological principles were employed for the derivation. It is shown that the model honors the equipartition theorem in equilibrium and fulfills the Landau-Teller relaxation equations for internal degrees of freedom. The objective behind this approximate kinetic model is accuracy at reasonably low computational cost. This can be achieved due to the fact that the resulting stochastic differential equations are continuous in time; therefore, no collisions between the simulated particles have to be calculated. Besides, because of the devised energy conserving time integration scheme, it is not required to resolve the collisional scales, i.e., the mean collision time and the mean free path of molecules. This, of course, gives rise to much more efficient simulations with respect to other particle methods, especially the conventional direct simulation Monte Carlo (DSMC), for small and moderate Knudsen numbers. To examine the new approach, first the computational cost of the model was compared with respect to DSMC, where significant speed up could be obtained for small Knudsen numbers. Second, the structure of a high Mach shock (in nitrogen) was studied, and the good performance of the model for such out of equilibrium conditions could be demonstrated. At last, a hypersonic flow of nitrogen over a wedge was studied, where good agreement with respect to DSMC (with level to level transition model) for vibrational and translational temperatures is shown.

  10. Reduced kinetic mechanism of n-heptane oxidation in modeling polycyclic aromatic hydrocarbon formation in opposed-flow diffusion flames

    Institute of Scientific and Technical Information of China (English)

    Beijing ZHONG; Jun XI

    2008-01-01

    A reduced mechanism, which could couple with the multidimensional computational fluid dynamics code for quantitative description of a reacting flow, was developed for chemical kinetic modeling of polycyclic aro-matic hydrocarbon formation in an opposed-flow dif-fusion flame. The complete kinetic mechanism, which comprises 572 reactions and 108 species, was reduced to a simplified mechanism that includes only 83 reactions and 56 species through sensitivity analysis. The results computed via this reduced mechanism are nearly indistin-guishable from those via the detailed mechanism, which demonstrate that the model based on this reduced mech-anism can properly describe n-heptane oxidation chem-istry and quantitatively predict polycyclic aromatic hydrocarbon (such as benzene, naphthalene, phenan-threne and pyrene) formation in opposed-flow diffusion flames.

  11. Intestinal intraepithelial lymphocyte cytometric pattern is more accurate than subepithelial deposits of anti-tissue transglutaminase IgA for the diagnosis of celiac disease in lymphocytic enteritis.

    Directory of Open Access Journals (Sweden)

    Fernando Fernández-Bañares

    Full Text Available BACKGROUND & AIMS: An increase in CD3+TCRγδ+ and a decrease in CD3- intraepithelial lymphocytes (IEL is a characteristic flow cytometric pattern of celiac disease (CD with atrophy. The aim was to evaluate the usefulness of both CD IEL cytometric pattern and anti-TG2 IgA subepithelial deposit analysis (CD IF pattern for diagnosing lymphocytic enteritis due to CD. METHODS: Two-hundred and five patients (144 females who underwent duodenal biopsy for clinical suspicion of CD and positive celiac genetics were prospectively included. Fifty had villous atrophy, 70 lymphocytic enteritis, and 85 normal histology. Eight patients with non-celiac atrophy and 15 with lymphocytic enteritis secondary to Helicobacter pylori acted as control group. Duodenal biopsies were obtained to assess both CD IEL flow cytometric (complete or incomplete and IF patterns. RESULTS: Sensitivity of IF, and complete and incomplete cytometric patterns for CD diagnosis in patients with positive serology (Marsh 1+3 was 92%, 85 and 97% respectively, but only the complete cytometric pattern had 100% specificity. Twelve seropositive and 8 seronegative Marsh 1 patients had a CD diagnosis at inclusion or after gluten free-diet, respectively. CD cytometric pattern showed a better diagnostic performance than both IF pattern and serology for CD diagnosis in lymphocytic enteritis at baseline (95% vs 60% vs 60%, p = 0.039. CONCLUSIONS: Analysis of the IEL flow cytometric pattern is a fast, accurate method for identifying CD in the initial diagnostic biopsy of patients presenting with lymphocytic enteritis, even in seronegative patients, and seems to be better than anti-TG2 intestinal deposits.

  12. 应用流式微球检测黄曲霉毒素B1方法的建立%Determination of aflatoxin B1 based on a flow cytometric microsphere immunoassay

    Institute of Scientific and Technical Information of China (English)

    李泳宁; 吴海燕; 郑允权; 郭养浩

    2012-01-01

    采用活性酯法,将AFB1-BSA人工抗原交联于含有羧基表面的荧光微球,通过与游离AFB1竞争抗AFB1单克隆抗体后,再与异硫氰酸荧光素标记二抗的反应,建立基于微球的间接竞争免疫检测方法.检测结果表明,流式细胞仪检测AFB1的检测限为0.03 ng·mL-1,检测范围为0.05 ~ 1.0 ng· mL-1.与黄曲霉毒素B2、黄曲霉毒素G1、黄曲霉毒素G2、黄曲霉毒素M1和黄曲霉毒素M2的交叉反应率均低于1.0%.在玉米样品中的加标回收率为87% ~ 103%,变异系数为6.1%~8.4%.%Carboxyl - modified microspheres internally dyed with fluroscent dye were conjugated with the artificial antigen AFB, - BSA. Aflatoxin B, ( AFB,) was used as a positive control to compete with the AFB, - BSA antigen on the surface of the microspheres for the anti - AFB, McAb. A fluorescein isothiocyanate labeled IgG reporter antibody was added to react specifically with the anti - AFB, McAb on the microspheres. The detection limit of AFB, reached 0.03 ng · mL-1, with a good linearity ranging 0.05 ~ 1.0 ng mL-1. The cross - reactivity rates were less than 1.0% with other toxins such as aflatoxins B2, aflatoxins G,, aflatoxins G2, aflatoxins M, and aflatoxins M2. The recovery of AFB, from artificially contaminated corn samples was from 89% to 92% , with CVs from 6.8% to 9.0%. A novel method for the determination of aflatoxin B, by an indirect competitive immunoassay with a flow cytometer has been developed.

  13. Analysis of the Fine-Scale Population Structure of “Candidatus Accumulibacter phosphatis” in Enhanced Biological Phosphorus Removal Sludge, Using Fluorescence In Situ Hybridization and Flow Cytometric Sorting▿

    Science.gov (United States)

    Kim, Jeong Myeong; Lee, Hyo Jung; Kim, Sun Young; Song, Jae Jun; Park, Woojun; Jeon, Che Ok

    2010-01-01

    To investigate the fine-scale diversity of the polyphosphate-accumulating organisms (PAO) “Candidatus Accumulibacter phosphatis” (henceforth referred to as “Ca. Accumulibacter”), two laboratory-scale sequencing batch reactors (SBRs) for enhanced biological phosphorus removal (EBPR) were operated with sodium acetate as the sole carbon source. During SBR operations, activated sludge always contained morphologically different “Ca. Accumulibacter” strains showing typical EBPR performances, as confirmed by the combined technique of fluorescence in situ hybridization (FISH) and microautoradiography (MAR). Fragments of “Ca. Accumulibacter” 16S rRNA genes were retrieved from the sludge. Phylogenetic analyses together with sequences from the GenBank database showed that “Ca. Accumulibacter” 16S rRNA genes of the EBPR sludge were clearly differentiated into four “Ca. Accumulibacter” clades, Acc-SG1, Acc-SG2, Acc-SG3, and Acc-SG4. The specific FISH probes Acc444, Acc184, Acc72, and Acc119 targeting these clades and some helpers and competitors were designed by using the ARB program. Microbial characterization by FISH analysis using specific FISH probes also clearly indicated the presence of different “Ca. Accumulibacter” cell morphotypes. Especially, members of Acc-SG3, targeted by probe Acc72, were coccobacillus-shaped cells with a size of approximately 2 to 3 μm, while members of Acc-SG1, Acc-SG2, and Acc-SG4, targeted by Acc444, Acc184, and Acc119, respectively, were coccus-shaped cells approximately 1 μm in size. Subsequently, cells targeted by each FISH probe were sorted by use of a flow cytometer, and their polyphosphate kinase 1 (ppk1) gene homologs were amplified by using a ppk1-specific PCR primer set for “Ca. Accumulibacter.” The phylogenetic tree based on sequences of the ppk1 gene homologs was basically congruent with that of the 16S rRNA genes, but members of Acc-SG3 with a distinct morphology comprised two different ppk1 genes

  14. Mechanism of kinetic energy transfer in homogeneous bidisperse gas-solid flow and its implications for segregation

    Science.gov (United States)

    Mehrabadi, Mohammad; Subramaniam, Shankar

    2017-02-01

    Most gas-solid flows encountered in nature and industrial applications are polydisperse, and the segregation or mixing of particle classes in polydisperse gas-solid flows is a phenomenon of great practical importance. A statistically homogeneous gas-solid flow with a bidisperse distribution (in size or density) of particles is a canonical representation of polydisperse flows. A key feature that distinguishes the bidisperse flow from its monodisperse counterpart is the exchange of momentum and kinetic energy between the particle classes due to collisions, which are important for applications outside the very dilute regime. The average exchange of linear momentum between particle classes due to collisions occurs through the particle-particle drag term. The conservation equations for average momentum corresponding to each particle class can be used to deduce the average slip velocity between the particle size and density classes, which is the signature of particle segregation. In this canonical problem, the steady value of particle mean slip velocity results from a balance between three terms, each in turn involving the body force or the mean fluid pressure gradient, the gas-particle drag, and the particle-particle drag. The particle-particle drag depends on the particle velocity fluctuations in each class [Louge, M. Y. et al., "The role of particle collisions in pneumatic transport," J. Fluid Mech. 231, 345-359 (1991)], thereby coupling the mean and second-moment equations. For monodisperse gas-solid flows the transfer of kinetic energy from the mean to second-moment equations was explained by Subramaniam and co-workers who proposed the conservation of interphase turbulent kinetic energy transfer principle [Xu, Y. and Subramaniam, S., "Consistent modeling of interphase turbulent kinetic energy transfer in particle-laden turbulent flows," Phys. Fluids 19(8), 085101 (2007)], and this was subsequently verified by particle-resolved direct numerical simulation [Mehrabadi

  15. Experimental and detailed kinetic modeling study of PAH formation in laminar co-flow methane diffusion flames

    DEFF Research Database (Denmark)

    Cuoci, Alberto; Frassoldati, Alessio; Faravelli, Tiziano

    2013-01-01

    In the present paper, synchrotron VUV photoionization mass spectrometry is used to study the detailed chemistry of co-flow methane diffusion flames with different dilution ratios. The experimental results constitute a comprehensive characterization of species important for PAH and soot formation...... an original CFD code based on the operator-splitting technique, specifically conceived to handle large kinetic mechanisms. The detailed kinetic modeling was effectively used to describe and analyze the fuel consumption and the formation of PAH. Experimental measurements and numerical predictions were found...

  16. Flow Cytometric Evidence for Hydroxyl Radical-induced Apoptosis in Tobacco Protoplasts%羟自由基诱导的烟草原生质体的凋亡:流式细胞法的新证据

    Institute of Scientific and Technical Information of China (English)

    雷晓勇; 廖旭东; 张贵友; 戴尧仁

    2003-01-01

    用1.0 mmol/L FeSO4/0.5 mmol/L H202处理烟草(Nicotiana tabacum L.cultivar BY 2)原生质体,发现羟自由基能够诱导烟草原生质体的凋亡.具体表现为细胞核皱缩、DNA Ladder、TUNEL阳性反应等典型的凋亡特征.在动物细胞凋亡过程中,线粒体起着非常重要的作用,其中膜电位(△ψm)的变化以及由其引起的位于线粒体膜上的通透性孔(PTP)的开放与Cyt c的释放有关.另外,在动物凋亡细胞中,磷脂酰丝氨酸(phosphatidyl serine,PS)会从细胞膜内侧向外翻转.为了判断植物细胞凋亡过程中膜电位的变化情况以及PS的外翻程度,我们采用了流式细胞法.结果表明,随着处理时间的延长,烟草原生质体线粒体的膜电位逐渐降低;膜内PS大量外翻.说明由羟自由基和烟草原生质体组成的凋亡体系是一种可靠的凋亡组合,可以用来对植物细胞凋亡机理做进一步研究.%Protoplasts prepared from tobacco (Nicotiana tabacum L., cultivar BY-2) suspension cellshave similar morphological characteristics to those in animal cells. The hallmarks of apoptosis such ascondensation and peripheral distribution of nuclei, TUNEL positive reaction, and DNA ladders were ob-served when tobacco protoplasts were treated with the hydroxyl radical generating system (1.0 mmol/LFeSO4/0.5 mmol/L H2O2). In animals, the loss of transmembrane potential (△ψm ) and the exposure ofphospholipid phosphatidylserine (PS) are believed to be the main apoptosis events. To test whether thesesignificant processes take place in plants, flow cytometry was used to detect annexin V binding andchanges in △ψm. Results showed that the PS turned out from inner membrane and △ψm graduallydecreased during the apoptosis. All these apoptotic characteristics proved that hydroxyl radicals cancause typical programmed cell death (PCD) in tobacco protoplasts and this design can be served as aneffective experiment system to explore the mechanism of plant apoptosis.

  17. Chemical Kinetic Modeling of Dimethyl Carbonate in an Opposed-Flow Diffusion Flame

    Energy Technology Data Exchange (ETDEWEB)

    Glaude, P A; Pitz, W J; Thomson, M J

    2003-12-08

    Dimethyl carbonate (DMC) has been of interest as an oxygenate additive to diesel fuel because of its high oxygen content. In this study, a chemical kinetic mechanism for DMC was developed for the first time and used to understand its combustion under conditions in an opposed flow diffusion flame. Computed results were compared to experimental results from an opposed flow diffusion flame. It was found that the decomposition rate DMC {yields} H{sub 3}COC(=O)O. + CH{sub 3} in the flame was much slower than originally thought because resonance stabilization in the H{sub 3}COC(=O)O. radical was less than expected. Also, a new molecular elimination path for DMC is proposed and its rate calculated by quantum chemical methods. In the simulations of DMC in the flame, it was determined that much of the oxygen in dimethyl carbonate goes directly to CO{sub 2}. This characteristic indicates that DMC would not be an effective oxygenate additive for reducing soot emissions from diesel engines. In an ideal oxygenate additive for diesel fuel, each oxygen atom stays bonded to one carbon atom in the products thereby preventing the formation of carbon-carbon bonds that can lead to soot. When CO2 is formed directly, two oxygen atoms are bonded to one carbon atom thereby wasting one oxygen atom in the oxygenate additive. To determine how much CO{sub 2} is formed directly, the branching ratio of the key reaction, CH{sub 3}OC.=O going to the products CH{sub 3} + CO{sub 2} or CH{sub 3}O + CO was determined by ab initio methods. The A-factors of the rate constant of this reaction were found to be about 20 times higher than previous factors estimates. The new reaction rate constants obtained can be used as reaction rate rules for all oxygenates that contain the ester moiety including biodiesel.

  18. Experimental and Kinetic Modeling Study of Extinction and Ignition of Methyl Decanoate in Laminar Nonpremixed Flows

    Energy Technology Data Exchange (ETDEWEB)

    Seshadri, K; Lu, T; Herbinet, O; Humer, S; Niemann, U; Pitz, W J; Law, C K

    2008-01-09

    Methyl decanoate is a large methyl ester that can be used as a surrogate for biodiesel. In this experimental and computational study, the combustion of methyl decanoate is investigated in nonpremixed, nonuniform flows. Experiments are performed employing the counterflow configuration with a fuel stream made up of vaporized methyl decanoate and nitrogen, and an oxidizer stream of air. The mass fraction of fuel in the fuel stream is measured as a function of the strain rate at extinction, and critical conditions of ignition are measured in terms of the temperature of the oxidizer stream as a function of the strain rate. It is not possible to use a fully detailed mechanism for methyl decanoate to simulate the counterflow flames because the number of species and reactions is too large to employ with current flame codes and computer resources. Therefore a skeletal mechanism was deduced from a detailed mechanism of 8555 elementary reactions and 3036 species using 'directed relation graph' method. This skeletal mechanism has only 713 elementary reactions and 125 species. Critical conditions of ignition were calculated using this skeletal mechanism and are found to agree well with experimental data. The predicted strain rate at extinction is found to be lower than the measurements. In general, the methyl decanoate mechanism provides a realistic kinetic tool for simulation of biodiesel fuels.

