CD1d expression by B cells is required for NKT-enhanced Ab recall responses. (A) Splenocytes from C57Bl/6 and CD1d^−/− mice were assessed for TCRβ⁺, CD1d tetramer⁺ cells (dot plots) and CD1d⁺ (histograms). Empty histograms show anti-CD1d and filled show isotype control mAb. Data are representative of several (> 20) determinations. (B) Shows CD1d status of the T- and NKT-depleted B220⁺ cells used for adoptive transfer. Data are representative of several (> 12) determinations. (C) B cells from C57Bl/6 and CD1d^−/− donors were adoptively transferred to μMT recipients before immunization with NP-KLH or NP-KLH plus α-GC 24 hours after transfer. All groups were boosted with NP-KLH on day 28 and sera collected on day 33. Each mouse was immunized and boosted with 10 μg (experiment 1) and 20 μg (experiment 2) NP-KLH. Data show endpoint IgG1 titer (mean ± SD) for 3 mice per group from 2 independent experiments. * Significant differences in titer between mice receiving cells from C57Bl/6 and CD1d^−/− donors. (D) CFSE-labeled B cells from C57Bl/6 and CD1d^−/− donors were adoptively transferred into μMT mice. After 18 hours, splenocytes were prepared and assessed for CFSE⁺ cells. Data are representative of 3 independent experiments. (E) Experiment was performed as in panel C, except that mice were immunized with NP-KLH/Alum on day 0, bled (primary bleed), and then boosted with NP-KLH on day 21 and bled again on day 26 (secondary bleed). Results from a single experiment are shown in panel E.