Analysis of Intracellular Ca(2+) Mobilization in Human NK Cell Subsets by Flow Cytometry

In signaling cascades downstream of NK cell activating receptor engagement, Ca(2+) ions are pivotal second messengers for NK cell cytotoxicity as well as cytokine production. Upon cellular activation, intracellular mobilization of Ca(2+) ions initially involves depletion of endoplasmic reticulum stores, leading to subsequent Ca(2+) influx through specific plasma membrane Ca(2+) release activated Ca(2+) channels. Multiple probes and assays for detecting intracellular Ca(2+) concentrations have been developed. With the advance of multiparameter flow cytometry instrumentation, a thorough analysis of signaling in specific NK cell subsets is possible. Here, a flow cytometric method for dynamic measurements of intracellular Ca(2+) concentrations in human NK cells subsets is detailed and discussed. This assay can be further adapted for specific scientific and diagnostic questions, with implications for various immunopathological conditions.