  19. Fully Nonlinear Edge Gyrokinetic Simulations of Kinetic Geodesic-Acoustic Modes and Boundary Flows

    Energy Technology Data Exchange (ETDEWEB)

    Xu, X Q; Belli, E; Bodi, K; Candy, J; Chang, C S; Cohen, B I; Cohen, R H; Colella, P; Dimits, A M; Dorr, M R; Gao, Z; Hittinger, J A; Ko, S; Krasheninnikov, S; McKee, G R; Nevins, W M; Rognlien, T D; Snyder, P B; Suh, J; Umansky, M V

    2008-09-18

    We present edge gyrokinetic neoclassical simulations of tokamak plasmas using the fully nonlinear (full-f) continuum code TEMPEST. A nonlinear Boltzmann model is used for the electrons. The electric field is obtained by solving the 2D gyrokinetic Poisson Equation. We demonstrate the following: (1) High harmonic resonances (n > 2) significantly enhance geodesic-acoustic mode (GAM) damping at high-q (tokamak safety factor), and are necessary to explain both the damping observed in our TEMPEST q-scans and experimental measurements of the scaling of the GAM amplitude with edge q{sub 95} in the absence of obvious evidence that there is a strong q dependence of the turbulent drive and damping of the GAM. (2) The kinetic GAM exists in the edge for steep density and temperature gradients in the form of outgoing waves, its radial scale is set by the ion temperature profile, and ion temperature inhomogeneity is necessary for GAM radial propagation. (3) The development of the neoclassical electric field evolves through different phases of relaxation, including GAMs, their radial propagation, and their long-time collisional decay. (4) Natural consequences of orbits in the pedestal and scrape-off layer region in divertor geometry are substantial non-Maxwellian ion distributions and flow characteristics qualitatively like those observed in experiments.

  20. A Well-Balanced Unified Gas-Kinetic Scheme for Multiscale Flow Transport Under Gravitational Field

    CERN Document Server

    Xiao, Tianbai; Xu, Kun

    2016-01-01

    The gas dynamics under gravitational field is usually associated with the multiple scale nature due to large density variation and a wide range of local Knudsen number. It is chal- lenging to construct a reliable numerical algorithm to accurately capture the non-equilibrium physical effect in different regimes. In this paper, a well-balanced unified gas-kinetic scheme (UGKS) for all flow regimes under gravitational field will be developed, which can be used for the study of non-equilibrium gravitational gas system. The well-balanced scheme here is defined as a method to evolve an isolated gravitational system under any initial condition to an isothermal hydrostatic equilibrium state and to keep such a solution. To preserve such a property is important for a numerical scheme, which can be used for the study of slowly evolving gravitational system, such as the formation of star and galaxy. Based on the Boltzmann model with external forcing term, an analytic time evolving (or scale-dependent) solution is constru...

  1. On-line study of growth kinetics of single hyphae of Aspergillus oryzae in a flow-through cell

    DEFF Research Database (Denmark)

    Christiansen, Torben; Spohr, Anders Bendsen; Nielsen, Jens Bredal

    1999-01-01

    the outgrowth of a hyphal element from a single spore using a Monte Carlo simulation technique. The simulations shows that the observed kinetics for the individual hyphae result in an experimentally verified growth pattern with exponential growth in both total hyphal length and number of tips. (C) 1999 John......Using image analysis the growth kinetics of the single hyphae of the filamentous fungus Aspergillus oryzae has been determined on-line in a flow-through cell at different glucose concentrations in the range from 26 mg L-1 to 20 g L-1. The tip extension rate of the individual hyphae can be described...... branching occurs, it is observed that the tip extension rate decreases temporarily. The number of branches formed on a hypha is proportional to the length of the hypha that exceeds a certain minimum length required to support the growth of a new branch. The observed kinetics has been used to simulate...

  2. Seasonality in molecular and cytometric diversity of marine bacterioplankton: the reshuffling of bacterial taxa by vertical mixing

    KAUST Repository

    García, Francisca C.

    2015-07-17

    The ’cytometric diversity’ of phytoplankton communities has been studied based on single-cell properties, but the applicability of this method to characterize bacterioplankton has been unexplored. Here, we analysed seasonal changes in cytometric diversity of marine bacterioplankton along a decadal time-series at three coastal stations in the Southern Bay of Biscay. Shannon-Weaver diversity estimates and Bray-Curtis similarities obtained by cytometric and molecular (16S rRNA tag sequencing) methods were significantly correlated in samples from a 3.5-year monthly time-series. Both methods showed a consistent cyclical pattern in the diversity of surface bacterial communities with maximal values in winter. The analysis of the highly resolved flow cytometry time-series across the vertical profile showed that water column mixing was a key factor explaining the seasonal changes in bacterial composition and the winter increase in bacterial diversity in coastal surface waters. Due to its low cost and short processing time as compared to genetic methods, the cytometric diversity approach represents a useful complementary tool in the macroecology of aquatic microbes.

  3. Implicit gas-kinetic unified algorithm based on multi-block docking grid for multi-body reentry flows covering all flow regimes

    Science.gov (United States)

    Peng, Ao-Ping; Li, Zhi-Hui; Wu, Jun-Lin; Jiang, Xin-Yu

    2016-12-01

    Based on the previous researches of the Gas-Kinetic Unified Algorithm (GKUA) for flows from highly rarefied free-molecule transition to continuum, a new implicit scheme of cell-centered finite volume method is presented for directly solving the unified Boltzmann model equation covering various flow regimes. In view of the difficulty in generating the single-block grid system with high quality for complex irregular bodies, a multi-block docking grid generation method is designed on the basis of data transmission between blocks, and the data structure is constructed for processing arbitrary connection relations between blocks with high efficiency and reliability. As a result, the gas-kinetic unified algorithm with the implicit scheme and multi-block docking grid has been firstly established and used to solve the reentry flow problems around the multi-bodies covering all flow regimes with the whole range of Knudsen numbers from 10 to 3.7E-6. The implicit and explicit schemes are applied to computing and analyzing the supersonic flows in near-continuum and continuum regimes around a circular cylinder with careful comparison each other. It is shown that the present algorithm and modelling possess much higher computational efficiency and faster converging properties. The flow problems including two and three side-by-side cylinders are simulated from highly rarefied to near-continuum flow regimes, and the present computed results are found in good agreement with the related DSMC simulation and theoretical analysis solutions, which verify the good accuracy and reliability of the present method. It is observed that the spacing of the multi-body is smaller, the cylindrical throat obstruction is greater with the flow field of single-body asymmetrical more obviously and the normal force coefficient bigger. While in the near-continuum transitional flow regime of near-space flying surroundings, the spacing of the multi-body increases to six times of the diameter of the single

  4. A unified gas-kinetic scheme for continuum and rarefied flows IV: Full Boltzmann and model equations

    Science.gov (United States)

    Liu, Chang; Xu, Kun; Sun, Quanhua; Cai, Qingdong

    2016-06-01

    Fluid dynamic equations are valid in their respective modeling scales, such as the particle mean free path scale of the Boltzmann equation and the hydrodynamic scale of the Navier-Stokes (NS) equations. With a variation of the modeling scales, theoretically there should have a continuous spectrum of fluid dynamic equations. Even though the Boltzmann equation is claimed to be valid in all scales, many Boltzmann solvers, including direct simulation Monte Carlo method, require the cell resolution to the order of particle mean free path scale. Therefore, they are still single scale methods. In order to study multiscale flow evolution efficiently, the dynamics in the computational fluid has to be changed with the scales. A direct modeling of flow physics with a changeable scale may become an appropriate approach. The unified gas-kinetic scheme (UGKS) is a direct modeling method in the mesh size scale, and its underlying flow physics depends on the resolution of the cell size relative to the particle mean free path. The cell size of UGKS is not limited by the particle mean free path. With the variation of the ratio between the numerical cell size and local particle mean free path, the UGKS recovers the flow dynamics from the particle transport and collision in the kinetic scale to the wave propagation in the hydrodynamic scale. The previous UGKS is mostly constructed from the evolution solution of kinetic model equations. Even though the UGKS is very accurate and effective in the low transition and continuum flow regimes with the time step being much larger than the particle mean free time, it still has space to develop more accurate flow solver in the region, where the time step is comparable with the local particle mean free time. In such a scale, there is dynamic difference from the full Boltzmann collision term and the model equations. This work is about the further development of the UGKS with the implementation of the full Boltzmann collision term in the region

  5. Sensitivity of heat fluxes in hypersonic CO2 flows to the state-to-state kinetic schemes

    Science.gov (United States)

    Armenise, I.; Kustova, E.

    2016-11-01

    Kinetics and heat transfer in a CO2/CO/O2/O/C mixture in a hypersonic boundary layer is studied using a state-to-state vibrational-chemical kinetic model. The CO2 molecule is detailed in its symmetric stretching, bending and asymmetric stretching modes, which are strongly coupled through inter-mode vibrational energy transfers. Two sets of rate coefficients for the vibrational energy transitions are used. Different kinetic schemes including various physical and chemical processes are assessed. The heat flux is calculated, in the framework of the modified Chapman-Enskog theory, accounting for the vibrational states of involved molecules. Comparisons with results obtained using a simplified model, including mainly vibrational levels of the asymmetric stretching mode, are carried out. It is shown that VT transitions in the symmetric and asymmetric modes do not alter the flow and can be neglected. The heat flux is not sensitive to the rates of vibrational energy transitions but depends noticeably on the processes implemented to the kinetic scheme. Using the simplified model yields under-predicted surface heat fluxes; nevertheless we can recommend it for fast estimates of the fluid dynamic variables and heat transfer in hypersonic flows since its implementation essentially reduces computational costs.

  6. Mechanism of Cph1 phytochrome assembly from stopped-flow kinetics and circular dichroism.

    Science.gov (United States)

    Borucki, Berthold; Otto, Harald; Rottwinkel, Gregor; Hughes, Jonathan; Heyn, Maarten P; Lamparter, Tilman

    2003-11-25

    The kinetics and mechanism of the autocatalytic assembly of holo-Cph1 phytochrome (from Synechocystis) from the apoprotein and the bilin chromophores phycocyanobilin (PCB) and phycoerythrobilin (PEB) were investigated by stopped flow and circular dichroism. At 1:1 stoichiometry, pH 7.9, and 10 degrees C, SVD analysis of the kinetic data for PCB revealed three spectral components involving three transitions with time constants tau(1) approximately 150 ms, tau(2) approximately 2.5 s, and tau(3) approximately 50 s. Tau(1) was associated with a major red shift and transfer of oscillator strength from the Soret region to the 680 nm region. When the sulfhydryl group of cysteine 259 was blocked with iodoacetamide, preventing the formation of a covalent adduct, a noncovalent red-shifted complex (680 nm) was formed with a time constant of 200 ms. Tau(1) could thus be assigned to the formation of a noncovalent complex. The absorption changes during tau(1) are due to the formation of the extended conformation of the linear tetrapyrrole and to its protonation in the binding pocket. From the concentration and pH dependence of the kinetics we obtained a value of 1.5 microM for the K(D) of this noncovalent complex and a value of 8.4 for the pK(a) of the proton donor. The tau(2) component was associated with a blue shift of about 25 nm and was attributed to the formation of the covalent bond (P(r)), accompanied with the loss of the 3-3' double bond to ring A. Tau(3) was due to photoconversion to P(fr). For PEB, which is not photochromic, the formation of the noncovalent complex is faster (tau(1) = 70 ms), but the covalent bond formation is about 80 times slower (tau(2) = 200 s) than with the natural chromophore PCB. The CD spectra of the PCB adduct in the 250-800 nm range show that the chromophore geometries in P(r) and P(fr) are similar to those in plant phytochrome. The opposite rotational strengths of P(r) and P(fr) in the longest wavelength band suggest that the

  7. Flow cytometric detection of viruses in the Zuari estuary, Goa

    Digital Repository Service at National Institute of Oceanography (India)

    Mitbavkar, S.; Rajaneesh, K.M.; SathishKumar, P.

    and microalgae, the most abundant organisms in the ocean and also micro- zooplankton 2 . They have been implicated in phytoplankton mortality and the de- cline of phytoplankton blooms 1 . Marine phytoplankton is responsible for up to half of the total primary...

  8. Flow cytometric immunophenotyping test for staging/monitoring neuroblastoma patients

    National Research Council Canada - National Science Library

    Warzynski, Michael J; Graham, David M; Axtell, Richard A; Higgins, James V; Hammers, Yuki A

    2002-01-01

    .... Following the “rare event” philosophy of selecting one negative and two positive antigens, we initially tried a “cocktail” of CD45 − CD56 very bright+ neuron‐specific enolase (NSE) cytoplasmic...

  9. Flow Cytometric Ploidy Determination of Oral Premalignant and Malignant Lesions

    Science.gov (United States)

    1990-01-01

    However, subjectivity remains an inherent part of the diagnostic process.9 According to Dabelsteen,10 subjectivity is most apparent in the determination...of Silverman 16 and Pindborg43 and Mincer et al.21 that light microscopio features of premalignancy are often not present in original biopsy specimens...was caused either in whole or in part by the presence of doublets was eliminated by syringina,. filtering and visually examining the population. In

  10. Flow Cytometric Applicability of Fluorescent Vitality Probes on Phytoplankton

    NARCIS (Netherlands)

    Peperzak, L.; Brussaard, C.P.D.

    2011-01-01

    The applicability of six fluorescent probes (four esterase probes: acetoxymethyl ester of Calcein [Calcein-AM], 5-chloromethylfluorescein diacetate [CMFDA], fluorescein diacetate [FDA], and 2',7'-dichlorofluorescein diacetate [H(2)DCFDA]; and two membrane probes: bis-(1,3-dibutylbarbituric acid) tri

  11. Nonlinear interactions between the pumping kinetics, fluid dynamics and optical resonator of cw fluid flow lasers. Final technical report

    Energy Technology Data Exchange (ETDEWEB)

    Sentman, L.H.; Nayfeh, M.H.

    1983-12-01

    This research is an integrated theoretical and experimental investigation of the nonlinear interactions which may occur between the chemical kinetics, the fluid dynamics and the unstable resonator of a continuous wave fluid flow laser. The objectives of this grant were to measure the frequency and amplitude of the time dependent pulsations in the power spectral output which have been predicted to occur in cw chemical lasers employing unstable resonators to extract power.

  12. A kinetic study of solar wind electrons in the transition region from collision dominated to collisionless flow

    Science.gov (United States)

    Lie-Svendsen, O.; Leer, E.

    1995-01-01

    We have studied the evolution of the velocity distribution function of a test population of electrons in the solar corona and inner solar wind region, using a recently developed kinetic model. The model solves the time dependent, linear transport equation, with a Fokker-Planck collision operator to describe Coulomb collisions between the 'test population' and a thermal background of charged particles, using a finite differencing scheme. The model provides information on how non-Maxwellian features develop in the distribution function in the transition region from collision dominated to collisionless flow. By taking moments of the distribution the evolution of higher order moments, such as the heat flow, can be studied.

  13. Cytometric Approach for Detection of Encephalitozoon intestinalis, an Emergent Agent▿

    Science.gov (United States)

    Barbosa, Joana; Rodrigues, Acácio Gonçalves; Pina-Vaz, Cidália

    2009-01-01

    Encephalitozoon intestinalis is responsible for intestinal disease in patients with AIDS and immunocompetent patients. The infectious form is a small spore that is resistant to water treatment procedures. Its detection is very important, but detection is very cumbersome and time-consuming. Our main objective was to develop and optimize a specific flow cytometric (FC) protocol for the detection of E. intestinalis in hospital tap water and human feces. To determine the optimal specific antibody (Microspor-FA) concentration, a known concentration of E. intestinalis spores (Waterborne, Inc.) was suspended in hospital tap water and stool specimens with different concentrations of Microspor-FA, and the tap water and stool specimens were incubated under different conditions. The sensitivity limit and specificity were also evaluated. To study spore infectivity, double staining with propidium iodide (PI) and Microspor-FA was undertaken. Distinct approaches for filtration and centrifugation of the stool specimens were used. E. intestinalis spores stained with 10 μg/ml of Microspor-FA at 25°C overnight provided the best results. The detection limit was 5 × 104 spores/ml, and good specificity was demonstrated. Simultaneous staining with Microspor-FA and PI ensured that the E. intestinalis spores were dead and therefore noninfectious. With the stool specimens, better spore recovery was observed with a saturated solution of NaCl and centrifugation at 1,500 × g for 15 min. A new approach for the detection of E. intestinalis from tap water or human feces that ensures that the spores are not viable is now available and represents an important step for the prevention of this threat to public health. PMID:19439525

  14. Kinetic analysis of 18F-fluorodihydrorotenone as a deposited myocardial flow tracer: Comparison to thallium-201.

    Energy Technology Data Exchange (ETDEWEB)

    Marshall, Robert C.; Powers-Risius, Patricia; Reutter, Bryan W.; O' Neil, James P.; La Belle, Michael; Huesman, Ronald H.; VanBrocklin, Henry F.

