MBTP 20: Immunophenotyping of the activation status of NK cells from human PBMCs using flow cytometry

Miltenyi Biotec-tested panel 20 (MBTP 20)

This application protocol describes the flow cytometric analysis of the activation status of NK cells isolated from PBMCs from whole blood of a healthy donor. Immunophenotyping of NK cells was performed on all PBMCs, directly after isolation via CD3 depletion and CD56 enrichment, and after 14 days of culture.

Protocol

Flow cytometric analysis of the activation status of NK cells within PBMCs, freshly isolated NK cells, and cultured NK cells. PBMCs, freshly isolated NK cells, and isolated NK cells cultured for 14 days were stained using the previously described antibody panel. Samples were initially gated on single cells using FSC-H/FSC-A (data not shown). Live CD45+ CD3 CD14 cells (A) were further analyzed for expression of CD56 (B) and different activation markers. Compared to NK cells within PBMCs and freshly isolated NK cells, NK cells cultured for 14 days demonstrated an increased expression of CD335 (B), CD336 (C), CD337 (D), and CD314 (F). In contrast, CD16 expression remained unchanged (E). 

Materials

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