Cd38 bv510

To immunophenotype cell populations, PBMC were thawed, washed, split into two panels, and stained for 10 min in the dark at room temperature with antibodies against the following human surface markers: CD3 Percp.Cy5.5 (BD-Biosciences, San Jose, CA, USA), CD4, CD8-FITC, CD45RO-APC, CD197-AlexaFluor-700, CD38-BV510, HLA-DR-PE.Cy7, CD28-BV711, CD27 PE-Dazzle-594, CD57-APC, and CD107a BV421(Biolegend, San Diego, CA, USA). The cells were fixed using Perm/fix (BD-Biosciences, San Jose, CA, USA) for 20 min at 4 °C in the dark, and then stained for intracellular Ki67 (Biolegend, San Diego, CA, USA) or IFN-γ (Biolegend, San Diego, CA, USA). Raw data were quantified by FACSAria (BD-Biosciences, San Jose, CA, USA) and analyzed with FlowJo version-10.

In order to analyze NK cell subsets and their receptor expression profile, PBMCs were labeled with anti-CD3-BV785, -CD16-BV570, -CD38-BV510, -CD56-BV711, -CD57-FITC, -NKG2A-PE, -NKG2D-PE/Cy7, -NKp30-BV421, -NKp44-PE/Cy7 and -NKp46-AlexaFluor700 mAbs (all from Biolegend, San Jose, CA, USA) for 20 minutes at room temperature in the dark. The data were collected and analyzed with NovoCyte flow cytometer with NovoExpress operating system software (Agilent Technologies, USA).