    2004-03-01

    The goal of this investigation was to assess the accuracy of 18F-fluorodihydrorotenone (18F-FDHR) as a new deposited myocardial flow tracer and compare the results to those for 201Tl. Methods. The kinetics of these flow tracers were evaluated in 22 isolated, erythrocyte- and albumin-perfused rabbit hearts over a flow range encountered in patients. The two flow tracers plus a vascular reference tracer (131I-albumin) were introduced as a bolus through a port just above the aortic cannula. Myocardial extraction, retention, washout, and uptake parameters were computed from the venous outflow curves using the multiple indicator dilution technique and spectral analysis. Results. The mean initial extraction fractions of 18F-FDHR (0.85 +- 0.07) and 201Tl (0.87 +- 0.05) were not significantly different, although the initial extraction fraction for 18F-FDHR declined with flow (P < 0.0001), whereas the initial extraction fraction of 201Tl did not. Washout of 201Tl was faster (P < 0.001) and more affected by flow (P < 0.05) than 18F-FDHR washout. Except for initial extraction fraction, 18F-FDHR retention was greater (P < 0.001) and less affected by flow (P < 0.05) than 201Tl retention. Reflecting its superior retention, net uptake of 18F-FDHR was better correlated with flow than 201Tl uptake at both one and fifteen minutes after tracer introduction (P < 0.0001 for both comparisons). Conclusion. The superior correlation of 18F-FDHR uptake with flow indicates that it is a better flow tracer than 201Tl in the isolated rabbit heart. Compared to the other currently available positron-emitting flow tracers (82Rb, 13N-ammonia, and 15O-water), 18F-FDHR has the potential of providing excellent image resolution without the need for an on-site cyclotron.

  15. An immersed-boundary method for compressible viscous flow and its application in gas-kinetic BGK scheme

    CERN Document Server

    Yuan, Ruifeng

    2016-01-01

    An immersed-boundary (IB) method is proposed and applied in the gas-kinetic BGK scheme to simulate incompressible/compressible viscous flow with stationary/moving boundary. In present method the ghost-cell technique is adopted to fulfill the boundary condition on the immersed boundary. A novel idea "local boundary determination" is put forward to identify the ghost cells, each of which may have several different ghost-cell constructions corresponding to different boundary segments, thus eliminating the singularity of the ghost cell. Furthermore, the so-called "fresh-cell" problem when implementing the IB method in moving-boundary simulation is resolved by a simple extrapolation in time. The method is firstly applied in the gas-kinetic BGK scheme to simulate the Taylor-Couette flow, where the second-order spatial accuracy of the method is validated and the "super-convergence" of the BGK scheme is observed. Then the test cases of supersonic flow around a stationary cylinder, incompressible flow around an oscill...

  16. New Chemical Kinetics Approach for DSMC Applications to Nonequilibrium Flows Project

    Data.gov (United States)

    National Aeronautics and Space Administration — A new chemical kinetics model and database will be developed for aerothermodynamic analyses on entry vehicles. Unique features of this model include (1) the ability...

  17. Monodomain dynamics for rigid rod and platelet suspensions in strongly coupled coplanar linear flow and magnetic fields. II. Kinetic theory

    Science.gov (United States)

    Forest, M. Gregory; Sircar, Sarthok; Wang, Qi; Zhou, Ruhai

    2006-10-01

    We establish reciprocity relations of the Doi-Hess kinetic theory for rigid rod macromolecular suspensions governed by the strong coupling among an excluded volume potential, linear flow, and a magnetic field. The relation provides a reduction of the flow and field driven Smoluchowski equation: from five parameters for coplanar linear flows and magnetic field, to two field parameters. The reduced model distinguishes flows with a rotational component, which map to simple shear (with rate parameter) subject to a transverse magnetic field (with strength parameter), and irrotational flows, for which the reduced model consists of a triaxial extensional flow (with two extensional rate parameters). We solve the Smoluchowski equation of the reduced model to explore: (i) the effect of introducing a coplanar magnetic field on each sheared monodomain attractor of the Doi-Hess kinetic theory and (ii) the coupling of coplanar extensional flow and magnetic fields. For (i), we show each sheared attractor (steady and unsteady, with peak axis in and out of the shearing plane, periodic and chaotic orbits) undergoes its own transition sequence versus magnetic field strength. Nonetheless, robust predictions emerge: out-of-plane degrees of freedom are arrested with increasing field strength, and a unique flow-aligning or tumbling/wagging limit cycle emerges above a threshold magnetic field strength or modified geometry parameter value. For (ii), irrotational flows coupled with a coplanar magnetic field yield only steady states. We characterize all (generically biaxial) equilibria in terms of an explicit Boltzmann distribution, providing a natural generalization of analytical results on pure nematic equilibria [P. Constantin, I. Kevrekidis, and E. S. Titi, Arch. Rat. Mech. Anal. 174, 365 (2004); P. Constantin, I. Kevrekidis, and E. S. Titi, Discrete and Continuous Dynamical Systems 11, 101 (2004); P. Constantin and J. Vukadinovic, Nonlinearity 18, 441 (2005); H. Liu, H. Zhang, and P

  18. Iron oxidation kinetics and phosphate immobilization along the flow-path from groundwater into surface water

    Directory of Open Access Journals (Sweden)

    B. van der Grift

    2014-06-01

    Full Text Available The retention of phosphorus in surface waters though co-precipitation of phosphate with Fe-oxyhydroxides during exfiltration of anaerobic Fe(II rich groundwater is not well understood. We developed an experimental field set-up to study Fe(II oxidation and P immobilization along the flow-path from groundwater to surface water in an agricultural experimental catchment of a small lowland river. We physically separated tube drain effluent from groundwater discharge before it entered a ditch in an agricultural field. Through continuous discharge measurements and weekly water quality sampling of groundwater, tube drain water, exfiltrated groundwater, and ditch water, we investigated Fe(II oxidation kinetics and P immobilization processes. The oxidation rate inferred from our field measurements closely agreed with the general rate law for abiotic oxidation of Fe(II by O2. Seasonal changes in climatic conditions affected the Fe(II oxidation process. Lower pH and lower temperatures in winter (compared to summer resulted in low Fe oxidation rates. After exfiltration to the surface water, it took a couple of days to more than one week before complete oxidation of Fe(II is reached. In summer time, Fe oxidation rates were much higher. The Fe concentrations in the exfiltrated groundwater were low, indicating that dissolved Fe(II is completely oxidized prior to inflow into a ditch. While the Fe oxidation rates reduce drastically from summer to winter, P concentrations remained high in the groundwater and an order of magnitude lower in the surface water throughout the year. This study shows very fast immobilisation of dissolved P during the initial stage of the Fe(II oxidation proces which results in P-depleted water before Fe(II is competly depleted. This cannot be explained by surface complexation of phosphate to freshly formed Fe-oxyhydroxides but indicates the formation of Fe(III-phosphate precipitates. The formation of Fe(III-phosphates at redox gradients

  19. Electro-hydrodynamics and kinetic modelling of polluted air flow activated by multi-tip-to-plane corona discharge

    Energy Technology Data Exchange (ETDEWEB)

    Meziane, M.; Eichwald, O.; Ducasse, O.; Marchal, F. [Universite de Toulouse, UPS, INPT, LAPLACE (Laboratoire Plasma et Conversion d' Energie), Toulouse Cedex 9 F-31062 (France); Sarrette, J. P.; Yousfi, M. [Universite de Toulouse, UPS, INPT, LAPLACE (Laboratoire Plasma et Conversion d' Energie), Toulouse Cedex 9 F-31062 (France); CNRS, LAPLACE, Toulouse F-31062 (France)

    2013-04-21

    The present paper is devoted to the 2D simulation of an Atmospheric Corona Discharge Reactor (ACDR) involving 10 pins powered by a DC high voltage and positioned 7 mm above a grounded metallic plane. The corona reactor is periodically crossed by thin mono filamentary streamers with a natural repetition frequency of some tens of kHz. The simulation involves the electro-dynamic, chemical kinetic, and neutral gas hydrodynamic phenomena that influence the kinetics of the chemical species transformation. Each discharge stage (including the primary and the secondary streamers development and the resulting thermal shock) lasts about one hundred nanoseconds while the post-discharge stages occurring between two successive discharge phases last one hundred microseconds. The ACDR is crossed by a lateral air flow including 400 ppm of NO. During the considered time scale of 10 ms, one hundred discharge/post-discharge cycles are simulated. The simulation involves the radical formation and thermal exchange between the discharges and the background gas. The results show how the successive discharges activate the flow gas and how the induced turbulence phenomena affect the redistribution of the thermal energy and the chemical kinetics inside the ACDR.

  20. Electro-hydrodynamics and kinetic modelling of polluted air flow activated by multi-tip-to-plane corona discharge

    Science.gov (United States)

    Meziane, M.; Eichwald, O.; Sarrette, J. P.; Ducasse, O.; Yousfi, M.; Marchal, F.

    2013-04-01

    The present paper is devoted to the 2D simulation of an Atmospheric Corona Discharge Reactor (ACDR) involving 10 pins powered by a DC high voltage and positioned 7 mm above a grounded metallic plane. The corona reactor is periodically crossed by thin mono filamentary streamers with a natural repetition frequency of some tens of kHz. The simulation involves the electro-dynamic, chemical kinetic, and neutral gas hydrodynamic phenomena that influence the kinetics of the chemical species transformation. Each discharge stage (including the primary and the secondary streamers development and the resulting thermal shock) lasts about one hundred nanoseconds while the post-discharge stages occurring between two successive discharge phases last one hundred microseconds. The ACDR is crossed by a lateral air flow including 400 ppm of NO. During the considered time scale of 10 ms, one hundred discharge/post-discharge cycles are simulated. The simulation involves the radical formation and thermal exchange between the discharges and the background gas. The results show how the successive discharges activate the flow gas and how the induced turbulence phenomena affect the redistribution of the thermal energy and the chemical kinetics inside the ACDR.

  1. Relationship between changes in pulmonary V̇O₂ kinetics and autonomic regulation of blood flow.

    Science.gov (United States)

    McNarry, M A; Kingsley, M I C; Lewis, M J

    2014-08-01

    Various regulatory mechanisms of pulmonary oxygen uptake (V̇O2) kinetics have been postulated. The purpose of this study was to investigate the relationship between vagal withdrawal, measured using RMSSDRR, the root mean square of successive differences in cardiac interval (RR) kinetics, a mediator of oxygen delivery, and V̇O2 kinetics. Forty-nine healthy adults (23 ± 3 years; 72 ± 13 kg; 1.80 ± 0.08 m) performed multiple repeat transitions to moderate- and heavy-intensity exercise. Electrocardiography, impedance cardiography, and pulmonary gas exchange parameters were measured throughout; time domain measures of heart rate variability were subsequently derived. The parameters describing the dynamic response of V̇O2, cardiac output (Q) and RMSSDRR were determined using a mono-exponential model. During heavy-intensity exercise, the phase II τ of V̇O2 was significantly correlated with the τ of RR (r = 0.36, P kinetics and those of Q, RR, or RMSSDRR during moderate exercise. Vagal withdrawal kinetics support the concept of a centrally mediated oxygen delivery limitation partly regulating V̇O2 kinetics during heavy-, but not moderate-, intensity exercise.

  2. Two corrections to the drift-wave kinetic equation in the context of zonal-flow physics

    CERN Document Server

    Ruiz, D E; Shi, E L; Dodin, I Y

    2016-01-01

    The drift-wave (DW) kinetic equation, that is commonly used in studies of zonal flows (ZF), excludes the exchange of enstrophy between DW and ZF and also effects beyond the geometrical-optics limit. Using the quasilinear approximation of the generalized Hasegawa--Mima model, we propose a modified theory that accounts for these effects within a wave kinetic equation (WKE) of the Wigner--Moyal type, which is commonly known in quantum mechanics. In the geometrical-optics limit, this theory features additional terms beyond the traditional WKE that ensure exact conservation of the \\textit{total} enstrophy and energy in the DW-ZF system. Numerical simulations are presented to illustrate the importance of these additional terms. The proposed theory can be viewed as a reformulation of the second-order cumulant expansion (also known as the CE2) in a more intuitive manner, namely, in terms of canonical phase-space variables.

  3. A stochastic asymptotic-preserving scheme for a kinetic-fluid model for disperse two-phase flows with uncertainty

    Science.gov (United States)

    Jin, Shi; Shu, Ruiwen

    2017-04-01

    In this paper we consider a kinetic-fluid model for disperse two-phase flows with uncertainty. We propose a stochastic asymptotic-preserving (s-AP) scheme in the generalized polynomial chaos stochastic Galerkin (gPC-sG) framework, which allows the efficient computation of the problem in both kinetic and hydrodynamic regimes. The s-AP property is proved by deriving the equilibrium of the gPC version of the Fokker-Planck operator. The coefficient matrices that arise in a Helmholtz equation and a Poisson equation, essential ingredients of the algorithms, are proved to be positive definite under reasonable and mild assumptions. The computation of the gPC version of a translation operator that arises in the inversion of the Fokker-Planck operator is accelerated by a spectrally accurate splitting method. Numerical examples illustrate the s-AP property and the efficiency of the gPC-sG method in various asymptotic regimes.

  4. A stochastic asymptotic-preserving scheme for a kinetic-fluid model for disperse two-phase flows with uncertainty

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Shi, E-mail: sjin@wisc.edu [Department of Mathematics, University of Wisconsin–Madison, Madison, WI 53706 (United States); Institute of Natural Sciences, School of Mathematical Science, MOELSEC and SHL-MAC, Shanghai Jiao Tong University, Shanghai 200240 (China); Shu, Ruiwen, E-mail: rshu2@math.wisc.edu [Department of Mathematics, University of Wisconsin–Madison, Madison, WI 53706 (United States)

    2017-04-15

    In this paper we consider a kinetic-fluid model for disperse two-phase flows with uncertainty. We propose a stochastic asymptotic-preserving (s-AP) scheme in the generalized polynomial chaos stochastic Galerkin (gPC-sG) framework, which allows the efficient computation of the problem in both kinetic and hydrodynamic regimes. The s-AP property is proved by deriving the equilibrium of the gPC version of the Fokker–Planck operator. The coefficient matrices that arise in a Helmholtz equation and a Poisson equation, essential ingredients of the algorithms, are proved to be positive definite under reasonable and mild assumptions. The computation of the gPC version of a translation operator that arises in the inversion of the Fokker–Planck operator is accelerated by a spectrally accurate splitting method. Numerical examples illustrate the s-AP property and the efficiency of the gPC-sG method in various asymptotic regimes.

  5. Effect of leaching-driven flow on the alteration kinetics of an ideal crack in SON68 glass

    Energy Technology Data Exchange (ETDEWEB)

    Chomat, Laure [CEA, DEN, DPC, SECR, Laboratoire d' Etude du Comportement des Betons et des Argiles, F-91191 Gif-sur-Yvette (France); Bouyer, Frederic, E-mail: frederic.bouyer@cea.fr [CEA, DEN, DTCD/SECM/LCLT, Marcoule, F-30207 Bagnols-sur-Ceze (France); Gin, Stephane [CEA, DEN, DTCD/SECM/LCLT, Marcoule, F-30207 Bagnols-sur-Ceze (France); Roux, Stephane [LMT-Cachan, ENS de Cachan/CNRS UMR 8535/Universite Paris 6/PRES UniverSud Paris, 61 Avenue du President-Wilson, 94235 Cachan Cedex (France)

    2012-07-15

    The kinetics of glass alteration is studied in a reactor cell containing parallel glass plates with a small gap, representing an ideal crack geometry. The calibrated aperture of the crack is varied from a few tens to a few hundred micrometers. The glass coupons are immersed in a highly alkaline solution to obtain fast alteration. Although the cell is closed and isothermal, it is observed that vertical or horizontal configurations lead to very different alteration gel thickness profiles. This effect results from gravity-driven flow taking place in the vertical geometry due to glass leaching as opposed to essentially diffusive transport in the horizontal case. The advection velocity is estimated from a one-dimensional model of the coupled reaction/transport mechanism, based on a zero or first order reaction. The coupling between reaction and transport has a clear influence on the apparent alteration kinetics, a phenomenon generally ignored.

