Absence of IL-12 p40 and CD8α expression in ICSBP^−/− DCs generated in vitro.

(A) DCs generated in vitro were stimulated with LPS (1 μg/mL), CpG (5 μg/mL), or STAg (2 μg/mL) for 24 hours for expression of ICSBP, CD8α, and IL-12 p40 transcripts by semiquantitative RT-PCR. mRNA levels were quantified by the NIH Image software and normalized to β-actin levels (numbers in parentheses). (B) Intracellular IL-12 protein expressed in indicated DCs was detected using APC-conjugated anti–IL-12 antibody. Cells were prestained with anti-CD11c antibody. The numbers indicate the percentages of IL-12⁺ cells. (C) Supernatants from DCs generated in vitro stimulated with LPS were tested for IL-12 p40 by ELISA. (D) Allo-MLR was performed with CD11c⁺ DCs as a stimulator and BALB/c spleen cells as a responder. Values represent ³H-TdR incorporation during the final 8 hours of reaction. Control (⋄) denotes MLR by ICSBP^+/+ fresh BM mononuclear cells. Values are shown as means ± SDs.