Fitc anti human foxp3 antibody

Harvested CD4⁺ T cells were counted for Th17 and Treg percentages using flow cytometry (FACScalibur). Before detection of Th17 cells, cells were stimulated with 50 ng/ml PMA (Sigma-Aldrich) and 1 µg/ml Ionomycin (Sigma-Aldrich) in the presence of Brefeldin A (BD Pharmingen) at 37°C for 4 hours. Cells were stained with FITC anti-human CD4 antibody (Biolegend). Afterwards, cells were fixed, permeabilized, and labelled with and PerCP/Cy5.5 anti-human IL-17A (Biolegend). Th17 were cells which expressed CD4⁺ IL-17A⁺. For detection of Treg, cells were labelled with PerCP anti-human CD4 antibody (Biolegend) and PE anti-human CD25 antibody (Biolegend). FITC anti-human FoxP3 antibody (Biolegend) was added later after cells were fixed and permeabilized. Treg were cells which expressed CD4⁺ CD25⁺ FoxP3⁺.