Quantifying the effects of IL-12 and IL-2 on human NK cell responses to HLA class I–deficient cells. Purified NK cells were tested by ELISPOT assay for secretion of IFN-γ (A) and granzyme B (B) in a 20- to 24-hour response to the HLA class I–deficient B-cell line 221. Comparison was made between assays performed in the absence of cytokines (▪) with those made in the presence of different concentrations of IL-2 (▦) and IL-12 (▧). Cytokine concentrations are in units per milliliter for IL-2 and nanograms per milliliter for IL-12. Data are presented as the mean number of spots counted in duplicate wells and are representative of at least 5 independent experiments. Using single concentrations of cytokine (IL-12 at 10 ng/mL, IL-2 at 250 U/mL), similar assays compared NK cell secretion of IFN-γ (C) and granzyme B (D) in response to 221 transfectants expressing HLA-Cw*0304 (▪) with that in response to untransfected 221 cells (▦). From the results in panels C and D the proportion of NK cells that responded to 221 but were inhibited by 221-Cw*0304 was calculated (E and F, respectively). ▪ shows the inhibition of NK cells incubated with IL-12; ▨ shows the inhibition of NK cells incubated with IL-2.