Anti pr h 190 sc 7208

Co-immunoprecipitations were performed as described in [17 (link)] using the anti-CHD8 antibody (A301-224A) from Bethyl Laboratories. Rabbit or mouse purified IgG (Sigma-Aldrich) were used as a control. 3% Input and precipitated proteins were separated by SDS/PAGE, and visualized by Western blotting with the indicated antibodies using ECL Plus (GE Healthcare), according to the manufacturer’s instructions. Antibodies used for western blotting were: anti-BRG1 (H88, sc-10768), anti-BAF155 (R-18, sc-9746), anti-BAF170 (E-6, sc-17838), anti-PR (H-190) (sc-7208) and anti-FOXA1 (H-120) (sc-22841), and anti-hSNF5 (C20, sc-16189) from Santa Cruz Biotechnology; anti-CHD8 (A301-224A) and anti-BAF180 (A301-590A) from Bethyl; anti-BAF250 (04–080) from Millipore; α-tubulin antibody (DM1A, T9026) from Sigma Aldrich and anti-BRM (ab15597) from Abcam.

The following antibodies were used: anti-GATA3 (HG3-31, sc-268), anti-cyclin A (C-19, sc-596), anti-cyclin E (HE12, sc-247), anti-ER (MC-20, sc-542) and anti-PR (H-190, sc-7208), from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-pSer308-GATA3 (ab61052), anti-Histone H3 (tri-methyl K27) (ab6002) and anti-Histone H3 (acetyl K9) (ab4441) from Abcam (Cambridge, MA, USA); anti-PR (Ab7), anti-actin (clone ACTN05) and anti-cyclin D1 (RB 9041-P1) from Neomarkers (Freemont, CA, USA); anti-GAPDH (D16H11), anti-phospho-PKA Substrate (RRXS*/T*)(100G7E) and anti-PKA C-α (4782) from Cell Signaling (Beverly, MA, USA); anti-EZH2 (#39933) from Active Motif (Carlsbad, CA,USA); anti-acetyl-Histone H4 (#06-866) from Millipore (Temecula, CA, USA); and anti-β-tubulin from Sigma.

ChIP assays were performed as described (Strutt and Paro, 1999 (link)) using anti-PR (H190 SC-7208, Santa Cruz,); anti-RNApol II (#2629, Cell Signaling); anti-CTCF (07-729, Merck); anti-H3K27ac (ab4729, Abcam); anti-H3K18ac (#39693, Active Motif). Quantification of chromatin immunoprecipitation was performed by real-time PCR using Roche Lightcycler (Roche). The fold enrichment of target sequence in the immunoprecipitated (IP) compared to input (Ref) fractions was calculated using the comparative Ct (the number of cycles required to reach a threshold concentration) method with the Eq. (2) ^{Ct(IP)-Ct(Ref)}. Each of these values was corrected by the human β-globin gene and referred as relative abundance over time zero. Primers sequences for target regions are available on request.