Neutralization of IL-12 on day 6 after birth nullifies IFN-γ response through up-regulation of IL-13Rα1 and apoptosis of Th1 cells. Newborn BALB/c mice were given 3 x 10⁴ CD4 neonatal DO11.10 T cells within 24 h after birth. On day 6, the hosts were given 60 μg/g Ig-OVA and 50 μg anti–IL-12 antibody per mouse or rat IgG. Another injection of anti–IL-12 antibody or rat IgG was given on days 7 and 8. A group of mice that received Ig-OVA on day 1 and no anti–IL-12 antibody at any time was included as a control. 7 wk later, all groups were challenged with 125 μg OVA peptide in CFA. 10 d later, the splenic cells (10⁶ cells/well) were stimulated with 10 μM OVA peptide and assayed for (A) IFN-γ production by ELISPOT and (B) apoptosis by Annexin V staining, as in Fig. 3. Each bar represents the mean ± SD of triplicate wells. (C and D) Mice that received neonatal DO11.10 T cells were given Ig-OVA and anti–IL-12 antibody as in A, and 2 wk later, primary Th1 cells were isolated and IL-13Rα1 expression was assessed by spot blot (C) and real-time PCR (D), as described in Fig. 3. For the spot blot in C, each bar represents the mean ± SD of duplicate spots. For the real-time PCR, the bars represent the comparative threshold cycle (C_T). The value of the sample from mice that received rat IgG was set as 1. (E and F) Newborn BALB/c from A and B that received anti–IL-12 antibody or rat IgG control during exposure to Ig-OVA on day 6 were killed 2 wk later, and the splenic cells were stained with KJ1-26 and rabbit anti–IL-13Rα1 antiserum (1:100 dilution). The Th1 cells were identified by staining for intracellular IFN-γ with anti–IFN-γ antibody, and expression of surface IL-13Rα1 was determined by flow cytometry on cells gated for KJ1-26 expression and intracellular IFN-γ production.