Resting KIR2DS1⁺ NK cells lacking expression of inhibitory KIRs and CD94/NKG2A are hyporesponsive to stimulation with MHC class I–negative target cells. (A) Gating strategy to identify expression of KIR2DS1, KIR2DL1, KIR2DL3, KIR3DL1, KIR3DL2, and NKG2A on CD56^dim NK cells simultaneously by flow cytometry. CD56^dim NK cells were identified within single live CD3⁻ lymphocytes based on CD56 expression. (B) Representative example of degranulation by NK cells from a HLA-C2 homozygous donor, measured as CD107a cell-surface expression after stimulation with K562 cells. Degranulation by NKG2A⁻KIR⁻ NK cells, NKG2A⁻KIR2DS1sp NK cells, NKG2A⁻KIR2DL1sp NK cells, and NKG2A⁻KIR2DL3sp NK cells is shown. (C) Summary of NK-cell degranulation measured as CD107a cell-surface expression after stimulation with K562 cells within NKG2A⁻KIR⁻, NKG2A⁻KIR2DS1sp, NKG2A⁻KIR2DL1sp, and NKG2A⁻KIR2DL3sp NK cells. In each graph, HLA-C1 homozygous (C1), HLA-C1/C2 heterozygous (C1/C2), and HLA-C2 homozygous (C2/C2) donors are compared. Horizontal bars represent the mean frequency of CD107a⁺ NK cells within each subset. P values indicate a statistically significant difference between the groups, tested by a nonparametric Kruskal-Wallis test with Dunn posttest.