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Human peripheral blood mononuclear cells (PBMCs) were incubated with or without (left images) PMA and ionomycin for 6 hours. After 2 hours, brefeldin A was added. The cells were harvested, fixed, permeabilized, and intracellularly stained with Anti-IL-21 antibodies and analyzed by flow cytometry using the MACSQuant®
Analyzer. Cell surface staining was performed with CD4 antibodies and CD154 antibodies. Gating was performed according to CD4 expression and side scatter properties of the cells. Cell debris was excluded from the analysis in a fluorescence 3 (FL-3/B2) versus fluorescence 4 (FL-4/B3) dot plot.
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