  6. The effect of water temperature and flow on respiration in barnacles: patterns of mass transfer versus kinetic limitation.

    Science.gov (United States)

    Nishizaki, Michael T; Carrington, Emily

    2014-06-15

    In aquatic systems, physiological processes such as respiration, photosynthesis and calcification are potentially limited by the exchange of dissolved materials between organisms and their environment. The nature and extent of physiological limitation is, therefore, likely to be dependent on environmental conditions. Here, we assessed the metabolic sensitivity of barnacles under a range of water temperatures and velocities, two factors that influence their distribution. Respiration rates increased in response to changes in temperature and flow, with an interaction where flow had less influence on respiration at low temperatures, and a much larger effect at high temperatures. Model analysis suggested that respiration is mass transfer limited under conditions of low velocity (mass transfer and kinetic limitation are important. Behavioral monitoring revealed that barnacles fully extend their cirral appendages at low flows and display abbreviated 'testing' behaviors at high flows, suggesting some form of mechanical limitation. In low flow-high temperature treatments, however, barnacles displayed distinct 'pumping' behaviors that may serve to increase ventilation. Our results suggest that in slow-moving waters, respiration may become mass transfer limited as temperatures rise, whereas faster flows may serve to ameliorate the effects of elevated temperatures. Moreover, these results underscore the necessity for approaches that evaluate the combined effects of multiple environmental factors when examining physiological and behavioral performance.

  7. Dispersion in oscillatory electro-osmotic flow through a parallel-plate channel with kinetic sorptive exchange at walls

    Institute of Scientific and Technical Information of China (English)

    吴朝安

    2014-01-01

    Dispersion in time-oscillatory electro-osmotic flows in a slit micro-channel under the effect of kinetic sorptive exchange at walls is theoretically investigated using the homogenization method. The two walls of the channel are considered to be made up of different materials, and therefore have different zeta potentials and sorption coefficients. A general expression for the Taylor disper-sion coefficient under different zeta potentials as well as various sorption conditions at the walls is derived analytically. The disper-sion coefficient is found to be dependent on the oscillation frequency, the Debye parameter, the species partition coefficient, the rea-ction kinetics and the ratio of the wall potentials. The results demonstrate that the presence of wall sorption tends to enhance the dispersion when the oscillation frequency is low, but the effect is negligible in high-frequency oscillatory flows. Moreover, it is found that the dispersion coefficient could be significantly changed by adjusting the relative wall potentials for low-frequency flows.

  8. Multi-GPU unsteady 2D flow simulation coupled with a state-to-state chemical kinetics

    Science.gov (United States)

    Tuttafesta, Michele; Pascazio, Giuseppe; Colonna, Gianpiero

    2016-10-01

    In this work we are presenting a GPU version of a CFD code for high enthalpy reacting flow, using the state-to-state approach. In supersonic and hypersonic flows, thermal and chemical non-equilibrium is one of the fundamental aspects that must be taken into account for the accurate characterization of the plasma and state-to-state kinetics is the most accurate approach used for this kind of problems. This model consists in writing a continuity equation for the population of each vibrational level of the molecules in the mixture, determining at the same time the species densities and the distribution of the population in internal levels. An explicit scheme is employed here to integrate the governing equations, so as to exploit the GPU structure and obtain an efficient algorithm. The best performances are obtained for reacting flows in state-to-state approach, reaching speedups of the order of 100, thanks to the use of an operator splitting scheme for the kinetics equations.

  9. Experimental study of the structure of flow regions with negative turbulent kinetic energy production in confined three-dimensional shear flows with and without buoyancy

    Science.gov (United States)

    Liberzon, A.; Lüthi, B.; Guala, M.; Kinzelbach, W.; Tsinober, A.

    2005-09-01

    Regions of negative turbulent kinetic energy (TKE) production are observed and studied in two different flows, namely in turbulent thermal Rayleigh-Bénard convection in a cubic cell, and in a mechanically agitated shear flow in absence of buoyancy, with a main focus on the small scale structure of the flow. The experimental investigation is performed using three-dimensional (3D) particle tracking velocimetry, which allows for measuring the three velocity components and the full tensor of velocity derivatives in a finite 3D volume. The capability to compute the TKE production term in its complete form P =-⟨uiuj⟩Sij is crucial due to the three dimensionality of the flows. A comparative analysis of four different flow situations is performed in regions with positive and negative TKE production with and without buoyancy effects. In both, convective shear flow and shear flow without buoyancy, negative TKE production is associated with the unusual, more pronounced alignment of the velocity vector u with the first eigenvector λ1S of the mean rate-of-strain tensor, related to the stretching eigenvalue, Λ1S, in contrast to the positive TKE production associated with the alignment with the third eigenvector (i.e., related to the negative, compressing eigenvalue). In the negative TKE production region of convective flow we find (i) increased values for mean strain, (ii) increased values of the first contribution PΛ1 in the eigenframe of the mean rate-of-strain tensor, and decreased values of the vertical contribution to the production term in a fixed frame of reference, (iii) stronger anisotropy of u, (iv) higher levels of fluctuating strain s2 and enstrophy ω2, as well as (v) higher rates of their production, -sijsjkski and ωiωjsij, compared to the respective values in positive TKE production region. In the shear flow without buoyancy, all the mentioned quantities are lower in the negative TKE production region than in the positive TKE production region. From this

  10. Kinetics and product studies of the reaction ClO + BrO using discharge-flow mass spectrometry

    Science.gov (United States)

    Friedl, Randall R.; Sander, Stanley P.

    1989-01-01

    The kinetics and product branching ratios of the reaction between ClO and BrO were studied at 1 torr pressure over the temperature range 220-400 K, using the method of discharge-flow mass spectrometry. Three product channels were identified and quantified: Br + ClOO, Br + OClO, and BrCl + O2, indicating that the reaction mechanism of ClO + BrO involves metastable intermediates. The overall reaction rate coefficient and the rate coefficients for the three channel reactions are given.

  11. Kinetics of quartz dissolution in electrolyte solutions using a hydrothermal mixed flow reactor

    Science.gov (United States)

    Dove, Patricia M.; Crerar, David A.

    1990-04-01

    A hydrothermal mixed flow reactor has been developed to study the reaction kinetics of a wide variety of mineral/solution systems. The reactor is constructed of commercially pure titanium to minimize corrosion and operates at temperatures of 25 to 300°C and pressures up to 124 bars. This system is used to measure the dissolution rates of quartz at near-neutral pH in 0.0 to 0.15 m solutions of NaCl, KCl, LiCl, MgCl 2 over a temperature range of 100 to 300°C. In all cases, small concentrations of electrolytes increase the rate, some by as much as 1.5 orders of magnitude above the values measured for deionized water. The effect is greatest for solutions of NaCl and KCl where reaction rates increase with increasing electrolyte concentrations up to 0.05 molal and become constant at higher molalities. Smaller rate increases are observed for LiCl and MgCl 2 solutions. The first-order rate equation for quartz dissolution in pure water at temperatures of 100 to 300°C is given by r H 4sio 4 = k +(a sio2)(a H 2o ) 2(1 - Q/K) for a standard system of 1 m 2 of surface area and 1 kg of solution. The addition of electrolytes to reacting solutions at near-neutral pH accelerates the rate according to a Langmuir adsorption model and has the form r H 4sio 4 = (k + + k adK me +/1 + k me +)(a sio2)(a H 2o ) 2(1 - Q/K). m me + Analysis of the data indicates that the observed rate increases are controlled by the identity and concentration of the cation where alkali cations coordinate with the surface to increase the reactivity of siloxane groups by disrupting the structure of the mineral-solution interface. The rate-limiting step for the dissolution mechanism is described by (Si - O - Si) + H 2O = (Si - O - Si · OH 2)† → 2(Si - O - H) where the intermediate species is probably the same in deionized water and electrolyte solutions, but the reaction frequency is higher in electrolyte solutions due to increases in the accessibility of water to the mineral surface structures

  12. Kinetic modeling of a high power fast-axial-flow CO2 laser with computational fluid dynamics method

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    A new computational fluid dynamics (CFD) method for the simulation of fast-axial-flow CO2 laser is developed.The model which is solved by CFD software uses a set of dynamic differential equations to describe the dynamic process in one discharge tube.The velocity,temperature,pressure and turbulence energy distributions in discharge passage are presented.There is a good agreement between the theoretical prediction and the experimental results.This result indicates that the parameters of the laser have significant effect on the flow distribution in the discharge passage.It is helpful to optimize the output of high power CO2 laser by mastering its kinetic characteristics.

  13. Enhanced reaction kinetics and reactive mixing scale dynamics in mixing fronts under shear flow for arbitrary Damk\\"ohler numbers

    CERN Document Server

    Bandopadhyay, Aditya; Méheust, Yves; Dentz, Marco

    2016-01-01

    Mixing fronts, where fluids of different chemical compositions mix with each other, are typically subjected to velocity gradients, ranging from the pore scale to the catchment scale due to permeability variations and flow line geometries. A common trait of these processes is that the mixing interface is strained by shear. Depending on the P\\'eclet number $Pe$, which represents the ratio of the characteristic diffusion time to the characteristic advection time, and the Damk\\"ohler number $Da$, which represents the ratio of the characteristic diffusion time to the characteristic reaction time, the local reaction rates can be strongly impacted by the dynamics of the mixing interface. This impact has been characterized mostly either in kinetics-limited or in mixing-limited conditions, that is, for either very low or very high $Da$. Here the coupling of shear flow and chemical reactivity is investigated for arbitrary Damk\\"ohler numbers, for a bimolecular reaction and an initial interface with separated reactants....

  14. Quantitative retinal blood flow mapping from fluorescein videoangiography using tracer kinetic modeling.

    Science.gov (United States)

    Tichauer, Kenneth M; Guthrie, Micah; Hones, Logan; Sinha, Lagnojita; St Lawrence, Keith; Kang-Mieler, Jennifer J

    2015-05-15

    Abnormal retinal blood flow (RBF) has been associated with numerous retinal pathologies, yet existing methods for measuring RBF predominantly provide only relative measures of blood flow and are unable to quantify volumetric blood flow, which could allow direct patient to patient comparison. This work presents a methodology based on linear systems theory and an image-based arterial input function to quantitatively map volumetric blood flow from standard fluorescein videoangiography data, and is therefore directly translatable to the clinic. Application of the approach to fluorescein retinal videoangiography in rats (4 control, 4 diabetic) demonstrated significantly higher RBF in 4-5 week diabetic rats as expected from the literature.

  15. Kinetic characteristics of debris flows as exemplified by field investigations and discrete element simulation of the catastrophic Jiweishan rockslide, China

    Science.gov (United States)

    Zou, Zongxing; Tang, Huiming; Xiong, Chengren; Su, Aijun; Criss, Robert E.

    2017-10-01

    The Jiweishan rockslide of June 5, 2009 in China provides an important opportunity to elucidate the kinetic characteristics of high-speed, long-runout debris flows. A 2D discrete element model whose mechanical parameters were calibrated using basic field data was used to simulate the kinetic behavior of this catastrophic landslide. The model output shows that the Jiweishan debris flow lasted about 3 min, released a gravitational potential energy of about 6 × 10^13 J with collisions and friction dissipating approximately equal amounts of energy, and had a maximum fragment velocity of 60-70 m/s, almost twice the highest velocity of the overall slide mass (35 m/s). Notable simulated characteristics include the high velocity and energy of the slide material, the preservation of the original positional order of the slide blocks, the inverse vertical grading of blocks, and the downslope sorting of the slide deposits. Field observations that verify these features include uprooted trees in the frontal collision area of the air-blast wave, downslope reduction of average clast size, and undamaged plants atop huge blocks that prove their lack of downslope tumbling. The secondary acceleration effect and force chains derived from the numerical model help explain these deposit features and the long-distance transport. Our back-analyzed frictions of the motion path in the PFC model provide a reference for analyzing and predicting the motion of similar geological hazards.

  16. Atmospheric pressure flow reactor: Gas phase chemical kinetics under tropospheric conditions without wall effects

    Science.gov (United States)

    Koontz, Steven L. (Inventor); Davis, Dennis D. (Inventor)

    1991-01-01

    A flow reactor for simulating the interaction in the troposphere is set forth. A first reactant mixed with a carrier gas is delivered from a pump and flows through a duct having louvers therein. The louvers straighten out the flow, reduce turbulence and provide laminar flow discharge from the duct. A second reactant delivered from a source through a pump is input into the flowing stream, the second reactant being diffused through a plurality of small diffusion tubes to avoid disturbing the laminar flow. The commingled first and second reactants in the carrier gas are then directed along an elongated duct where the walls are spaced away from the flow of reactants to avoid wall interference, disturbance or turbulence arising from the walls. A probe connected with a measuring device can be inserted through various sampling ports in the second duct to complete measurements of the first and second reactants and the product of their reaction at selected XYZ locations relative to the flowing system.

  17. Non-modal theory of the kinetic ion temperature gradient driven instability of plasma shear flows across the magnetic field

    Science.gov (United States)

    Mikhailenko, V. V.; Mikhailenko, V. S.; Lee, Hae June

    2016-06-01

    The temporal evolution of the kinetic ion temperature gradient driven instability and of the related anomalous transport of the ion thermal energy of plasma shear flow across the magnetic field is investigated analytically. This instability develops in a steady plasma due to the inverse ion Landau damping and has the growth rate of the order of the frequency when the ion temperature is equal to or above the electron temperature. The investigation is performed employing the non-modal methodology of the shearing modes which are the waves that have a static spatial structure in the frame of the background flow. The solution of the governing linear integral equation for the perturbed potential displays that the instability experiences the non-modal temporal evolution in the shearing flow during which the unstable perturbation becomes very different from a canonical modal form. It transforms into the non-modal structure with vanishing frequency and growth rate with time. The obtained solution of the nonlinear integral equation, which accounts for the random scattering of the angle of the ion gyro-motion due to the interaction of ions with ensemble of shearing waves, reveals similar but accelerated process of the transformations of the perturbations into the zero frequency structures. It was obtained that in the shear flow the anomalous ion thermal conductivity decays with time. It is a strictly non-modal effect, which originates from the temporal evolution of the shearing modes turbulence.

  18. Development of an electrochemically integrated SR-GIXRD flow cell to study FeCO3 formation kinetics

    Science.gov (United States)

    Burkle, D.; De Motte, R.; Taleb, W.; Kleppe, A.; Comyn, T.; Vargas, S. M.; Neville, A.; Barker, R.

    2016-10-01

    An electrochemically integrated Synchrotron Radiation-Grazing Incidence X-Ray Diffraction (SR-GIXRD) flow cell for studying corrosion product formation on carbon steel in carbon dioxide (CO2)-containing brines typical of oil and gas production has been developed. The system is capable of generating flow velocities of up to 2 m/s at temperatures in excess of 80 °C during SR-GIXRD measurements of the steel surface, enabling flow to be maintained over the course of the experiment while diffraction patterns are being collected. The design of the flow cell is presented, along with electrochemical and diffraction pattern transients collected from an initial experiment which examined the precipitation of FeCO3 onto X65 carbon steel in a CO2-saturated 3.5 wt. % NaCl brine at 80 °C and 0.1 m/s. The flow cell is used to follow the nucleation and growth kinetics of FeCO3 using SR-GIXRD linked to the simultaneous electrochemical response of the steel surface which were collected in the form of linear polarisation resistance measurements to decipher in situ corrosion rates. The results show that FeCO3 nucleation could be detected consistently and well before its inhibitive effect on the general corrosion rate of the system. In situ measurements are compared with ex situ scanning electron microscopy (SEM) observations showing the development of an FeCO3 layer on the corroding steel surface over time confirming the in situ interpretations. The results presented demonstrate that under the specific conditions evaluated, FeCO3 was the only crystalline phase to form in the system, with no crystalline precursors being apparent. The numerous capabilities of the flow cell are highlighted and presented in this paper.

