Isolation of human regulatory T lymphocytes by fluorescence-activated cell sorting

Milward, Kate ORCID: https://orcid.org/0000-0003-3019-4443, Hester, Joanna and Wood, Kathryn J. 2019. Isolation of human regulatory T lymphocytes by fluorescence-activated cell sorting. Immunological Tolerance, Vol. 1899. Methods in Molecular Biology, New York: Humana Press, pp. 43-54. (10.1007/978-1-4939-8938-6_4)

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Abstract

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.

Item Type:	Book Section
Date Type:	Published Online
Status:	Published
Schools:	Schools > Medicine
Publisher:	Humana Press
ISBN:	9781493989362
Last Modified:	07 Nov 2022 09:57
URI:	https://orca.cardiff.ac.uk/id/eprint/130742

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