Generation and validation of the anti-B7-H6 mAbs 1.18 and 5.51.18. (A) Binding of mAbs 1.18 and 5.51.18 at the indicated concentrations to immobilized B7-H6-FP was measured by enzyme-linked immunosorbent assay. The mean ± standard deviation absorbance at 492 nm of duplicates is shown. n.d., not detectable. (B) Binding of 1.18, 5.51.18, mIgG1 isotype control, NKp30-FP, and a control-FP to Ba/F3 and Ba/F3-B7-H6 transductants in flow cytometry. (C) Binding of NKp30-FP and a control-FP after preincubation with 1.18, 5.51.18 anti–B7-H6 mAbs, or mIgG1 isotype control to Ba/F3-B7-H6 cells was assessed by flow cytometry. (D) B7-H6 surface expression was analyzed on Ramos, K562, HeLa, SK-Mel-37, and HepG2 tumor cell lines; PBMCs and foreskin keratinocytes from healthy donors by the mAb 1.18 in flow cytometry. Representative histograms from 1 out of at least 3 experiments are shown.