  19. Numerical Simulations of Two-Phase Flow in a Self-Aerated Flotation Machine and Kinetics Modeling

    KAUST Repository

    Fayed, Hassan E.

    2015-03-30

    A new boundary condition treatment has been devised for two-phase flow numerical simulations in a self-aerated minerals flotation machine and applied to a Wemco 0.8 m3 pilot cell. Airflow rate is not specified a priori but is predicted by the simulations as well as power consumption. Time-dependent simulations of two-phase flow in flotation machines are essential to understanding flow behavior and physics in self-aerated machines such as the Wemco machines. In this paper, simulations have been conducted for three different uniform bubble sizes (db = 0.5, 0.7 and 1.0 mm) to study the effects of bubble size on air holdup and hydrodynamics in Wemco pilot cells. Moreover, a computational fluid dynamics (CFD)-based flotation model has been developed to predict the pulp recovery rate of minerals from a flotation cell for different bubble sizes, different particle sizes and particle size distribution. The model uses a first-order rate equation, where models for probabilities of collision, adhesion and stabilization and collisions frequency estimated by Zaitchik-2010 model are used for the calculation of rate constant. Spatial distributions of dissipation rate and air volume fraction (also called void fraction) determined by the two-phase simulations are the input for the flotation kinetics model. The average pulp recovery rate has been calculated locally for different uniform bubble and particle diameters. The CFD-based flotation kinetics model is also used to predict pulp recovery rate in the presence of particle size distribution. Particle number density pdf and the data generated for single particle size are used to compute the recovery rate for a specific mean particle diameter. Our computational model gives a figure of merit for the recovery rate of a flotation machine, and as such can be used to assess incremental design improvements as well as design of new machines.

  20. Kinetics of selenate sorption in soil as influenced by biotic and abiotic conditions: a stirred flow-through reactor study.

    Science.gov (United States)

    Garcia-Sanchez, L; Loffredo, N; Mounier, S; Martin-Garin, A; Coppin, F

    2014-12-01

    This study (i) quantified the kinetics of selenate sorption and (ii) measured the influence of biotic processes in soil selenate stabilisation. Stirred flow-through reactor experiments were conducted on samples of a silty clay soil (pH = 8, Eh = 240-300 mV) from Bure (France) in both non-sterile and sterile conditions. Parameters of the proposed two-site sorption model (EK), adapted from van Genuchten and Wagenet (1989), were estimated by nonlinear regression. Fast selenate sorption on type-1 sites was moderate, with an equilibrium constant of 25.5 and 39.1 L/kg for non-sterile and sterile conditions. Rate-limited sorption on type-2 sites increased with time, and was predominant for longer periods of time in non-sterile conditions. At equilibrium, it would represent over 96% of the sorbed inventory, with mean sorption times of 17 h and 191 h for non-sterile and sterile conditions. Our results showed for Bure soil that (i) selenate sorption in flowing and mildly-oxidising conditions was strongly kinetically controlled, especially in non-sterile conditions, (ii) selenate desorption was much slower than sorption, which suggests its pseudo-irreversible stabilisation, and (iii) microbial activity increased the contribution of rate-limited sorption on type-2 sites, for which it increased sorption rate by a factor 7 but also facilitated its reversibility. This work stresses the limits of the Kd approach to represent selenate sorption in flowing conditions and supports an alternative formulation like the EK model, but also points out that biotic conditions are significant sources of variability for sorption parameters.

  1. Flow: Statistics, visualization and informatics for flow cytometry

    Directory of Open Access Journals (Sweden)

    Kepler Thomas B

    2008-06-01

    Full Text Available Abstract Flow is an open source software application for clinical and experimental researchers to perform exploratory data analysis, clustering and annotation of flow cytometric data. Flow is an extensible system that offers the ease of use commonly found in commercial flow cytometry software packages and the statistical power of academic packages like the R BioConductor project.

  2. First Kinetic Reactive-Flow and Melting Calculations for Entropy Budget and Major Elements in Heterogeneous Mantle Lithologies (Invited)

    Science.gov (United States)

    Asimow, P. D.

    2009-12-01

    The consequences of source heterogeneity and reactive flow during melt transport in the mantle can be classified by scale. At the smallest spatial and longest temporal scales, we can assume complete equilibrium and use batch melting of homogenized sources or equilibrium porous flow treatments. At large enough spatial scale or short enough temporal scale to prevent any thermal or chemical interaction between heterogeneities or between melt and matrix, we can assume perfectly fractional melting and transport and apply simple melt-mixing calculations. At a somewhat smaller spatial or longer temporal scale, thermal but not chemical interactions are significant and various lithologies and channel/matrix systems must follow common pressure-temperature paths, with energy flows between them. All these cases are tractable to model with current tools, whether we are interested in the energy budget, major elements, trace elements, or isotopes. There remains, however, the very important range of scales where none of these simple theories applies because of partial chemical interaction among lithologies or along the flow path. Such disequilibrium or kinetic cases have only been modeled, in the case of mantle minerals and melts, for trace elements and isotopes, with fixed melting rates instead of complete energy budgets. In order to interpret volumes of magma production and major element basalt and residue compositions that might emerge from a heterogeneous mantle in this last range of scales, we must develop tools that can combine a kinetic formulation with a major element and energy-constrained thermodynamic calculation. The kinetics can be handled either with a chemical kinetic approach with rate constants for various net transfer and exchange reactions, or with a physical diffusion-limited approach. A physical diffusion-limited approach can be built with the following elements. At grain scale, spherical grains of an arbitrary number of solid phases can evolve zoning profiles

  3. KINETIC MODELING OF CONSTITUTIVE RELATIONS FOR PARTICLE MOTION IN LOW TO MODERATELY CONCENTRATED FLOWS

    Institute of Scientific and Technical Information of China (English)

    Guangqian WANG; Xudong FU; Xingkui WANG

    2005-01-01

    Formulating underlying mechanisms of concentrated solid-liquid flows is essential for simulation of various industrial processes and natural phenomena. A generalized constitutive model for particle motion in flows with low to moderate solids concentrations is developed. This generalized model facilitates characterization of inelastic collisions, particle-fluid interactions, and shearing effects.Moderately concentrated simple shear flows of a sand-water mixture are analyzed, and comparisons of model predictions and experimental data are in good agreement. This model exhibits sound performance in characterizing particle motion for wide ranges of concentration and shear rate, and may supply a reasonable and competent alternative to previous models developed for dilute and rapid-granular flows when applied to moderately concentrated situations. The concentration approaches zero (C → 0) asymptote is observed at a relatively high shear rate in model predictions.Assumption of low collisional dissipation of the particle phase as C → 0 is more reasonable for this observation, compared to that without the interstitial fluid effect. Accurately modeling energy dissipation is important for characterizing the stability of dilute simple shear flows of solid-liquid mixtures. Incorporating friction forces will also facilitate improvement of the applicability of this generalized model to flows at extremely high concentrations.

  4. Structure and kinetics of shear aggregation in turbulent flows. I. Early stage of aggregation.

    Science.gov (United States)

    Bäbler, Matthäus U; Moussa, Amgad S; Soos, Miroslav; Morbidelli, Massimo

    2010-08-17

    Aggregation of rigid colloidal particles leads to fractal-like structures that are characterized by a fractal dimension d(f) which is a key parameter for describing aggregation processes. This is particularly true in shear aggregation where d(f) strongly influences aggregation kinetics. Direct measurement of d(f) in the early stages of shear aggregation is however difficult, as the aggregates are small and few in number. An alternative method for determining d(f) is to use an aggregation model that when fitted to the time evolution of the cluster mass distribution allows for estimating d(f). Here, we explore three such models, two of which are based on an effective collision sphere and one which directly incorporates the permeable structure of the aggregates, and we apply them for interpreting the initial aggregate growth measured experimentally in a turbulent stirred tank reactor. For the latter, three polystyrene latexes were used that differed only in the size of the primary particles (d(p) = 420, 600, and 810 nm). It was found that all three models describe initial aggregation kinetics reasonably well using, however, substantially different values for d(f). To discriminate among the models, we therefore also studied the regrowth of preformed aggregates where d(f) was experimentally accessible. It was found that only the model that directly incorporates the permeable structure of the aggregates is able to predict correctly this second type of experiments. Applying this model to the initial aggregation kinetics, we conclude that the actual initial fractal dimension is d(f) = 2.07 +/- 0.04 as found from this model.

  5. Enhanced thermal stability and mismatch discrimination of mutation-carrying DNA duplexes and their kinetic and thermodynamic properties in microchannel laminar flow.

    Science.gov (United States)

    Nagata, Maria Portia B; Yamashita, Kenichi; Miyazaki, Masaya; Nakamura, Hiroyuki; Maeda, Hideaki

    2009-07-01

    This article reports the enhancement of thermal stability involving normal duplex and mutation-carrying DNA duplexes in microchannel laminar flow. The application of an in-house temperature-controllable microchannel-type flow cell is demonstrated for improved discrimination of mismatch base pairs such as A-G and T-G that are difficult to distinguish due to the rather small thermal destabilizations. Enhancement in thermal stability is reflected by an increased thermal melting temperature achieved in microchannel laminar flow as compared with batch reactions. To examine the kinetics and thermodynamics of duplex-coil equilibrium of DNA oligomers, denaturation-renaturation hysteresis curves were measured. The influence of microchannel laminar flow on DNA base mismatch analysis was described from the kinetic and thermodynamic perspectives. An increasing trend was observed for association rate constant as flow rate increased. In contrast, an apparent decrease in dissociation rate constant was observed with increasing flow rate. The magnitudes of the activation energies of dissociation were nearly constant for both the batch and microchannel laminar flow systems at all flow rates. In contrast, the magnitudes of activation energies of association decreased as flow rate increased. These results clearly show how microchannel laminar flow induces change in reaction rate by effecting change in activation energy. We anticipate, therefore, that this approach based on microchannel laminar flow system holds great promise for improved mismatch discrimination in DNA analyses, particularly on single-base-pair mismatch, by pronouncedly enhancing thermal stability.

  6. Flow cytometric monitoring of minimal residual diseases in patients with acute leukemia after allogeneic hemapoietic stem cell transplantation%急性白血病异基因造血干细胞移植后流式细胞术监测微量残留病的意义

    Institute of Scientific and Technical Information of China (English)

    高雁群; 孙媛; 张耀臣; 纪树荃; 陆道培; 吴彤; 王卉; 童春容; 张维婕; 王静波; 卢岳; 赵艳丽; 周葭蕤

    2012-01-01

    目的 研究急性白血病异基因造血干细胞移植(allo-HSCT)后采用流式细胞术(FCM)监测微量残留病(MRD)的意义.方法 自2007年1月至2008年1月采用FCM对102例初诊时未检测出白血病基因和染色体改变的急性白血病allo-HSCT后患者进行骨髓MRD检测(移植后1、2、3、6、12个月,部分高危患者增加检测频率),观察MRD结果与临床转归的关系,对有意义的MRD增高患者予以临床干预并采用FCM监测疗效.MRD> 0.01%为阳性.结果 ①移植后MRD持续阴性者71例,均为血液学完全缓解(CR),仅3例髓外复发,其无病生存( DFS)及总生存(0S)率分别为66.2%及90.1%.②移植后MRD阳性者27例,经过干预治疗(化疗加供者淋巴细胞输注、多种细胞因子诱导的杀伤细胞和NK细胞治疗),11例患者转阴,其DFS及OS率分别为63.6%及72.7%.另外16例血液学复发,其DFS及OS率分别为11.1%及25.0%.从MRD增高至血液学复发的中位时间为48(7~69)d.③移植后直接血液学复发者共4例,均死亡.结论 移植后采用FCM检测MRD:①MRD持续阴性组患者其DFS及OS率均明显高于MRD阳性组.②移植后出现MRD阳性的患者,通过干预性治疗,MRD再次转阴后,其DFS及OS率仍然高于持续阳性组.③移植后直接血液学复发的患者,其DFS及OS率极低,预后极差.采用FCM监测急性白血病allo-HSCT后MRD是一种敏感、特异、快速、简便的方法,可及时提示复发倾向,便于早期干预治疗,降低血液学复发风险,提高allo-HSCT后患者的DFS率.%Objective To study the significance of flow cytometric monitoring minimal residual diseases (MRD) in patients with acute leukemia (AL) after allogeneic hemapoietic stem cell transplantation (HSCT).Methods From January 2007 and January 2008 MRD were detected by flow cytometry (FCM) in 402 bone marrow (BM) in 102 AL patients without leukemic gene and chromosomal changes at first diagnosis after HSCT( 1,2,3,6,12 months after HSCT

  7. Quantification of left and right atrial kinetic energy using four-dimensional intracardiac magnetic resonance imaging flow measurements.

    Science.gov (United States)

    Arvidsson, Per M; Töger, Johannes; Heiberg, Einar; Carlsson, Marcus; Arheden, Håkan

    2013-05-15

    Kinetic energy (KE) of atrial blood has been postulated as a possible contributor to ventricular filling. Therefore, we aimed to quantify the left (LA) and right (RA) atrial blood KE using cardiac magnetic resonance (CMR). Fifteen healthy volunteers underwent CMR at 3 T, including a four-dimensional phase-contrast flow sequence. Mean LA KE was lower than RA KE (1.1 ± 0.1 vs. 1.7 ± 0.1 mJ, P energy better than nonrotational flow did. The KE increase in early diastole was higher in the LA (P < 0.001). Systolic KE correlated with the combination of atrial volume and systolic velocity of the atrioventricular plane displacement (r(2) = 0.57 for LA and r(2) = 0.64 for RA). Early diastolic KE of the LA correlated with left ventricle (LV) mass (r(2) = 0.28), however, no such correlation was found in the right heart. This suggests that LA KE increases during early ventricular diastole due to LV elastic recoil, indicating that LV filling is dependent on diastolic suction. Right ventricle (RV) relaxation does not seem to contribute to atrial KE. Instead, RA KE generated during ventricular systole may be conserved in a hydraulic "flywheel" and transferred to the RV through helical flow, which may contribute to RV filling.

  8. On a consistent high-order finite difference scheme with kinetic energy conservation for simulating turbulent reacting flows

    Science.gov (United States)

    Trisjono, Philipp; Kang, Seongwon; Pitsch, Heinz

    2016-12-01

    The main objective of this study is to present an accurate and consistent numerical framework for turbulent reacting flows based on a high-order finite difference (HOFD) scheme. It was shown previously by Desjardins et al. (2008) [4] that a centered finite difference scheme discretely conserving the kinetic energy and an upwind-biased scheme for the scalar transport can be combined into a useful scheme for turbulent reacting flows. With a high-order spatial accuracy, however, an inconsistency among discretization schemes for different conservation laws is identified, which can disturb a scalar field spuriously under non-uniform density distribution. Various theoretical and numerical analyses are performed on the sources of the unphysical error. From this, the derivative of the mass-conserving velocity and the local Péclet number are identified as the primary factors affecting the error. As a solution, an HOFD stencil for the mass conservation is reformulated into a flux-based form that can be used consistently with an upwind-biased scheme for the scalar transport. The effectiveness of the proposed formulation is verified using two-dimensional laminar flows such as a scalar transport problem and a laminar premixed flame, where unphysical oscillations in the scalar fields are removed. The applicability of the proposed scheme is demonstrated in an LES of a turbulent stratified premixed flame.

  9. Kinetic freeze-out, particle spectra, and harmonic-flow coefficients from mode-by-mode hydrodynamics

    Science.gov (United States)

    Floerchinger, Stefan; Wiedemann, Urs Achim

    2014-03-01

    The kinetic freeze-out for the hydrodynamical description of relativistic heavy-ion collisions is discussed using a background-fluctuation splitting of the hydrodynamical fields. For a single event, the particle spectrum, or its logarithm, can be written as the sum of the background part that is symmetric with respect to azimuthal rotations and longitudinal boosts and a part containing the contribution of fluctuations or deviations from the background. Using a complete orthonormal basis to characterize the initial state allows one to write the double differential harmonic-flow coefficients determined by the two-particle correlation method as matrix expressions involving the initial fluid correlations. We discuss the use of these expressions for a mode-by-mode analysis of fluctuating initial conditions in heavy-ion collisions.

