Altered class-switch recombination in IgH 3′RR-deficient mice. B cells were cultured for 4 days with LPS plus or minus cytokines at 1 × 10⁶ cells/mL and stained with anti-B220-PC5 and anti-isotype fluorescein isothiocyanate antibodies: anti-IgG₁ (A), anti-IgG_2a (B), anti-IgG_2b (C), anti-IgG₃ (D), and anti-IgA (E). Representative results from 6 IgH 3′RR-deficient mice and wild-type mice are shown. (F) Iμ-Cx hybrid transcripts (x being any constant gene after CSR) induced in stimulated splenocytes from wild-type and IgH 3′RR-deficient mice. Total RNA was isolated on day 3. RT-PCR for Iμ → γ2b, Iμ → γ3 was performed on LPS-induced splenocytes RNA; Iμ → γ1 and Iμ → ϵ on LPS plus IL4, Iμ → α on LPS plus TGF-β; and Iμ → γ2a on LPS plus interferon-γ. Fivefold dilutions of cDNA were used per assay. β-Actin amplification was used as control. (G) Germinal transcription in stimulated splenocytes from wild-type and IgH 3′RR-deficient mice. Total RNA was isolated on day 3. RT-PCR for germline μ, γ2b, γ3 transcripts was performed on LPS-induced splenocytes RNA; germline γ1 and ϵ on LPS plus IL4; and germline α on LPS plus TGF-β. Fivefold dilutions of cDNA were used per assay. β-Actin.