Flow Cytometry
For phenotypic characterization, immunostaining of 200 μl of fresh peripheral whole blood with following fluorescently labelled monoclonal antibodies, anti-CD3 (Clone:SK7), anti-CD4 (Clone: RPA-T4), anti-CD8 (Clone: SK1), anti-CD25 (Clone: M-A251), anti-CD127 (Clone: HIL-7R-M21), anti-CCR7 (Clone: 150503), anti-CD45RA (Clone: HI100), anti-CD14 (Clone: M5E2 ) and anti-16 (Clone: 3G8) was performed using stain/lyse/wash protocol as described earlier (11). Data acquisition was performed on BD FACS Aria Fusion (SORP) flow cytometer (BD Biosciences) where at least 100,000 events in lymphocyte scatter gate were acquired. Data analysis was carried out using FlowJo software. IL-6 determination was carried out using a cytometric bead array (CBA, BD Biosciences) to quantify cytokine levels in plasma of study participants as per manufacturers’ protocol.