  10. Determinants of kinetic energy of blood flow in the four-chambered heart in athletes and sedentary controls.

    Science.gov (United States)

    Steding-Ehrenborg, K; Arvidsson, P M; Töger, J; Rydberg, M; Heiberg, E; Carlsson, M; Arheden, H

    2016-01-01

    The kinetic energy (KE) of intracardiac blood may play an important role in cardiac function. The aims of the present study were to 1) quantify and investigate the determinants of KE, 2) compare the KE expenditure of intracardiac blood between athletes and control subjects, and 3) quantify the amount of KE inside and outside the diastolic vortex. Fourteen athletes and fourteen volunteers underwent cardiac MRI, including four-dimensional phase-contrast sequences. KE was quantified in four chambers, and energy expenditure was calculated by determining the mean KE/cardiac index. Left ventricular (LV) mass was an independent predictor of diastolic LVKE (R(2) = 0.66, P energy expenditure for intracardiac blood flow, indicating similar pumping efficiency, likely explained by the lower heart rate that cancels the higher KE per heart beat in athletes. The majority of LVKE is found within the LV diastolic vortex, in contrast to earlier findings. Copyright © 2016 the American Physiological Society.

  11. Drag force and transport property of a small cylinder in free molecule flow: A gas-kinetic theory analysis

    Science.gov (United States)

    Liu, Changran; Li, Zhigang; Wang, Hai

    2016-08-01

    Analytical expressions are derived for aerodynamic drag force on small cylinders in the free molecule flow using the gas-kinetic theory. The derivation considers the effect of intermolecular interactions between the cylinder and gas media. Two limiting collision models, specular and diffuse scattering, are investigated in two limiting cylinder orientations with respect to the drift velocity. The earlier solution of Dahneke [B. E. Dahneke, J. Aerosol Sci. 4, 147 (1973), 10.1016/0021-8502(73)90066-9] is shown to be a special case of the current expressions in the rigid-body limit of collision. Drag force expressions are obtained for cylinders that undergo Brownian rotation and for those that align with the drift velocity. The validity of the theoretical expressions is tested against experimental mobility data available for carbon nanotubes.

  12. A kinetic-theory approach for computing chemical-reaction rates in upper-atmosphere hypersonic flows.

    Science.gov (United States)

    Gallis, Michael A; Bond, Ryan B; Torczynski, John R

    2009-09-28

    Recently proposed molecular-level chemistry models that predict equilibrium and nonequilibrium reaction rates using only kinetic theory and fundamental molecular properties (i.e., no macroscopic reaction-rate information) are investigated for chemical reactions occurring in upper-atmosphere hypersonic flows. The new models are in good agreement with the measured Arrhenius rates for near-equilibrium conditions and with both measured rates and other theoretical models for far-from-equilibrium conditions. Additionally, the new models are applied to representative combustion and ionization reactions and are in good agreement with available measurements and theoretical models. Thus, molecular-level chemistry modeling provides an accurate method for predicting equilibrium and nonequilibrium chemical-reaction rates in gases.

  13. Hydrogen/Oxygen Reactions at High Pressures and Intermediate Temperatures: Flow Reactor Experiments and Kinetic Modeling

    DEFF Research Database (Denmark)

    Hashemi, Hamid; Christensen, Jakob Munkholt; Glarborg, Peter

    A series of experimental and numerical investigations into hydrogen oxidation at high pressures and intermediate temperatures has been conducted. The experiments were carried out in a high pressure laminar flow reactor at 50 bar pressure and a temperature range of 600–900 K. The equivalence ratio......, ignition occurs at the temperature of 775–800 K. In general, the present model provides a good agreement with the measurements in the flow reactor and with recent data on laminar burning velocity and ignition delay time.......A series of experimental and numerical investigations into hydrogen oxidation at high pressures and intermediate temperatures has been conducted. The experiments were carried out in a high pressure laminar flow reactor at 50 bar pressure and a temperature range of 600–900 K. The equivalence ratio......, the mechanism is used to simulate published data on ignition delay time and laminar burning velocity of hydrogen. The flow reactor results show that at reducing, stoichiometric, and oxidizing conditions, conversion starts at temperatures of 750–775 K, 800–825 K, and 800–825 K, respectively. In oxygen atmosphere...

  14. Diagnosis and identification of respirometer dynamics and sludge kinetics in continuous-flow respirometers

    NARCIS (Netherlands)

    Lukasse, L.J.S.; Keesman, K.J.; Spanjers, H.; Bloemen, M.

    2000-01-01

    This paper deals with excitation of respiration chamber dynamics in a continuous-flow respirometer with the objective of extracting additional information from its dissolved oxygen (DO) sensor readings. A measurement strategy is proposed from which it is theoretically possible to identify

  15. Kinetics of gravity-driven slug flow in partially wettable capillaries of varying cross section

    Science.gov (United States)

    Nissan, Alon; Wang, Qiuling; Wallach, Rony

    2016-11-01

    A mathematical model for slug (finite liquid volume) motion in not-fully-wettable capillary tubes with sinusoidally varying cross-sectional areas was developed. The model, based on the Navier-Stokes equation, accounts for the full viscous terms due to nonuniform geometry, the inertial term, the slug's front and rear meniscus hysteresis effect, and dependence of contact angle on flow velocity (dynamic contact angle). The model includes a velocity-dependent film that is left behind the advancing slug, reducing its mass. The model was successfully verified experimentally by recording slug movement in uniform and sinusoidal capillary tubes with a gray-scale high-speed camera. Simulation showed that tube nonuniformity has a substantial effect on slug flow pattern: in a uniform tube it is monotonic and depends mainly on the slug's momentary mass/length; an undulating tube radius results in nonmonotonic flow characteristics. The static nonzero contact angle varies locally in nonuniform tubes owing to the additional effect of wall slope. Moreover, the nonuniform cross-sectional area induces slug acceleration, deceleration, blockage, and metastable-equilibrium locations. Increasing contact angle further amplifies the geometry effect on slug propagation. The developed model provides a modified means of emulating slug flow in differently wettable porous media for intermittent inlet water supply (e.g., raindrops on the soil surface).

  16. Chemical Kinetics in the expansion flow field of a rotating detonation-wave engine

    Science.gov (United States)

    Kailasanath, Kazhikathra; Schwer, Douglas

    2014-11-01

    Rotating detonation-wave engines (RDE) are a form of continuous detonation-wave engines. They potentially provide further gains in performance than an intermittent or pulsed detonation-wave engine (PDE). The overall flow field in an idealized RDE, primarily consisting of two concentric cylinders, has been discussed in previous meetings. Because of the high pressures involved and the lack of adequate reaction mechanisms for this regime, previous simulations have typically used simplified chemistry models. However, understanding the exhaust species concentrations in propulsion devices is important for both performance considerations as well as estimating pollutant emissions. A key step towards addressing this need will be discussed in this talk. In this approach, an induction parameter model is used for simulating the detonation but a more detailed finite-chemistry model is used in the expansion flow region, where the pressures are lower and the uncertainties in the chemistry model are greatly reduced. Results show that overall radical concentrations in the exhaust flow are substantially lower than from earlier predictions with simplified models. The performance of a baseline hydrogen/air RDE increased from 4940 s to 5000 s with the expansion flow chemistry, due to recombination of radicals and more production of H2O, resulting in additional heat release.

  17. Iron oxidation kinetics and phosphate immobilization along the flow-path from groundwater into surface water.

    NARCIS (Netherlands)

    Grift, van der B.; Rozemeijer, J.C.; Griffioen, J.; Velde, van der Y.

    2014-01-01

    The retention of phosphorus in surface waters though co-precipitation of phosphate with Fe-oxyhydroxides during exfiltration of anaerobic Fe(II) rich groundwater is not well understood. We developed an experimental field set-up to study Fe(II) oxidation and 5 P immobilization along the flow-path fro

  18. Iron oxidation kinetics and phosphate immobilization along the flow-path from groundwater into surface water

    NARCIS (Netherlands)

    van der Grift, B.; Rozemeijer, J. C.; Griffioen, J.; van der Velde, Y.

    2014-01-01

    The retention of phosphorus in surface waters though co-precipitation of phosphate with Fe-oxyhydroxides during exfiltration of anaerobic Fe(II) rich groundwater is not well understood. We developed an experimental field set-up to study Fe(II) oxidation and P immobilization along the flow-path from

  19. Improved Predictions of Carbon Tetrachloride Contaminant Flow and Transport: Implementation of Kinetic Volatilization and Multicomponent NAPL Behavior

    Energy Technology Data Exchange (ETDEWEB)

    Oostrom, Martinus; Zhang, Z. F.; Freedman, Vicky L.; Tartakovsky, Guzel D.

    2008-09-29

    Carbon tetrachloride (CT) was discharged to waste sites that are included in the 200-PW-1 Operable Unit in Hanford 200 West Area. Fluor Hanford, Inc. is conducting a Comprehensive Environmental Response, Compensation, and Liability Act (CERCLA) remedial investigation/feasibility study (RI/FS) for the 200-PW-1 Operable Unit. The RI/FS process and remedial investigations for the 200-PW-1, 200 PW-3, and 200-PW-6 Operable Units are described in the Plutonium/Organic-Rich Process Condensate/Process Waste Groups Operable Unit RI/FS Work Plan. As part of this overall effort, Pacific Northwest National Laboratory (PNNL) was contracted to improve the STOMP simulator (White and Oostrom, 2006) by incorporating kinetic volatilization of nonaqueous phase liquids (NAPL) and multicomponent flow and transport. This work supports the U.S. Department of Energy's (DOE's) efforts to characterize the nature and distribution of CT in the 200 West Area and subsequently select an appropriate final remedy. Previous numerical simulation results with the STOMP simulator have overestimated the effect of soil vapor extraction (SVE) on subsurface CT, showing rapid removal of considerably more CT than has actually been recovered so far. These previous multiphase simulations modeled CT mass transfer between phases based on equilibrium partitioning. Equilibrium volatilization can overestimate volatilization because mass transfer limitations present in the field are not considered. Previous simulations were also conducted by modeling the NAPL as a single component, CT. In reality, however, the NAPL mixture disposed of at the Hanford site contained several non-volatile and nearly insoluble organic components, resulting in time-variant fluid properties as the CT component volatilized or dissolved over time. Simulation of CT removal from a DNAPL mixture using single-component DNAPL properties typically leads to an overestimation of CT removal. Other possible reasons for the discrepancy

  20. Observation by flow sup 1 H NMR and dimerization kinetics and products of reactive ortho-quinodimethanes and benzocyclobutadiene

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, D.

    1990-09-21

    The reactive o-quinodimethanes, 1,2-dimethylene-1,2-dihydronaphthalene (9) and o-xylylene (1) were observed by flow {sup 1}H NMR spectroscopy at room temperature. The {sup 1}H NMR spectrum of 9 was obtained in the absence of precursor and dimers. However, the {sup 1}H NMR spectrum of the more reactive 1, generated in a similar manner from (o-((trimethylsilyl)methyl)benzyl)trimethylammonium iodide (5.) could be obtained only in the presence of its stable (4 + 2) and (4 + 4) dimers. The dimerization kinetics of 3-methyl- (5{prime}), 3,6-dimethyl- (11), 3-isopropyl- (12), and 3,6-diisoproply-1,2-xylylene (13) in acetonitrile (CH{sub 3}CN) were studied by stopped-flow UV-visible spectroscopy. Fluoride ion induced 1,2-elimination from 2-elimination from 2-trimethylsilylbenzocyclobutenyl-1 mesylate (26) was used to generate the reactive molecule benzocyclobutadiene (1{prime}) in CD{sub 3}CN, which was observed by flow {sup 1}H NMR spectroscopy at room temperature. The {sup 1}H NMR spectrum (in CD{sub 3}CN) of 1,2-dimethylene-1,2-dihydrothiophene (1{double prime}), obtained by fluoride ion induced 1,4-elimination from 3-(trimethylammoniummethyl)-2-(trimethylsilylmethyl)thiophene iodine was observed by flow {sup 1}H NMR spectroscopy at room temperature. The dimerization rate of 1{double prime} in CH{sub 3}CN, generated in the same manner, was measured by UV-visible spectroscopy. 166 refs., 7 figs., 7 tabs.

  1. Stopped-flow kinetic studies of sphere-to-rod transitions of sodium alkyl sulfate micelles induced by hydrotropic salt.

    Science.gov (United States)

    Zhang, Jingyan; Ge, Zhishen; Jiang, Xiaoze; Hassan, P A; Liu, Shiyong

    2007-12-15

    The kinetics and mechanism of sphere-to-rod transitions of sodium alkyl sulfate micelles induced by hydrotropic salt, p-toluidine hydrochloride (PTHC), were investigated by stopped-flow with light scattering detection. Spherical sodium dodecyl sulfate (SDS) micelles transform into short ellipsoidal shapes at low salt concentrations ([PTHC]/[SDS], chi(PTHC)=0.3 and 0.4). Upon stopped-flow mixing aqueous solutions of spherical SDS micelles with PTHC, the scattered light intensity gradually increases with time. Single exponential fitting of the dynamic traces leads to characteristic relaxation time, tau(g), for the growth process from spherical to ellipsoidal micelles, and it increases with increasing SDS concentrations. This suggests that ellipsoidal micelles might be produced by successive insertion of unimers into spherical micelles, similar to the case of formation of spherical micelles as suggested by Aniansson-Wall (A-W) theory. At chi(PTHC) > or = 0.5, rod-like micelles with much higher axial ratio form. The scattered light intensity exhibits an initially abrupt increase and then levels off. The dynamic curves can be well fitted with single exponential functions, and the obtained tau(g) decreases with increasing SDS concentration. Thus, the growth from spherical to rod-like micelles might proceed via fusion of spherical micelles, in agreement with mechanism proposed by Ikeda et al. At chi(PTHC)=0.3 and 0.6, the apparent activation energies obtained from temperature dependent kinetic studies for the micellar growth are 40.4 and 3.6 kJ/mol, respectively. The large differences between activation energies for the growth from spherical to ellipsoidal micelles at low chi(PTHC) and the sphere-to-rod transition at high chi(PTHC) further indicate that they should follow different mechanisms. Moreover, the sphere-to-rod transition kinetics of sodium alkyl sulfate with varying hydrophobic chain lengths (n=10, 12, 14, and 16) are also studied. The longer the carbon chain

  2. Adherence and kinetics of biofilm formation of Staphylococcus epidermidis to different types of intraocular lenses under dynamic flow conditions.

    Science.gov (United States)

    Baillif, Stéphanie; Ecochard, René; Casoli, Emmanuelle; Freney, Jean; Burillon, Carole; Kodjikian, Laurent

    2008-01-01

    To compare the adherence and biofilm formation of Staphylococcus epidermidis under in vitro flow conditions on intraocular lenses (IOLs) made of 4 biomaterials: poly(methyl methacrylate) (PMMA), silicone, hydrophilic acrylic, and hydrophobic acrylic. Department of Ophthalmology, Croix-Rousse University Hospital and University Research Laboratory, Lyon, France. Intraocular lenses were placed in a bioreactor designed to replicate intraocular conditions. The model consisted of Tygon tubing connected to a vial. Three septa allowed the entry and elimination of the artificial aqueous humor and inoculation of the bacterial suspension. The first of 2 pumps moved the aqueous humor along the circuit; the second pump regulated the flow at which the nutritive environment was regenerated. At various times (12, 16, 24, 40, 48, 60, and 72 hours), IOLs were taken from this environment and the bound bacteria were removed and counted. The distribution of bacterial adhesion on the IOLs was modeled using polynomial Poisson regression. To test the effect of the IOL biomaterial on bacterial adhesion, likelihood ratio tests were performed. The model provided the kinetics of S epidermidis biofilm growth on IOLs. The biofilm growth on each of the 4 biomaterials occurred in 3 phases: latent, dynamic or accelerated growth, and linear growth. The extent of bacterial binding to IOLs increased from hydrophilic acrylic polymer to PMMA, hydrophobic acrylic, and silicone. The differences were statistically significant. Bacterial adhesion to and biofilm development on the IOL surface depended on the characteristics of the biomaterial.

