Isolation of viable antigen-specific CD8+ T cells based on membrane-bound tumor necrosis factor (TNF)-? expression.

ABSTRACT: Current technology to isolate viable cytokine-producing antigen-specific primary human T cells is limited to bi-specific antibody capture systems, which suffer from limited sensitivity and high background. Here, we describe a novel procedure for isolating antigen-specific human T cells based on their ability to produce tumor necrosis factor (TNF)-?. Unlike many cytokines, TNF-? is initially produced in a biologically active membrane-bound form that is subsequently cleaved by TNF-? converting enzyme (TACE) to release the soluble form of TNF-?. By preventing this cleavage event, we show that TNF-? can be 'trapped' on the surface of the T cells from which it originates and directly labeled for viable isolation of these antigen-specific T cells. Together with other existing sorting procedures to isolate activated T cells, this new technique should permit the direct isolation of multi-functional T lymphocytes for further protein and gene expression analyses, as well as a detailed functional assessment of the potential role that TNF-? producing T cells play in the adaptive immune system.

SUBMITTER: Haney D 

PROVIDER: S-EPMC3116017 | biostudies-literature | 2011 Jun

REPOSITORIES: biostudies-literature