Flow cytometric-gating strategies for cell lineages from Indian and US cord blood mononuclear cells (PBMCs).

After primary gating for CD45, various cell subsets were gated as shown and percentage values for each subset calculated. (A) First row: Gating for monocytes (CD45+CD66b-CD14+CD11c+), and among them, for patrolling monocytes (CD14dimCD16+), inflammatory monocytes (CD14+CD16+), and classical monocytes (CD14+CD16-). Second row: CD45+CD20+ B cells were further gated for B1 B cells (CD45+CD20+CD27+CD43+), and naïve B cells (CD45+CD20+CD27-). Naïve B cells were further gated as immature naïve B cells (CD45+CD20+CD27-CD19+CD10+). Third row: Gating for dendritic cells (DCs) (CD45+Lineage- [CD3-CD14-CD16-CD19-CD20-CD56-] HLA-DR+) and among them, for myeloid DCs (mDCs; CD45+lineage-HLADR+CD11c+CD123-) and plasmacytoid DCs (pDCs; CD45+lineage-HLA-DR+CD11c-CD123+). Fourth row: Gating for Monocytes (CD45+CD14+), NK (Natural Killer) cells (CD45+CD3-CD56+), B cells (CD45+CD19+) and Transitional B cells (CD45+CD19+CD24+CD38bright). (B) CD45+CD3+ cells identified as T cells. First row: Classical T cells (CD45+CD3+CD25-CD56-TCRg/d-) which were sub-divided into CD4 and CD8 cells. CD4 and CD8 T cells were further characterized using CD45RA and CCR7 as naïve (CCR7+CD45RA+) cells, central memory (CCR7+CD45RA-) and TEMRA (CD8+CCR7-CD45RA+). Second row: CD45+CD3+ T cells were further gated for putative regulatory T cells (CD45+CD3+CD4+CD25+CD127-). Third row: Gating for gamma-delta T cells (CD45+CD3+TCRg/d+), NKT cells (CD45+CD3+CD56+) and invariant NKT (iNKT) cells (CD45+ CD3+V alpha 24-J alpha 18 TCR+).