  3. An MPI-CUDA approach for hypersonic flows with detailed state-to-state air kinetics using a GPU cluster

    Science.gov (United States)

    Bonelli, Francesco; Tuttafesta, Michele; Colonna, Gianpiero; Cutrone, Luigi; Pascazio, Giuseppe

    2017-10-01

    This paper describes the most advanced results obtained in the context of fluid dynamic simulations of high-enthalpy flows using detailed state-to-state air kinetics. Thermochemical non-equilibrium, typical of supersonic and hypersonic flows, was modeled by using both the accurate state-to-state approach and the multi-temperature model proposed by Park. The accuracy of the two thermochemical non-equilibrium models was assessed by comparing the results with experimental findings, showing better predictions provided by the state-to-state approach. To overcome the huge computational cost of the state-to-state model, a multiple-nodes GPU implementation, based on an MPI-CUDA approach, was employed and a comprehensive code performance analysis is presented. Both the pure MPI-CPU and the MPI-CUDA implementations exhibit excellent scalability performance. GPUs outperform CPUs computing especially when the state-to-state approach is employed, showing speed-ups, of the single GPU with respect to the single-core CPU, larger than 100 in both the case of one MPI process and multiple MPI process.

  4. Kinetic Theory of Thermal Transpiration and Mechanocaloric Effects: Flow of Rarefied Polyatomic Gases.

    Science.gov (United States)

    Lo, Sui-Sang

    The polyatomic gas model equations developed by Hansen and Morse were used in the linearized Wang-Chang and Uhlenbeck equation to study the thermal transpiration and mechanocaloric effects in various geometries with different boundary conditions. The phenomenological coefficients for mass and energy flows at all degrees of rarefaction are reported. It is shown, on a theoretical basis, that the thermal transpiration effect ratio has a strong dependence on f(,tr), the translational Eucken factor, and only a weak dependence on f(,tr), the total Eucken factor. The cross coefficients (,MQ) = (,QM) for all calculations but the energy flux near the wall was found negative for inverse Knudsen numbers greater than 3. Flow of polyatomic gases was studied in a long cylindrical tube with the Maxwell's diffuse scattering boundary conditions. Experimental thermal transpiration effect ratios ((DELTA)p/p(,0))/((DELTA)T/T(,0)) are nearly represented by the Hansen-Morse model for simple gases, argon, air and carbon dioxide. The sulfur dioxide data are not quantitatively given by the theory due to inadequacy of the Hansen-Morse model for strongly polar gases. The work was also extended to a cylindrical annulus where comparisons were made with the Poiseuille flow data for helium, air, hydrogen and carbon dioxide. The cross effects were also studied for arbitrary (alpha), the fraction of molecules that are diffusely reflected from the surface, for a gas confined between parallel plates and in a cylindrical tube. The comparisons of the experimental thermal transpiration data with the theoretical results yielded (alpha) values which lie between 0.8 and 1.0, as expected for the "engineering surfaces.".

  5. Kinetics of lignite pyrolysis in fixed bed and entrained flow reactors. Technical report No. 17

    Energy Technology Data Exchange (ETDEWEB)

    Scaroni, A. W.; Walker, Jr., P. L.

    1979-08-01

    A laminar flow isothermal furnace has been constructed and used to study lignite pyrolysis in nitrogen at temperatures between 700/sup 0/ and 1000/sup 0/C. Particles of a Texas lignite (Darco Seam) between 41 and 201 microns in mean diameter, are found to flow down the furnace tube with velocities approximated by the summation of the gas plug-flow velocity and particle free-fall velocities. Some particle shrinkage and density changes occur during pyrolysis. Pyrolysis rate is particle size independent and increases with increase in temperature over the range of operating conditions. Ultimate yield of volatiles in the isothermal furnace, which is calculated from the linear relationship between weight loss and change in proximate volatile matter, is 66% of the original dry-ash-free coal and is particle size independent and relatively temperature independent. Ultimate yields of volatiles from fixed beds of pulverized coal are smaller than for dispersed particles of the same size. Proximate volatile matter for the lignite is, for example, 51% of the original dry-ash-free coal. Heating rates drop from about 10,000/sup 0/C/s in the isothermal furnace to about 20/sup 0/C/s in the proximate volatile matter test. Pyrolysis rates decrease and display particle size dependency in fixed beds. This implication of physical rate control is attributed to heat transfer limitations. It is proposed that pyrolysis rate and therefore residence time of volatiles in the fixed bed are important parameters affecting the preponderance of secondary char forming reactions.Also important is the total particle external surface area in the bed. Secondary char formation is considered responsible for yields of volatiles lower than the true volatile content of the lignite as measured in the isothermal furnace.

  6. Analysis of Flow Cytometry DNA Damage Response Protein Activation Kinetics Following X-rays and High Energy Iron Nuclei Exposure

    Energy Technology Data Exchange (ETDEWEB)

    Universities Space Research Association; Chappell, Lori J.; Whalen, Mary K.; Gurai, Sheena; Ponomarev, Artem; Cucinotta, Francis A.; Pluth, Janice M.

    2010-12-15

    We developed a mathematical method to analyze flow cytometry data to describe the kinetics of {gamma}H2AX and pATF2 phosphorylations ensuing various qualities of low dose radiation in normal human fibroblast cells. Previously reported flow cytometry kinetic results for these DSB repair phospho-proteins revealed that distributions of intensity were highly skewed, severely limiting the detection of differences in the very low dose range. Distributional analysis reveals significant differences between control and low dose samples when distributions are compared using the Kolmogorov-Smirnov test. Radiation quality differences are found in the distribution shapes and when a nonlinear model is used to relate dose and time to the decay of the mean ratio of phosphoprotein intensities of irradiated samples to controls. We analyzed cell cycle phase and radiation quality dependent characteristic repair times and residual phospho-protein levels with these methods. Characteristic repair times for {gamma}H2AX were higher following Fe nuclei as compared to X-rays in G1 cells (4.5 {+-} 0.46 h vs 3.26 {+-} 0.76 h, respectively), and in S/G2 cells (5.51 {+-} 2.94 h vs 2.87 {+-} 0.45 h, respectively). The RBE in G1 cells for Fe nuclei relative to X-rays for {gamma}H2AX was 2.05 {+-} 0.61 and 5.02 {+-} 3.47, at 2 h and 24-h postirradiation, respectively. For pATF2, a saturation effect is observed with reduced expression at high doses, especially for Fe nuclei, with much slower characteristic repair times (>7 h) compared to X-rays. RBEs for pATF2 were 0.66 {+-} 0.13 and 1.66 {+-} 0.46 at 2 h and 24 h, respectively. Significant differences in {gamma}H2AX and pATF2 levels comparing irradiated samples to control were noted even at the lowest dose analyzed (0.05 Gy) using these methods of analysis. These results reveal that mathematical models can be applied to flow cytometry data to uncover important and subtle differences following exposure to various qualities of low dose radiation.

  7. Multivalued fundamental diagrams of traffic flow in the kinetic Fokker-Planck limit

    CERN Document Server

    Visconti, Giuseppe; Puppo, Gabriella; Tosin, Andrea

    2016-01-01

    Starting from interaction rules based on two levels of stochasticity we study the influence of the microscopic dynamics on the macroscopic properties of vehicular flow. In particular, we study the qualitative structure of the resulting flux-density and speed-density diagrams for different choices of the desired speeds. We are able to recover multivalued diagrams as a result of the existence of a one-parameter family of stationary distributions, whose expression is analytically found by means of a Fokker-Planck approximation of the initial Boltzmann-type model.

  8. Hydrodynamic modeling of dense gas-fluidised beds using the kinetic theory of granular flow: effect of coefficient of restitution on bed dynamics

    NARCIS (Netherlands)

    Goldschmidt, M.J.V.; Kuipers, J.A.M.; van Swaaij, Willibrordus Petrus Maria

    2000-01-01

    A two-dimensional multi-fluid Eulerian CFD model with closure laws according to the kinetic theory of granular flow has been applied to study the influence of the coefficient of restitution on the hydrodynamics of dense gas-fluidised beds. It is demonstrated that hydrodynamics of dense gas-fluidised

  9. Hydrodynamic modeling of dense gas-fluidised beds using the kinetic theory of granular flow: effect of coefficient of restitution on bed dynamics.

    NARCIS (Netherlands)

    Goldschmidt, M.J.V.; Kuipers, J.A.M.; van Swaaij, Willibrordus Petrus Maria

    2001-01-01

    A two-dimensional multi-fluid Eulerian CFD model with closure laws according to the kinetic theory of granular flow has been applied to study the influence of the coefficient of restitution on the hydrodynamics of dense gas-fluidised beds. It is demonstrated that hydrodynamics of dense gas-fluidised

  10. Hydrodynamic modeling of dense gas-fluidised beds using the kinetic theory of granular flow: effect of coefficient of restitution on bed dynamics

    NARCIS (Netherlands)

    Goldschmidt, M.J.V.; Kuipers, J.A.M.; Swaaij, van W.P.M.

    2000-01-01

    A two-dimensional multi-fluid Eulerian CFD model with closure laws according to the kinetic theory of granular flow has been applied to study the influence of the coefficient of restitution on the hydrodynamics of dense gas-fluidised beds. It is demonstrated that hydrodynamics of dense gas-fluidised

  11. Hydrodynamic modelling of dense gas-fluidised beds using the kinetic theory of granular flow: effect of coefficient of restitution on bed dynamics.

    NARCIS (Netherlands)

    Goldschmidt, M.J.V.; Kuipers, J.A.M.; Swaaij, van W.P.M.

    2001-01-01

    A two-dimensional multi-fluid Eulerian CFD model with closure laws according to the kinetic theory of granular flow has been applied to study the influence of the coefficient of restitution on the hydrodynamics of dense gas-fluidised beds. It is demonstrated that hydrodynamics of dense gas-fluidised

  12. Limits on Alpha Particle Temperature Anisotropy and Differential Flow from Kinetic Instabilities: Solar Wind Observations

    CERN Document Server

    Bourouaine, Sofiane; Chandran, Benjamin D G; Maruca, Bennett A; Kasper, Justin C

    2013-01-01

    Previous studies have shown that the observed temperature anisotropies of protons and alpha particles in the solar wind are constrained by theoretical thresholds for pressure-anisotropy-driven instabilities such as the Alfv\\'en/ion-cyclotron (A/IC) and fast-magnetosonic/whistler (FM/W) instabilities. In this letter, we use a long period of in-situ measurements provided by the {\\em Wind} spacecraft's Faraday cups to investigate the combined constraint on the alpha-proton differential flow velocity and the alpha-particle temperature anisotropy due to A/IC and FM/W instabilities. We show that the majority of the data are constrained to lie within the region of parameter space in which A/IC and FM/W waves are either stable or have extremely low growth rates. In the minority of observed cases in which the growth rate of the A/IC (FM/W) instability is comparatively large, we find relatively higher values of $T_{\\perp\\alpha}/T_{\\perp p}$ ($T_{\\parallel\\alpha}/T_{\\parallel p}$) when alpha-proton differential flow vel...

  13. Real time kinetic flow cytometry measurements of cellular parameter changes evoked by nanosecond pulsed electric field.

    Science.gov (United States)

    Orbán, Csaba; Pérez-García, Esther; Bajnok, Anna; McBean, Gethin; Toldi, Gergely; Blanco-Fernandez, Alfonso

    2016-05-01

    Nanosecond pulsed electric field (nsPEF) is a novel method to increase cell proliferation rate. The phenomenon is based on the microporation of cellular organelles and membranes. However, we have limited information on the effects of nsPEF on cell physiology. Several studies have attempted to describe the effects of this process, however no real time measurements have been conducted to date. In this study we designed a model system which allows the measurement of cellular processes before, during and after nsPEF treatment in real time. The system employs a Vabrema Mitoplicator(TM) nsPEF field generating instrument connected to a BD Accuri C6 cytometer with a silicon tube led through a peristaltic pump. This model system was applied to observe the effects of nsPEF in mammalian C6 glioblastoma (C6 glioma) and HEK-293 cell lines. Viability (using DRAQ7 dye), intracellular calcium levels (using Fluo-4 dye) and scatter characteristics were measured in a kinetic manner. Data were analyzed using the FACSKin software. The viability and morphology of the investigated cells was not altered upon nsPEF treatment. The response of HEK-293 cells to ionomycin as positive control was significantly lower in the nsPEF treated samples compared to non-treated cells. This difference was not observed in C6 cells. FSC and SSC values were not altered significantly by the nsPEF treatment. Our results indicate that this model system is capable of reliably investigating the effects of nsPEF on cellular processes in real time. © 2016 International Society for Advancement of Cytometry.

  14. Effect of External Pressure and Catheter Gauge on Flow Rate, Kinetic Energy, and Endothelial Injury During Intravenous Fluid Administration in a Rabbit Model.

    Science.gov (United States)

    Hu, Mei-Hua; Chan, Wei-Hung; Chen, Yao-Chang; Cherng, Chen-Hwan; Lin, Chih-Kung; Tsai, Chien-Sung; Chou, Yu-Ching; Huang, Go-Shine

    2016-01-01

    The effects of intravenous (IV) catheter gauge and pressurization of IV fluid (IVF) bags on fluid flow rate have been studied. However, the pressure needed to achieve a flow rate equivalent to that of a 16 gauge (G) catheter through smaller G catheters and the potential for endothelial damage from the increased kinetic energy produced by higher pressurization are unclear. Constant pressure on an IVF bag was maintained by an automatic adjustable pneumatic pressure regulator of our own design. Fluids running through 16 G, 18 G, 20 G, and 22 G catheters were assessed while using IV bag pressurization to achieve the flow rate equivalent to that of a 16 G catheter. We assessed flow rates, kinetic energy, and flow injury to rabbit inferior vena cava endothelium. By applying sufficient external constant pressure to an IVF bag, all fluids could be run through smaller (G) catheters at the flow rate in a 16 G catheter. However, the kinetic energy increased significantly as the catheter G increased. Damage to the venous endothelium was negligible or minimal/patchy cell loss. We designed a new rapid infusion system, which provides a constant pressure that compresses the fluid volume until it is free from visible residual fluid. When large-bore venous access cannot be obtained, multiple smaller catheters, external pressure, or both should be considered. However, caution should be exercised when fluid pressurized to reach a flow rate equivalent to that in a 16 G catheter is run through a smaller G catheter because of the profound increase in kinetic energy that can lead to venous endothelium injury.

  15. Characterization of performance reference compound kinetics and analyte sampling rate corrections under three flow regimes using nylon organic chemical integrative samplers.

    Science.gov (United States)

    Morrison, Shane A; Belden, Jason B

    2016-09-30

    Performance reference compounds (PRCs) can be spiked into passive samplers prior to deployment. If the dissipation kinetics of PRCs from the sampler corresponds to analyte accumulation kinetics, then PRCs can be used to estimate in-situ sampling rates, which may vary depending on environmental conditions. Under controlled laboratory conditions, the effectiveness of PRC corrections on prediction accuracy of water concentrations were evaluated using nylon organic chemical integrative samplers (NOCIS). Results from PRC calibrations suggest that PRC elimination occurs faster under higher flow conditions; however, minimal differences were observed for PRC elimination between fast flow (9.3cm/s) and slow flow (5.0cm/s) conditions. Moreover, minimal differences were observed for PRC elimination from Dowex Optipore L-493; therefore, PRC corrections did not improve results for NOCIS configurations containing Dowex Optipore L-493. Regardless, results suggest that PRC corrections were beneficial for NOCIS configurations containing Oasis HLB; however, due to differences in flow dependencies of analyte sampling rates and PRC elimination rates across the investigated flow regimes, the use of multiple PRC corrections was necessary. As such, a "Best-Fit PRC" approach was utilized for Oasis HLB corrections using caffeine-(13)C3, DIA-d5, or no correction based on the relative flow dependencies of analytes and these PRCs. Although PRC corrections reduced the variability when in-situ conditions differed from laboratory calibrations (e.g. static versus moderate flow), applying PRC corrections under similar flow conditions increases variability in estimated values.

  16. Kinetics of the initial stage of immunoagglutionation studied with the scanning flow cytometer

    CERN Document Server

    Surovtsev, Ivan V; Shvalov, Alexander N; Nekrasov, Vyacheslav M; Sivolobova, Galina F; Grazhdantseva, Antonina A; Maltsev, Valeri P; Chernyshev, Andrey V

    2008-01-01

    The use of a scanning flow cytometer (SFC) to study the evolution of monomers, dimers and higher multimers of latex particles at the initial stage of the immunoagglutination is described. The SFC can measure the light-scattering pattern (indicatrix) of an individual particle over an angular range of 10-60 deg. A comparison of the experimentally measured and theoretically calculated indicatrices allows one to discriminate different types of latex particles (i.e. monomers, dimers, etc.) and, therefore, to study the evolution of immunoagglutination process. Validity of the approach was verified by simultaneous measurements of light-scattering patterns and fluorescence from individual polymer particles. Immunoagglutination was initiated by mixing bovine serum albumin (BSA)-covered latex particles (of 1.8 um in diameter) with anti-BSA IgG. The analysis of experimental data was performed on the basis of a mathematical model of diffusion-limited immunoagglutination aggregation with a steric factor. The steric factor...

