Parallel detection of multiple effector functions in live T cells using a short coculture assay v1

Assays measuring T cell effector function are powerful tools for determining the antigenic specificity of T cells. Conventional flow cytometric detection of cytokine production by T cells via intracellular staining necessarily compromises cell viability. In contrast, the use of a TNFa converting enzyme (TACE) inhibitor paired with monensin enables detection of both TNFa production and degranulation (CD107a/b) in viable T cells, enabling isolation of activated T cells by MACS or FACS for downstream applications such as cell culture or transcriptomic analysis. Herein, we describe the use of this assay to perform antigen rechallenge experiments in human T cells previously sensitized to antigen ex vivo.