NKC^KD NK cells display normal IFN-γ production and degranulation responses. (A) Splenocytes from WT or NKC^KD mice pretreated with poly(I:C) were incubated with the indicated tumor cells, on plates coated with anti-NKp46 mAb or with phorbol 12-myristate 13-acetate (PMA)/ionomycin. After 5 hours intracellular staining was performed to assess IFN-γ production by flow cytometry. (B) The frequency of IFN-γ⁺ NK cells was assessed in the spleens of the indicated mouse strains after 36 hours of infection with Smith strain murine CMV (MCMV). (C) The same assay as in panel A was performed, but splenocytes were additionally stained with anti-Ly49 and NKG2A mAb. Data are shown as the percentage of IFN-γ⁺ cells among Ly49/NKG2⁺ and Ly49/NKG2⁻ subsets. (D) CD107a levels on the surface of NK cells were evaluated by flow cytometry after the indicated tumor or mAb stimulations. Mice were pretreated with poly(I:C) or with a PBS control. Data are representative of ≥ 3 similar experiments, except for degranulation assays which were performed twice. Three individual mice were used as a source of splenocytes for each experiment (n = 3). These experiments were performed with mice on the B6 background.