In vitro generated NK cells are functional. The function of in vitro differentiated NK cells grown in culture (Cult) for 19 days then rested in LD IL15 was compared with the function of Rag1^−/− splenic and thymic NK cells cultured in LD IL15. Sorted NK1.1⁺ cells were stimulated with IL-12/IL-18 or target YAC-1 cells. (A) IFNγ production was measured by intracellular flow cytometry. For negative controls, cells were either not stained (gray-filled histograms) or not stimulated (black-dotted histograms). All histograms are gated on NK1.1⁺ cells. (B) Cytokines secreted were assessed by cytometric bead array. (C) CD107 degranulation on target encounter was measured by flow cytometry. Data are representative of 3 experiments.