Photostimulation of KIR2DL2 does not block ongoing Ca²⁺ responses. Fluo-4AM–loaded NKL cells expressing KIR2DL2 were imaged and UV irradiated on bilayers containing the indicated proteins. (A) Time-lapse montages (∼4-min intervals) showing Fluo-4AM responses before and after UV irradiation. Fluo-4AM fluorescence is proportional to intracellular Ca²⁺ concentration. (B) Ca²⁺ responses of two individual cells, which are indicated by asterisks in A. (C) Mean calcium responses for the entire population of cells. Each curve was derived from ≥30 cells. Error bars show SEM. In B and C, shaded purple bars denote UV irradiation. (D) Antibody cross-linking of KIR2DL2 does not inhibit ongoing Ca²⁺ responses. Fluo-4AM–loaded NKL cells expressing either wild-type KIR2DL2 or KIR2DL2(mut) were incubated with the indicated antibodies and subjected to flow cytometry. (top) SA (to cross-link α-NK and α-2B4) and α-Mouse (to cross-link α-KIR with α-NK and α-2B4) were added simultaneously as indicated. (bottom) α-Mouse was added after SA as indicated. All data are representative of at least two independent experiments. Norm., normalized. Bars, 10 µm.