Nine-color flow cytometry for accurate measurement of T cell subsets and cytokine responses. Part II: Panel performance across different instrument platforms.
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McLaughlin, Bridget E, Baumgarth, Nicole, Bigos, Martin, Roederer, Mario, De Rosa, Stephen C, Altman, John D, Nixon, Douglas F, Ottinger, Janet, Li, Judy, Beckett, Laurel, Shacklett, Barbara L, Evans, Thomasg and Asmuth, David M (2008) Nine-color flow cytometry for accurate measurement of T cell subsets and cytokine responses. Part II: Panel performance across different instrument platforms. Cytometry. Part A : the journal of the International Society for Analytical Cytology, 73 (5). pp. 411-420. ISSN 1552-4930
Abstract
Cellular immune responses elicited by vaccination are complex and require polychromatic analysis to accurately characterize the phenotype and function of rare, responding cells. Technical challenges and a lack of instrument standardization between research sites have limited the application of polychromatic cytometry in multicenter clinical trials. Two previously developed six-color T cell subset immunophenotyping reagent panels deliberately designed to accommodate three additional low frequency functional measurements were compared for their reproducibility of staining across three different flow cytometers. We repeatedly measured similar T cell subset frequencies between the two reagent panels and across the three different cytometers. Spectral overlap reduced sensitivity in two of the three open measurement channels (PE [IL-2] and APC [IFN gamma]) for one reagent combination, particularly in subsets with low cytokine expression. There was no significant interassay variation for measurements across instrument platforms. Careful panel design will identify reagent combinations that minimize spectral spillover into channels reserved for cytokine measurement and comparable results can be achieved using different cytometers, however, it is important to establish standardized quality control procedures for each instrument to minimize variation between cytometers.
| Item Type: | Article |
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| Additional Information: | archiving not allowed on institutional repository |
| Keywords: | polychromatic flow cytometry; multicolor flow cytometry; high-resolution immunophenotyping; fluorochrome conjugated antibody; compensation |
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| Date Deposited: | 14 Dec 2009 13:52 |
| Last Modified: | 14 Dec 2009 13:52 |
| URI: | https://oak.novartis.com/id/eprint/865 |