  17. Modeling the kinetics of microbial degradation of deicing chemicals in porous media under flow conditions.

    Science.gov (United States)

    Wehrer, Markus; Jaesche, Philipp; Totsche, Kai Uwe

    2012-09-01

    A quantitative knowledge of the fate of deicing chemicals in the subsurface can be provided by joint analysis of lab experiments with numerical simulation models. In the present study, published experimental data of microbial degradation of the deicing chemical propylene glycol (PG) under flow conditions in soil columns were simulated inversely to receive the parameters of degradation. We evaluated different scenarios of an advection-dispersion model including different terms for degradation, such as zero order, first order and inclusion of a growing and decaying biomass for their ability to explain the data. The general break-through behavior of propylene glycol in soil columns can be simulated well using a coupled model of solute transport and degradation with growth and decay of biomass. The susceptibility of the model to non-unique solutions was investigated using systematical forward and inverse simulations. We found that the model tends to equifinal solutions under certain conditions.

  18. Growth from Solutions: Kink dynamics, Stoichiometry, Face Kinetics and stability in turbulent flow

    Science.gov (United States)

    Chernov, A. A.; DeYoreo, J. J.; Rashkovich, L. N.; Vekilov, P. G.

    2005-01-01

    bunching if solution flows parallel to the step flow. The bunch height increases with the distance the bunch travels, i.e. with the face size. However, when the flow rate, u, increases, at u greater than approx. 1 m / s , the average step bunch height decreases as 1/u. The pheonomenon is attributed to the turbulent rather than laminar viscous boundary layer where diffusivity Dt = 0.5u(sub tau),y, i.e. increases linearly with the distance y from the solid face. Friction velocity, u(sub tau) approx. u(sup 7/8). Dramatically larger rate of the mass/heat transport within the turbulent, as compared to the laminar, viscous layer will be discussed.

  19. Flow-Induced New Channels of Energy Exchange in Multi-Scale Plasma Dynamics – Revisiting Perturbative Hybrid Kinetic-MHD Theory

    Science.gov (United States)

    Shiraishi, Junya; Miyato, Naoaki; Matsunaga, Go

    2016-01-01

    It is found that new channels of energy exchange between macro- and microscopic dynamics exist in plasmas. They are induced by macroscopic plasma flow. This finding is based on the kinetic-magnetohydrodynamic (MHD) theory, which analyses interaction between macroscopic (MHD-scale) motion and microscopic (particle-scale) dynamics. The kinetic-MHD theory is extended to include effects of macroscopic plasma flow self-consistently. The extension is realised by generalising an energy exchange term due to wave-particle resonance, denoted by δ WK. The first extension is generalisation of the particle’s Lagrangian, and the second one stems from modification to the particle distribution function due to flow. These extensions lead to a generalised expression of δ WK, which affects the MHD stability of plasmas. PMID:27160346

  20. Effect of acute hypoxia on muscle blood flow, VO₂p, and [HHb] kinetics during leg extension exercise in older men.

    Science.gov (United States)

    Zerbini, Livio; Spencer, Matthew D; Grey, Tyler M; Murias, Juan M; Kowalchuk, John M; Schena, Federico; Paterson, Donald H

    2013-07-01

    The adjustment of pulmonary oxygen uptake (VO2p), heart rate (HR), limb blood flow (LBF), and muscle deoxygenation [HHb] was examined during the transition to moderate-intensity, knee-extension exercise in six older adults (70 ± 4 years) under two conditions: normoxia (FIO₂ = 20.9 %) and hypoxia (FIO₂ = 15 %). The subjects performed repeated step transitions from an active baseline (3 W) to an absolute work rate (21 W) in both conditions. Phase 2 VO₂p, HR, LBF, and [HHb] data were fit with an exponential model. Under hypoxic conditions, no change was observed in HR kinetics, on the other hand, LBF kinetics was faster (normoxia 34 ± 3 s; hypoxia 28 ± 2), whereas the overall [HHb] adjustment (τ' = TD + τ) was slower (normoxia 28 ± 2; hypoxia 33 ± 4 s). Phase 2 VO₂p kinetics were unchanged (p < 0.05). The faster LBF kinetics and slower [HHb] kinetics reflect an improved matching between O₂ delivery and O₂ utilization at the microvascular level, preventing the phase 2 VO₂p kinetics from become slower in hypoxia. Moreover, the absolute blood flow values were higher in hypoxia (1.17 ± 0.2 L min(-1)) compared to normoxia (0.96 ± 0.2 L min(-1)) during the steady-state exercise at 21 W. These findings support the idea that, for older adults exercising at a low work rate, an increase of limb blood flow offsets the drop in arterial oxygen content (CaO₂) caused by breathing an hypoxic mixture.

  1. Collaborative Research: A Model of Partially Ionized Plasma Flows with Kinetic Treatment of Neutral Atoms and Nonthermal Ions

    Energy Technology Data Exchange (ETDEWEB)

    Pogorelov, Nikolai [Univ. of Alabama, Huntsville, AL (United States); Zhang, Ming [Florida Inst. of Technology, Melbourne, FL (United States)

    2016-07-31

    Interactions of flows of partially ionized, magnetized plasma are frequently accompanied by the presence of both thermal and non-thermal (pickup) ion components. Such interactions cannot be modeled using traditional MHD equations and require more advanced approaches to treat them. If a nonthermal component of ions is formed due to charge exchange and collisions between the thermal (core) ions and neutrals, it experiences the action of magnetic field, its distribution function is isotropized, and it soon acquires the velocity of the ambient plasma without being thermodynamically equilibrated. This situation, e. g., takes place in the outer heliosphere –- the part of interstellar space beyond the solar system whose properties are determined by the solar wind interaction with the local interstellar medium. This is also possible in laboratory, at million degrees and above, when plasma is conducting electricity far too well, which makes Ohmic heating ineffective. To attain the target temperatures one needs additional heating eventually playing a dominant role. Among such sources is a so-called neutral particle beam heating. This is a wide-spread technique (Joint European Torus and International Thermonuclear Experimental Reactor experiments) based on the injection of powerful beams of neutral atoms into ohmically preheated plasma. In this project we have investigated the energy and density separation between the thermal and nonthermal components in the solar wind and interstellar plasmas. A new model has been developed in which we solve the ideal MHD equations for mixture of all ions and the kinetic Boltzmann equation to describe the transport of neutral atoms. As a separate capability, we can treat the flow of neutral atoms in a multi-component fashion, where neutral atoms born in each thermodynamically distinct region are governed by the Euler gas dynamic equations. We also describe the behavior of pickup ions either kinetically, using the Fokker--Planck equation

  2. Collaborative Research: A Model of Partially Ionized Plasma Flows with Kinetic Treatment of Neutral Atoms and Nonthermal Ions

    Energy Technology Data Exchange (ETDEWEB)

    Pogorelov, Nikolai [Univ. of Alabama, Huntsville, AL (United States). Dept. of Space Science. Center for Space Plasma and; Zhang, Ming [Florida Inst. of Technology, Melbourne, FL (United States). Physics and Space Sciences Dept.; Borovikov, Sergey [Univ. of Alabama, Huntsville, AL (United States). Dept. of Space Science. Center for Space Plasma and Aeronomic Research; Heerikhuisen, Jacob [Univ. of Alabama, Huntsville, AL (United States). Dept. of Space Science. Center for Space Plasma and Aeronomic Research; Zank, Gary [Univ. of Alabama, Huntsville, AL (United States). Dept. of Space Science. Center for Space Plasma and Aeronomic Research; Gamayunov, Konstantin [Florida Inst. of Technology, Melbourne, FL (United States). Physics and Space Sciences Dept.; Colella, Phillip [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

    2016-07-31

    Interactions of flows of partially ionized, magnetized plasma are frequently accompanied by the presence of both thermal and non-thermal (pickup) ion components. Such interactions cannot be modeled using traditional MHD equations and require more advanced approaches to treat them. If a nonthermal component of ions is formed due to charge exchange and collisions between the thermal (core) ions and neutrals, it experiences the action of magnetic field, its distribution function is isotropized, and it soon acquires the velocity of the ambient plasma without being thermodynamically equilibrated. This situation, e. g., takes place in the outer heliosphere - the part of interstellar space beyond the solar system whose properties are determined by the solar wind interaction with the local interstellar medium. This is also possible in laboratory, at million degrees and above, when plasma is conducting electricity far too well, which makes Ohmic heating ineffective. To attain the target temperatures one needs additional heating eventually playing a dominant role. Among such sources is a so-called neutral particle beam heating. This is a wide-spread technique (Joint European Torus and International Thermonuclear Experimental Reactor experiments) based on the injection of powerful beams of neutral atoms into ohmically preheated plasma. In this project we have investigated the energy and density separation between the thermal and nonthermal components in the solar wind and interstellar plasmas. A new model has been developed in which we solve the ideal MHD equations for mixture of all ions and the kinetic Boltzmann equation to describe the transport of neutral atoms. As a separate capability, we can treat the flow of neutral atoms in a multi-component fashion, where neutral atoms born in each thermodynamically distinct regions are governed by the Euler gas dynamic equations. We also describe the behavior of pickup ions either kinetically, using the Fokker–Planck equation, or

  3. A semi-Lagrangian transport method for kinetic problems with application to dense-to-dilute polydisperse reacting spray flows

    Science.gov (United States)

    Doisneau, François; Arienti, Marco; Oefelein, Joseph C.

    2017-01-01

    For sprays, as described by a kinetic disperse phase model strongly coupled to the Navier-Stokes equations, the resolution strategy is constrained by accuracy objectives, robustness needs, and the computing architecture. In order to leverage the good properties of the Eulerian formalism, we introduce a deterministic particle-based numerical method to solve transport in physical space, which is simple to adapt to the many types of closures and moment systems. The method is inspired by the semi-Lagrangian schemes, developed for Gas Dynamics. We show how semi-Lagrangian formulations are relevant for a disperse phase far from equilibrium and where the particle-particle coupling barely influences the transport; i.e., when particle pressure is negligible. The particle behavior is indeed close to free streaming. The new method uses the assumption of parcel transport and avoids to compute fluxes and their limiters, which makes it robust. It is a deterministic resolution method so that it does not require efforts on statistical convergence, noise control, or post-processing. All couplings are done among data under the form of Eulerian fields, which allows one to use efficient algorithms and to anticipate the computational load. This makes the method both accurate and efficient in the context of parallel computing. After a complete verification of the new transport method on various academic test cases, we demonstrate the overall strategy's ability to solve a strongly-coupled liquid jet with fine spatial resolution and we apply it to the case of high-fidelity Large Eddy Simulation of a dense spray flow. A fuel spray is simulated after atomization at Diesel engine combustion chamber conditions. The large, parallel, strongly coupled computation proves the efficiency of the method for dense, polydisperse, reacting spray flows.

  4. A semi-Lagrangian transport method for kinetic problems with application to dense-to-dilute polydisperse reacting spray flows

    Energy Technology Data Exchange (ETDEWEB)

    Doisneau, François, E-mail: fdoisne@sandia.gov; Arienti, Marco, E-mail: marient@sandia.gov; Oefelein, Joseph C., E-mail: oefelei@sandia.gov

    2017-01-15

    For sprays, as described by a kinetic disperse phase model strongly coupled to the Navier–Stokes equations, the resolution strategy is constrained by accuracy objectives, robustness needs, and the computing architecture. In order to leverage the good properties of the Eulerian formalism, we introduce a deterministic particle-based numerical method to solve transport in physical space, which is simple to adapt to the many types of closures and moment systems. The method is inspired by the semi-Lagrangian schemes, developed for Gas Dynamics. We show how semi-Lagrangian formulations are relevant for a disperse phase far from equilibrium and where the particle–particle coupling barely influences the transport; i.e., when particle pressure is negligible. The particle behavior is indeed close to free streaming. The new method uses the assumption of parcel transport and avoids to compute fluxes and their limiters, which makes it robust. It is a deterministic resolution method so that it does not require efforts on statistical convergence, noise control, or post-processing. All couplings are done among data under the form of Eulerian fields, which allows one to use efficient algorithms and to anticipate the computational load. This makes the method both accurate and efficient in the context of parallel computing. After a complete verification of the new transport method on various academic test cases, we demonstrate the overall strategy's ability to solve a strongly-coupled liquid jet with fine spatial resolution and we apply it to the case of high-fidelity Large Eddy Simulation of a dense spray flow. A fuel spray is simulated after atomization at Diesel engine combustion chamber conditions. The large, parallel, strongly coupled computation proves the efficiency of the method for dense, polydisperse, reacting spray flows.

  5. Differential Responses of Post-Exercise Recovery of Leg Blood Flow and Oxygen Uptake Kinetics in HFpEF versus HFrEF.

    Science.gov (United States)

    Thompson, Richard B; Pagano, Joseph J; Mathewson, Kory W; Paterson, Ian; Dyck, Jason R; Kitzman, Dalane W; Haykowsky, Mark J

    2016-01-01

    The goals of the current study were to compare leg blood flow, oxygen extraction and oxygen uptake (VO2) after constant load sub-maximal unilateral knee extension (ULKE) exercise in patients with heart failure with reduced ejection fraction (HFrEF) compared to those with preserved ejection fraction (HFpEF). Previously, it has been shown that prolonged whole body VO2 recovery kinetics are directly related to disease severity and all-cause mortality in HFrEF patients. To date, no study has simultaneously measured muscle-specific blood flow and oxygen extraction post exercise recovery kinetics in HFrEF or HFpEF patients; therefore it is unknown if muscle VO2 recovery kinetics, and more specifically, the recovery kinetics of blood flow and oxygen extraction at the level of the muscle, differ between HF phenotypes. Ten older (68±10yrs) HFrEF (n = 5) and HFpEF (n = 5) patients performed sub-maximal (85% of maximal weight lifted during an incremental test) ULKE exercise for 4 minutes. Femoral venous blood flow and venous O2 saturation were measured continuously from the onset of end-exercise, using a novel MRI method, to determine off-kinetics (mean response times, MRT) for leg VO2 and its determinants. HFpEF and HFrEF patients had similar end-exercise leg blood flow (1.1±0.6 vs. 1.2±0.6 L/min, p>0.05), venous saturation (42±12 vs. 41±11%, p>0.05) and VO2 (0.13±0.08 vs. 0.11±0.05 L/min, p>0.05); however HFrEF had significantly delayed recovery MRT for flow (292±135sec. vs 105±63sec., p = 0.004) and VO2 (95±37sec. vs. 47±15sec., p = 0.005) compared to HFpEF. Impaired muscle VO2 recovery kinetics following ULKE exercise differentiated HFrEF from HFpEF patients and suggests distinct underlying pathology and potential therapeutic approaches in these populations.

  6. Gyrokinetic and kinetic particle-in-cell simulations of guide-field reconnection. Part I: macroscopic effects of the electron flows

    CERN Document Server

    Muñoz, P A; Kilian, P; Büchner, J; Jenko, F

    2015-01-01

    In this work, we extend a comparison between gyrokinetic (GK) and fully kinetic Particle-in-Cell (PIC) simulations of magnetic reconnection in the limit of strong guide field started by TenBarge et al. [Phys. Plasmas 21, 020708 (2014)]. By using a different set of kinetic PIC and GK simulation codes (ACRONYM and GENE, respectively), we analyze the limits of applicability of the GK approach when comparing to the force free kinetic simulations in the low guide field (bg) regime. Here we report the first part of a much more extended comparison, focusing on the macroscopic effects of the electron flows. For a low beta plasma (beta_i = 0.01), it is shown that magnetic reconnection only displays similar features between both plasma models for higher kinetic PIC guide fields (bg>30) in the secondary magnetic islands than in the region close to the X points or separatrices (bg>5). Kinetic PIC low guide field runs (53) to be negligible due to the reduced reconnection rate and fluctuation level.