PBMCs were isolated and cryopreserved. At the time of analysis, 5 x 105 thawed PBMCs were incubated with the following monoclonal antibodies: anti-CD3 VioBright R720, anti-CD8 APC, anti-CD4 APC-Vio770, anti-CD45RA PE-Vio615, anti-CD57 Vioblue, anti-PD1 ViobrightFITC, anti-CD49d PE770 (Miltenyi Biotech, Germany), anti-CCR7 BV785 (Biolegend, USA) and FVS510 (Fixable Viability Stain 510) (BD Biosciences, USA).
After 10 min on ice in the dark and one wash with PBSA (PBS supplemented with bovine serum albumin), the cells were acquired in an LSRFortessa flow cytometer (BD Biosciences, CA, USA). Dot plots were generated using FlowJo v10.6.2 software (Tree Star, USA).
After 10 min on ice in the dark and one wash with PBSA (PBS supplemented with bovine serum albumin), the cells were acquired in an LSRFortessa flow cytometer (BD Biosciences, CA, USA). Dot plots were generated using FlowJo v10.6.2 software (Tree Star, USA).