B7-H6 cell surface expression triggers NKp30-dependent cell activation. (A–C) DO.11.10, DOMSP30, and DOMSP46 cells were stimulated using the indicated plate-bound mAbs (A) or P815.B7-H1 versus P815.B7-H6 cells in the absence (B) or presence of indicated F(ab′)₂ mAbs or B7-H6 mouse antiserum (C). Data indicate the mean of triplicates + SD and are representative of three independent experiments. (D and E) NK-92 cells (D) and IL-2–stimulated primary NK cells (E) were used in a cytolytic assay against P815 cells (thin line) or P815.B7-H6 cells (bold line). Soluble NKp30-Fc (thin dashed line) and control Fc-fusion protein (B7-H3-Fc; bold dashed line) were used at 2 µg/ml. (F) Freshly isolated primary blood NK cells in PBMCs were tested for their activation induced by P815.B7-H1 or P815.B7-H6 cells in the presence or absence of anti-NKp30 F(ab′)₂ mAbs or control mouse Ig (mIg). 500,000 PBMCs were added to 100,000 tumor cells. The fraction of reactive NK cells (percentage of responding NK cells) was assessed by adding the percentage of CD107⁺IFN-γ⁻, CD107⁺IFN-γ⁺, and CD107⁻IFN-γ⁺ NK cells. Data are representative of three independent experiments. Error bars indicate SD. Statistical analysis was performed using a Mann-Whitney U test. *, P < 0.05.