Freshly isolated PBMCs (1 × 105) from Omicron COVID-19 cases (n = 19) were used for surface staining of NK cells, NKT-like cells, B cells, T cells, memory B, and memory T cells using cocktail of antihuman antibodies (CD19 PercpCy5.5 (clone HB-19), CD27 PECy7 (clone M-T271), CD3 APCH7 (clone SK7), CD4 BV480 (clone SK3), CD8 FITC (clone RPA-T8), CD45RA PECy7 (clone HI 100), CD62L APC (clone DREG-56), CCR7 PE (clone 2-LI-A) and NK Tritest (CD3 FITC: clone SK7, CD56 PE:clone NCAM 16.2, CD16 PE: clone B73.1, CD 45 RA: C8Mab-1)) all from BD Biosciences following a previously described protocol [18 (link)–23 (link)]. PE-Cy™7 Mouse IgG1 κ Isotype Control (BD Biosciences, San Jose, CA, USA) was used as the negative control.
Briefly, PBMCs were incubated with fluorochrome-tagged antihuman monoclonal antibodies for 30 min in the dark. After washing, the cells were fixed with 2% paraformaldehyde. The cells were acquired on FACS Aria II (BD Biosciences, San Jose, CA, USA). For each experiment, 50,000 events were acquired within the lymphocyte gate with appropriate isotype control. Data were analyzed using FACS Diva software (Becton Dickinson, San Jose, CA, USA), and results are expressed as the percentage of positive cells in the gated population. The gating strategy is depicted inFigure 1 .
Briefly, PBMCs were incubated with fluorochrome-tagged antihuman monoclonal antibodies for 30 min in the dark. After washing, the cells were fixed with 2% paraformaldehyde. The cells were acquired on FACS Aria II (BD Biosciences, San Jose, CA, USA). For each experiment, 50,000 events were acquired within the lymphocyte gate with appropriate isotype control. Data were analyzed using FACS Diva software (Becton Dickinson, San Jose, CA, USA), and results are expressed as the percentage of positive cells in the gated population. The gating strategy is depicted in