Regulatory T cells (Tregs) are a population of lymphocytes that exerts suppressive effects upon the immune system. In human peripheral blood, the major population of T lymphocytes with suppressive capacity are defined by expression of the T cell co-receptor CD4 and the interleukin-2 receptor α-chain (CD25), combined with minimal expression of the interleukin-7 receptor α subunit (CD127). We begin by outlining the method for isolating peripheral blood mononuclear cells (PBMCs) from human blood by centrifugation of whole blood overlayed on a hydrophilic polysaccharide, with an additional erythrocyte lysis step. The protocol that follows utilizes Fluorescence-Activated Cell Sorting (FACS) for the isolation of this CD4+CD25+CD127lo population of regulatory T cells, with high yield and purity, from immunostained PBMCs. Prior to FACS isolation, this protocol exploits magnetic immunoselection for pre-enrichment of CD25+ PBMC, which reduces the duration of the subsequent FACS isolation.
Journal article
Methods Mol Biol
2019
1899
43 - 54
Cell isolation, FACS, Fluorescence-activated cell sorting, Regulatory T cells, Treg, CD4 Antigens, Cell Separation, Flow Cytometry, Humans, Interleukin-2 Receptor alpha Subunit, Interleukin-7 Receptor alpha Subunit, Leukocytes, Mononuclear, Lymphocyte Count, T-Lymphocytes, Regulatory