The parental murine B-cell lymphoma NS0 is deficient for cell surface CD1d expression and does not stimulate NKT cells. (A) NS0-V and NS0-CD1 cells were stained for mouse CD1d (or K^d) and analyzed by flow cytometry. Open histogram: isotype control; filled histogram: anti-mouse CD1d or K^d. (B) NKT cells kill only CD1d-expressing NS0 cells. Splenic NKT cells were used as effectors against ⁵¹Cr-labeled NS0-V (■) or NS0-CD1 (□) target cells in a 24-hour ⁵¹Cr release assay. (C) NKT cells are activated only by CD1d-expressing NS0 cells. NS0-V or NS0-CD1 cells were cocultured with the NKT-cell hybridoma DN32.D3 for 48 hours. The supernatant was tested for IL-2 production as an indicator of NKT-cell activation. Each bar represents triplicate determinations, and the data are shown as mean plus SD. The data shown are representative of 3 independent experiments.