Fig. 1: Exclusion gating strategy for monocyte subset determination.

Example analysis of a raw data file provided by Center A. a Living cell selection on morphological parameters (forward scatter FCS and side scatter SSC). b Singlets selection. c Monocytes rough selection as CD45^high/SSC^int cells. d Exclusion of T cells selected as CD2⁺/SSC^low cells. e Exclusion of NK cells selected as CD56⁺/SSC^low/int cells (which overlap with ncMo CD14⁻/CD16⁺ subset on CD14/CD16 dot plot). f Exclusion of B cells selected as CD24⁺/SSC^low cells as well as both immature and mature granulocytes selected as CD24⁺/SSC^{int to high} cells. g Exclusion of residual mature granulocytes selected as CD16^high cells. h Exclusion of double-negative CD14⁻CD16⁻ cells. i Checking of the previous monocyte sub-population selection. j Separation of monocytes into CD14⁺⁺/CD16⁻ (cMo, classical), CD14⁺⁺/CD16⁺ (iMo, intermediate), and CD14⁻/CD16⁺ (ncMo, nonclassical) subsets