scholarly journals Rapid Quantification of NETs In Vitro and in Whole Blood Samples by Imaging Flow Cytometry

2019 ◽  
Vol 95 (5) ◽  
pp. 565-578 ◽  
Author(s):  
Patrick M. Lelliott ◽  
Masatoshi Momota ◽  
Michelle S.J. Lee ◽  
Etsushi Kuroda ◽  
Norifumi Iijima ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Whole Blood ◽  
Blood Samples ◽  
Imaging Flow Cytometry ◽  
Whole Blood Samples ◽  
Rapid Quantification

Blood Volumes of Dairy Calves Comparing Cr51-Tagged Red Blood Cells and T-1824 Plasma Dilution Methods

1958 ◽  
Vol 193 (2) ◽  
pp. 244-248 ◽  
Author(s):  
Perry Ruth Stahl ◽  
Homer E. Dale
Keyword(s):  
Body Weight ◽  
Red Blood Cells ◽  
Whole Blood ◽  
Blood Cells ◽  
Dairy Calves ◽  
Computation Method ◽  
Blood Samples ◽  
Whole Blood Samples ◽  
Blood Volumes

In a repeated study on 17 dairy calves, T-1824 dye plasma dilution showed significantly higher blood volumes than were found by any other technique or computation method using Cr51-tagged red blood cells. Five blood samples taken at 20-minute intervals after injection showed consistent decrease in radioactivity count from the first to the last sample, indicating greater accuracy in radioactivity dilution regressed to zero time figures than in average counts of several postinjection samples. In vitro studies suggest a loss of Cr51 from red blood cells to plasma after saline washings are Cr-free. Percentage blood volumes computed from whole blood samples of calves injected with Cr51-tagged red blood cells decreased in a straight line relationship with increase of body weight. Percentage plasma and whole blood volumes estimated with the T-1824 dye technique decreased regularly with body weight increase until a second determination was made when there was a rapid rise nearly to the level of the smallest calves, followed by another regular decrease with increase in weight. It is suggested that repeated dye injections do not always measure the same space. Regressed values of five whole blood samples taken at 20-minute intervals after injection of Cr51 tagged red blood cells gave more consistent blood volume determinations than either the weighed red cells or the plasma dye dilutions of the same samples.

Flow Cytometry Detection of Platelet Procoagulant Activity and Microparticles in Patients with Unstable Angina Treated by Percutaneous Coronary Angioplasty and Stent Implantation

2001 ◽  
Vol 86 (09) ◽  
pp. 784-790 ◽  
Author(s):  
Catherine Vidal ◽  
Christian Spaulding ◽  
Françoise Picard ◽  
Frédéric Schaison ◽  
Josiane Melle ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Platelet Activation ◽  
Stent Implantation ◽  
Whole Blood ◽  
Coronary Angioplasty ◽  
Annexin V ◽  
Percutaneous Coronary Angioplasty ◽  
Blood Samples ◽  
Percutaneous Coronary ◽  
Whole Blood Samples

SummaryPlatelet activation is known to participate to the pathogenesis of acute coronary syndromes. Aminophospholipid exposure and micro-particles shedding are hallmarks of full platelet activation and may account for the dissemination of prothrombotic seats. Using flow cytometry analysis of annexin V binding to externalized aminophospholipids, we followed platelet procoagulant activity (PPA) and platelet microparticles (PMP) shedding in venous and coronary whole blood samples from 30 patients with unstable angina before and after percutaneous coronary angioplasty (PTCA) and stent implantation. Baseline values of PPA and PMP were significantly more elevated in patients than in control subjects (p <0.005). PMP percentage was significantly higher in coronary than in venous blood, and in coronary blood of patients with proximal instead of mid/distal lesions of coronary arteries. No enhancement of platelet reactivity to TRAP and collagen was induced by procedure. Whereas activated GpIIb-IIIa and P-selectin expression decreased 24 h and 48 h after procedure, PPA and PMP remained as elevated as before. Thus, flow cytometry is a reliable method for detection of fully activated platelets in whole blood samples. Annexin V binding analysis demonstrates the persistance of in vivo platelet activation, despite the use of antiaggregating agents.

Long-Term Preservation of Whole Blood Samples for Flow Cytometry Analysis in Normal and HIV-Infected Individuals from Africa

1990 ◽  
Vol 1 (1) ◽  
pp. 38-45 ◽  
Author(s):  
Renu B Lal ◽  
Subhash K Hira ◽  
Rita R Dhawan ◽  
Peter L Perine
Keyword(s):  
Flow Cytometry ◽  
Whole Blood ◽  
Phenotypic Analysis ◽  
Blood Samples ◽  
Remote Areas ◽  
Immunodeficiency Virus ◽  
Whole Blood Samples ◽  
The Stability ◽  
Proliferation Assays ◽  
Lymphocyte Phenotypes

