Expression of HLA-C2 does not influence the overall frequency of KIR2DS1⁺ NK cells. (A) Gating strategy to identify KIR2DS1- and KIR2DL1-expressing NK cells. CD56^dim NK cells were defined within CD3⁻ single live lymphocytes in an anti-CD56 versus anti-KIR2DL1 plot. KIR2DS1⁺KIR2DL1⁻, KIR2DS1⁺KIR2DL1⁺, and KIR2DS1⁻KIR2DL1⁺ NK cells were identified based on binding of anti-KIR2DL1 and anti-KIR2DL1/S1 mAbs. (B) Bulk NK cells and sorted KIR2DS1⁻KIR2DL1⁻, KIR2DS1⁺KIR2DL1⁻, KIR2DS1⁺KIR2DL1⁺, and KIR2DS1⁻KIR2DL1⁺ NK cells were analyzed for expression of KIR2DL1 and KIR2DS1 by real-time PCR. Expression relative to KIR2DL1⁻KIR2DS1⁻ NK cells is shown. (C) The frequency of KIR2DS1⁺ NK cells among CD56^dim NK cells in HLA-C1 homozygous (C1/C1), HLA-C1/C2 heterozygous (C1/C2), and HLA-C2 homozygous donors (C2/C2). (D) The frequency of KIR2DL1⁺ NK cells among CD56^dim NK cells in HLA-C1 homozygous, HLA-C1/C2 heterozygous, and HLA-C2 homozygous donors. Horizontal bars represent the mean frequency of KIR2DS1⁺ and KIR2DL1⁺ NK cells. All graphs depict data on a log10 scale.