Protocol for flow cytometry immunophenotyping of human antigen-specific T cells by activation-induced marker and Th1 cytokine detection

Flow cytometry characterization of antigen-specific polyfunctional T cells is a valuable tool to study adaptive immunity. Here, we present a protocol for flow cytometry immunophenotyping of human antigen-specific T cells by activation-induced marker and Th1 cytokine detection. We describe steps for preparing peripheral blood mononuclear cells (PBMCs) for stimulation followed by washing and staining PBMCs for flow cytometry. We then detail procedures for acquisition and analysis. This protocol has potential applications in the field of vaccine immunology and immuno-oncology. For complete details on the use and execution of this protocol, please refer to Altosole et al.1 [Display omitted] •Antigen-specific T cell detection based on intracellular staining of CD137 and CD69•Activation-induced marker evaluation in conjunction with Th1 cytokine detection•T cell maturation curves based on CD45RA and CCR7 are included in the panel•Detection of antigen-specific T follicular helper cells is supported by CXCR5 staining Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Flow cytometry characterization of antigen-specific polyfunctional T cells is a valuable tool to study adaptive immunity. Here, we present a protocol for flow cytometry immunophenotyping of human antigen-specific T cells by activation-induced marker and Th1 cytokine detection. We describe steps for preparing peripheral blood mononuclear cells (PBMCs) for stimulation followed by washing and staining PBMCs for flow cytometry. We then detail procedures for acquisition and analysis. This protocol has potential applications in the field of vaccine immunology and immuno-oncology.