Isolation of PBMC was performed for the determination of NK cell subsets and NK cell surface markers. Peripheral blood mononuclear cells were obtained from 18 mm of blood samples with EDTA from all SLE patients by centrifugation of density Lymphoprep 1.07 (Axis-Shield, Norway), then separated and washed.
The purity of the isolation results was confirmed by FACSMelody (BD Bioscience) by staining PerCP-conjugated anti-CD3 antibody (BioLegend, San Diego, CA) and FITC-conjugated anti-CD56 antibody (BioLegend, San Diego, CA).
The number of cells was analyzed by BD Cell Quest software. The analysis resulted in the percentage (%) of cells. According to Lin et al. [12 (link)], the population of lymphocytes was gated to confirm CD3– and CD3+ lymphocytes.
Then, the population of CD3-negative lymphocytes was gated for the next analysis of CD56 expression. The results are expressed as the percentage of isolated NK cells (CD3–CD56+). The isolated NK cells were further sorted as CD56dim and CD56bright.
The purity of the isolation results was confirmed by FACSMelody (BD Bioscience) by staining PerCP-conjugated anti-CD3 antibody (BioLegend, San Diego, CA) and FITC-conjugated anti-CD56 antibody (BioLegend, San Diego, CA).
The number of cells was analyzed by BD Cell Quest software. The analysis resulted in the percentage (%) of cells. According to Lin et al. [12 (link)], the population of lymphocytes was gated to confirm CD3– and CD3+ lymphocytes.
Then, the population of CD3-negative lymphocytes was gated for the next analysis of CD56 expression. The results are expressed as the percentage of isolated NK cells (CD3–CD56+). The isolated NK cells were further sorted as CD56dim and CD56bright.
