NKT-cell activation by a microbial glycolipid. (A) Calcium dependence of IFN-γ and IL-4 production. NKT cells were stimulated by α-GalCer–pulsed CD1d transfectants in the presence of 111 nM CsA or of vehicle, and the indicated cytokines were measured by ELISA. The plot shows the amount of each cytokine produced in the presence of CsA as a percentage of the amount of the same cytokine induced in the absence of CsA. (B) A microbial antigen induces both IFN-γ and IL-4 secretion. NKT cells were stimulated by CD1d transfectants pulsed with 5 μg/mL BbGL, or 50 ng/mL α-GalCer, or left untreated (Autoreactive), and production of IL-4 (right y-axis) and IFN-γ (left y-axis) was quantitated by ELISA. (C) BbGL-pulsed APCs can stimulate calcium flux by NKT cells. Data are representative of 2 or 3 independent experiments using clone J24N.22 or clone J24L.17.