MBTP 28: Immunophenotyping of virus-specific T cells from human blood using flow cytometry 

Miltenyi Biotec-tested panel 28 (MBTP 28)

This application protocol describes the flow cytometric analysis of virus-specific T cells after enrichment of IFN-γ+ cells from human blood. IFN-γ+ cells were enriched using the CliniMACS® Cytokine Capture System (IFN-gamma) to obtain viable single cell suspensions ready for downstream applications, such as flow cytometric analysis.

Protocol

Gating strategy showing the analysis of virus-specific T cells from human blood. IFN-γ enriched cells were stained using the previously described panel. Debris was excluded using FSC-H/SSC-H gating (A). Cells were analyzed for viable cells using 7-AAD-negative gating (data not shown). Viable cells were gated on CD45 and CD3 expression to identify T cells (B, C). T helper cells and cytotoxic T cells were identified based on expression of CD4 and CD8 (D). In addition, naive, central memory (CM), effector (E), and effector memory (EM) T cells were detected within CD3+ cells by analysis of CD45RO and CD62L expression (E). Moreover, CD45RO and CD62L expression of CD3+ IFN-γ+ cells (F, I), CD3+ CD4+ IFN-γ+ cells (G, J), and CD8+ IFN-γ+ cells (H, K) was determined. 

Materials

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