A whole blood method requiring less than 4 ml of heparinized blood was developed to assess the practicality of preparing whole blood samples that could be easily stored, transported and readily used to determine the lymphocyte phenotypes and proliferation responses of individuals from remote areas who are infected with the human immunodeficiency virus. Minor modifications in standard whole blood procedure for lymphocyte phenotyping have significantly increased the stability of light scatter and fluorescence intensity of the cells for subsequent flow cytometry (FC) analysis. These changes include removal of lysis solution prior to fixation, fixation of monoclonal antibody-stained cells in 1% paraformaldehyde for 30 minutes and storage of fixed samples in medium containing 1% bovine serum albumin. Lymphocyte subsets and their functional subsets could reliably be determined on samples stored for up to 4 weeks. Further, blood samples could be kept at room temperature for up to 96 hours or at ambient temperature during transportation from Africa before staining for FC without affecting their quantitation. While samples could be processed for FC analysis under field-laboratory conditions, proliferation assays could only be performed on samples that were transported within 48 hours of their collection. The whole blood method saves time and expense and decreases the volumes of blood required to perform phenotypic analysis and functional assays on specimens collected in remote areas.

Sensitive Detection of Survival of Paroxysmal Nocturnal Hemoglobinuria (PNH) Clones in Red Blood Cells, Granulocytes and Monocytes by High Resolution Multiparameter Flow Cytometry.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3436-3436
Author(s):  
Mayur K Movalia ◽  
Andrea Illingworth
Keyword(s):  
Flow Cytometry ◽  
High Resolution ◽  
Red Blood Cells ◽  
Whole Blood ◽  
Blood Cells ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Clone Size ◽  
Blood Samples ◽  
Whole Blood Samples ◽  
Pnh Clone

Abstract Paroxysmal Nocturnal Hemoglobinuria (PNH) is an acquired clonal stem cell disorder characterized by intravascular hemolysis due to GPI-deficient red blood cells sensitive to complement-mediated lysis. Accurate and sensitive detection of PNH-type cells has become important not only to diagnose PNH but also because studies have shown PNH-type cells may indicate favorable response to therapy and favorable prognosis in patients with aplastic anemia and myelodysplastic syndrome. Previous studies have suggested optimal testing for PNH-type cells by flow cytometry should be limited to within 48 hours after collection of whole blood. Our laboratory has developed a very sensitive and specific high resolution flow cytometric method for detecting PNH-type cells based on testing over 3,000 patients with known PNH, aplastic anemia, myelodysplastic syndromes and other bone marrow failure syndromes. The aim for this study was to determine the longevity of PNH clones in whole blood samples, the day-to-day variability of these clones and the rate of deterioration of the PNH clones compared to normal blood cells. We analyzed 10 whole blood samples from patients known to have PNH-type cells on seven consecutive days utilizing a two-color assay with GPA-CD59 for the red blood cells, a 5-color assay with FLAER-CD24-CD14-CD15-CD45 for the granulocytes and a 5 color assay with FLAER-CD33-CD14-CD64-CD45 for the monocytes. The results are summarized in the table below. The initial PNH clone sizes ranged from 0.02% to 90.8%. The PNH cells showed an overall similar level of deterioration to the normal blood cells with even minor PNH clones of 0.02% able to be detected at day 7. The day-to-day variability of PNH clone sizes was generally less than 10%, with smaller clone sizes showing a higher degree of variation, up to 20%, due to their smaller absolute numbers. Interestingly, Type III PNH red blood cells showed slightly better overall survival than normal red blood cells and were detected in modestly increasing percentages throughout the study. Based on this data, we propose that accurate detection of PNH type cells can be achieved up to seven days after collection of whole blood when utilizing high resolution flow cytometry. PNH Clone Size on Sequential Days as Percentage of Original PNH Clone Size Original PNH Clone Sizes PNH Clone Sizes as Percentage of Original PNH Clone Size Cell Type Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 Type III RBCs .02%–58.6% 102% 106% 107% 104% 108% 103% Granulocytes .29%–90.8% 100% 100% 93% 89% 79% 86% Monocytes .52%–89.9% 96% 96% 92% 94% 97% 85%

scholarly journals Measurement of Circulating Forms of Prostate-specific Antigen in Whole Blood Immediately after Venipuncture: Implications for Point-of-Care Testing

2001 ◽  
Vol 47 (4) ◽  
pp. 703-711 ◽  
Author(s):  
Timo Piironen ◽  
Martti Nurmi ◽  
Kerttu Irjala ◽  
Olli Heinonen ◽  
Hans Lilja ◽  
...  
Keyword(s):  
Physical Exercise ◽  
Prostate Specific Antigen ◽  
Whole Blood ◽  
Point Of Care ◽  
Prostate Gland ◽  
Specific Antigen ◽  
Point Of Care Testing ◽  
Blood Samples ◽  
Whole Blood Samples

