Histone H3 acetylation and Stat1 binding at the T-bet enhancer are impaired in Ets-1^−/− B cells. (A) ChIP analysis of histone H3 acetylation. B cells were purified from the spleens of Ets-1^−/− (KO) or wild-type (WT) chimeras and either stimulated for 1 hour with 25 μg/mL of LPS and 50 ng/mL of IFN-γ (LPS + IFN-γ) or left untreated (Med). ChIP was performed using either normal rabbit serum (Ig) or anti-acetylated histone H3 (H3) Abs. Quantification of immunoprecipitated DNA fragments was performed by real-time PCR using primers for the IgH intronic enhancer Eμ (striped), CNS I promoter region (light gray), the CNS IV enhancer (dark gray), and the irrelevant Foxp3 gene (white). Bar graphs show the values normalized to the corresponding input control and are expressed as the fold enrichment relative to normal rabbit serum. Results are representative of 3 individual experiments. (B) ChIP analysis of Stat1 binding. B cells were treated as in panel A and ChIP was performed using either normal rabbit serum (Ig) or ant-Stat1 (Stat1) Abs. Quantification of immunoprecipitated DNA fragments was performed by real-time PCR with primers for the CNS IV enhancer (dark gray) or the irrelevant Foxp3 gene (white). Bar graphs show the values normalized to corresponding input control and are expressed as the fold enrichment relative to normal rabbit serum. Results are representative of 3 individual experiments.