Anti foxp3 apc 236a e7

Peripheral blood mononuclear cells (PBMCs) were isolated from whole blood using Ficoll–Paque PLUS (GE Healthcare, Buckingham, UK) density gradient centrifugation. CD4+ T cells were purified using EasySep^® Human CD4+ T‐cell Enrichment Kit (StemCell Technologies, Meylan, France) according to the manufacturer's instructions. Cells were washed with FACS buffer (2% FBS in PBS) and then stained with anti‐CD4 Alexa Fluor 700 (RPA‐T4, BD Biosciences), anti‐CD45RA PerCP Cy5.5 (HI100, eBioscience, California, USA) and anti‐CTLA‐4 PE (BNI3, BD) in FACS buffer at 4°C for 30 min in the dark. Cells were washed and stained using anti‐FoxP3‐APC (236A/E7, eBioscience) and the eBioscience FoxP3 Staining Kit according to the manufacturer's instructions.