Impaired generation of DSBs in switching Mlh1^GR/GR B cells. (A) ChIP experiments using anti-53BP1 antibodies were conducted on WT, Mlh1^−/−, and Mlh1^GR/GR splenic B cells activated with LPS + IL4 for 50 h. Specific DNA fragments corresponding to the Sμ, Sγ3, and Sγ1 regions of the IgH locus were quantitated in the immunoprecipitates by real-time PCR relative to input DNA. The bars denote the enrichment of specific DNA fragments present in the corresponding immunoprecipitates in stimulated (t = 50 h) over unstimulated (t = 0 h) cells. Error bars indicate the SD of triple PCRs (*, P < 0.05; ***, P < 0.001). (B) Western blotting analysis was performed on WT and Mlh1^GR/GR splenic B cells activated with either LPS (left) or with LPS + IL4 (right) for 96 h. Levels of γ-H2AX and anti-H2A were compared between the unstimulated (t = 0 h) and stimulated (t = 96 h) cells in the respective genotypes. A representative of three separate experiments is shown.