Cells were stained with 7-AAD viability dye, anti-CD3 eFluor450, anti-CD4 PE-eFluor 647 (eBioscience), anti-CD4 electron coupled dye (Beckmann Coulter), anti-CD4 fluorescein isothiocyanate (FITC), anti-CD3 PE, anti-CD3 APC-Cy7, anti-CD8 PE, anti-CD8 APC-Cy7, anti-CD25 PECy7, anti-CD127 PE, and anti-CD27 FITC (BD), anti-CD45RA FITC, FOXP3 Alexa Fluor 647 (BioLegend) specific antibodies. FOXP3 intracellular staining was performed using FOXP3 staining buffers (eBioscience). Data were acquired using a fluorescence activated cell sorting (FACS)CantoII and analysed using FACSDiva software (BD) and Flojo software (Treestar).