Abstract Background: The purpose of this study was to validate the use of whole-blood samples in the determination of circulating forms of prostate-specific antigen (PSA). Methods: Blood samples of hospitalized prostate cancer and benign prostatic hyperplasia patients were collected and processed to generate whole-blood and serum samples. Three different rapid two-site immunoassays were developed to measure the concentrations of total PSA (PSA-T), free PSA (PSA-F), and PSA-α1-antichymotrypsin complex (PSA-ACT) to detect in vitro changes in whole-blood samples immediately after venipuncture. The possible influence of muscle movement on the release of PSA from prostate gland was studied in healthy men by measuring the rapid in vitro whole-blood kinetics of PSA forms before and after 15 min of physical exercise on a stationary bicycle. Results: Rapid PSA-T, PSA-F, and PSA-ACT assays were designed using a 10-min sample incubation. No significant changes were detected in the concentrations of PSA-T, PSA-F, and PSA-ACT from the earliest time point of 12–16 min compared with measurements performed up to 4 h after venipuncture. Physical exercise did not influence the concentrations of the circulating forms of PSA. Hematocrit-corrected whole-blood values of PSA-T and PSA-F forms were comparable to the respective serum values. Calculation of the percentage of PSA-F (PSA F/T ratio × 100) was similar irrespective of the sample format used, i.e., whole blood or serum. Conclusions: We found that immunodetectable PSA forms are likely at steady state immediately after venipuncture, thus enabling the use of anticoagulated whole-blood samples in near-patient settings for point-of-care testing, whereas determinations of PSA (e.g., PSA-T, PSA-F, or PSA-ACT) performed within the time frame of the office visit would provide results equivalent to conventional analyses performed in serum.

Applicability of flow cytometry γH2AX assay in population studies: suitability of fresh and frozen whole blood samples

2021 ◽  
Author(s):  
Blanca Laffon ◽  
María Sánchez-Flores ◽  
Natalia Fernández-Bertólez ◽  
Eduardo Pásaro ◽  
Vanessa Valdiglesias
Keyword(s):  
Flow Cytometry ◽  
Whole Blood ◽  
Population Studies ◽  
Blood Samples ◽  
Whole Blood Samples

Effect of Ascorbic Acid in Vivo on Labeling of Red Cells by Radioactive Sodium Chromate

Blood ◽  
1963 ◽  
Vol 22 (3) ◽  
pp. 351-356 ◽  
Author(s):  
J. WILLIAM POPPELL ◽  
MARGARET E. HOOD ◽  
JEROME J. DECOSSE
Keyword(s):  
Ascorbic Acid ◽  
Acid Concentration ◽  
Whole Blood ◽  
Sodium Chromate ◽  
Ascorbic Acid Concentration ◽  
Blood Samples ◽  
Radioactive Sodium ◽  
Whole Blood Samples

Abstract Fifty-two whole blood samples from 17 patients were analyzed for ascorbic acid concentration and Cr51 tagging. Physiologic concentrations of ascorbic acid in whole blood in vivo reduced radioactive sodium chromate in vitro and impaired tagging.

scholarly journals An easy and reliable whole blood freezing method for flow cytometry immunophenotyping and functional analyses

2020 ◽  
Author(s):  
Cecile Braudeau ◽  
Nina Salabert-Le Guen ◽  
Chevreuil Justine ◽  
Rimbert Marie ◽  
Jerome C. Martin ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Whole Blood ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Immune Monitoring ◽  
Fresh Blood ◽  
Peripheral Blood Mononuclear ◽  
Blood Samples ◽  
Whole Blood Samples

ABSTRACTBackgroundImmune profiling by flow cytometry is not always possible on fresh blood samples due to time and/or transport constraints. Besides, the cryopreservation of peripheral blood mononuclear cells (PBMC) requires on-site specialized lab facilities, thus severely restricting the extent by which blood immune monitoring can be applied to multicenter clinical studies. These major limitations can be addressed through the development of simplified whole blood freezing methods.MethodsIn this report, we describe an optimized easy protocol for rapid whole blood freezing with the CryoStor® CS10 solution. Using flow cytometry, we compared cellular viability and composition on cryopreserved whole blood samples to matched fresh blood, as well as fresh and frozen PBMC.ResultsThough partial loss of neutrophils was observed, leucocyte viability was routinely >75% and we verified the preservation of viable T cells, NK cells, monocytes, dendritic cells and eosinophils in frequencies similar to those observed in fresh samples. A moderate decrease in B cell frequencies was observed. Importantly, we validated the possibility to analyze major intracellular markers, such as FOXP3 and Helios in regulatory T cells. Finally, we demonstrated good functional preservation of CS10-cryopreserved cells through the analysis of intracellular cytokine production in ex vivo stimulated T cells (IFNg, IL-4, IL-17A,) and monocytes (IL-1b, IL-6, TNFa).ConclusionsIn conclusion, our protocol provides a robust method to apply reliable immune monitoring studies to cryopreserved whole blood samples, hence offering new important opportunities for the design of future multicenter clinical trials.

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