An optimized multi-parameter flow cytometry protocol for human T regulatory cell analysis on fresh and viably frozen cells, correlation with epigenetic analysis, and comparison of cord and adult blood

L. Nettenstrom, K. Alderson, E. E. Raschke, M. D. Evans, P. M. Sondel, S. Olek, C. M. Seroogy

Research output: Contribution to journalArticlepeer-review

25 Scopus citations

Abstract

Multi-parameter flow cytometry analysis of T regulatory (Treg) cells is a widely used approach in basic and translational research studies. This approach has been complicated by a lack of specific markers for Treg cells and lack of uniformity in the quantification of Treg cells. Given the central role of Treg cells in the inception and perpetuation of diverse immune responses as well as its target as a therapeutic, it is imperative to have established methodologies for Treg cell analysis that are robust and usable for studies with multiple subjects as well as multicenter studies. In this study, we describe an optimized multi-parameter flow cytometry protocol for the quantification of human Treg cells from freshly obtained and viably frozen samples and correlations with epigenetic Treg cell analysis (TSDR demethylation). We apply these two methodologies to characterize Treg cell differences between cord blood and adult peripheral blood. In summary, the optimized protocol appears to be robust for Treg cell quantification from freshly isolated or viably frozen cells and the multi-parameter flow cytometry findings are strongly positively correlated with TSDR demethylation thus providing several options for the characterization of Treg cell frequency and function in large translational or clinical studies.

Original languageEnglish (US)
Pages (from-to)81-88
Number of pages8
JournalJournal of Immunological Methods
Volume387
Issue number1-2
DOIs
StatePublished - Jan 31 2013
Externally publishedYes

Keywords

  • Cord blood
  • Demethylation
  • Epigenetics
  • Flow cytometry
  • Foxp3
  • T regulatory cell

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