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OMIP-006

Phenotypic subset analysis of human T regulatory cells via polychromatic flow cytometry

David M. Murdoch, Janet S. Staats, Kent J. Weinhold Cytometry PART A, Volume 81A, Issue 4, 281-283 (2012)

PURPOSE: This panel was optimized for the enumeration and phenotypic characterization of T regulatory cells (Tregs) within the CD4+ T-cell pool using human peripheral blood mononuclear cells (PBMC) using intranuclear and intracellular staining methods. The panel was optimized for HIV+ clinical trial specimens through the use of HIV-infected and normal donor PBMC. Because the panel is to be used in the context of testing cryopreserved PBMC obtained from multiple sites participating in clinical trials, it was essential to develop an assay that performed well using cryopreserved PBMC. Other tissue types have not been tested. © 2012 International Society for Advancement of Cytometry.

CELL TYPE: Cryopreserved Peripheral Blood Mononuclear Cells

MACHINE: BD LSR2

Marker Target Clone Fluorochrome Purpose
Dead cells Human Live/Dead Fix Violet viability
CD45RO Human UCHL1 FITC maturation/differentiation
CD4 Human SK3 PerCP-Cy5.5 T cell subset
FoxP3 Human PCH101 PE Treg identification
CD25 Human B1.49.9 ECD T cell activation, Treg identification
CD49d Human 9F10 PE-Cy5 Treg subset
CD39 Human PE-Cy7 Treg subset
Helios Human 22F6 Alexa Fluor 647 Treg subset
CD3 Human UCHT1 V500 T cell subset