Optimization and validation of a flow cytometric method for immunophenotyping peripheral blood lymphocytes from cynomolgus monkeys (Macaca fascicularis)

BACKGROUND: Guidelines published by the Food and Drug Administration and Center for Human Medicinal Products describe the need to assess immunotoxic effects in nonclinical studies that evaluate drug toxicity, including the use of immunophenotyping to measure immunotoxicity. We are not aware of previ...

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Veröffentlicht in:Veterinary clinical pathology. - 1975. - 37(2008), 1 vom: 01. März, Seite 42-8
1. Verfasser: Baker, Deanna L (VerfasserIn)
Weitere Verfasser: Finco-Kent, Deborah L, Reagan, William J, Conklyn, Maryrose J, Kawabata, Thomas T
Format: Online-Aufsatz
Sprache:English
Veröffentlicht: 2008
Zugriff auf das übergeordnete Werk:Veterinary clinical pathology
Schlagworte:Journal Article
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245 1 0 |a Optimization and validation of a flow cytometric method for immunophenotyping peripheral blood lymphocytes from cynomolgus monkeys (Macaca fascicularis) 
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500 |a Date Revised 20.10.2016 
500 |a published: Print 
500 |a Citation Status MEDLINE 
520 |a BACKGROUND: Guidelines published by the Food and Drug Administration and Center for Human Medicinal Products describe the need to assess immunotoxic effects in nonclinical studies that evaluate drug toxicity, including the use of immunophenotyping to measure immunotoxicity. We are not aware of previous studies, however, that have validated methods for immunophenotyping peripheral blood lymphocyte subsets in whole blood samples from cynomolgus monkeys 
520 |a OBJECTIVE: The purpose of this study was to optimize and validate a flow cytometric assay for immunophenotyping lymphocytes in the peripheral blood of cynomolgus monkeys 
520 |a METHODS: A series of prevalidation experiments were done to determine optimal reagents, volumes, timing, and other procedural details of the flow cytometric assay. Using the optimized method, we then determined precision, interindividual variation, laboratory-to-laboratory variability, and sample stability. Stabilized human blood was used as a positive control for staining, processing, and analysis. The percentage and number of pan-T cells (CD3+), T-helper cells (CD3+4+), T cytotoxic/suppressor cells (CD3+8+), natural killer cells (CD3-16+), and B-cells (CD3-20+) were determined in 146 male and 140 female, clinically healthy monkeys and reference intervals were calculated 
520 |a RESULTS: By doing 4-color staining with a lyse-wash method, intra- and interassay precision were <5% for all lymphocyte subsets. Variability between technicians and laboratories was minimal (CVs<3%). Samples were stable for up to 24 hours after staining and fixing 
520 |a CONCLUSIONS: The validated method is extremely robust and can be performed under good laboratory practice conditions to support nonclinical studies. Reference intervals for lymphocyte subsets were similar to those previously reported 
650 4 |a Journal Article 
700 1 |a Finco-Kent, Deborah L  |e verfasserin  |4 aut 
700 1 |a Reagan, William J  |e verfasserin  |4 aut 
700 1 |a Conklyn, Maryrose J  |e verfasserin  |4 aut 
700 1 |a Kawabata, Thomas T  |e verfasserin  |4 aut 
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