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Histone H3 Antibody (MA3066) in WB
Western blot analysis of pan-acetyl Histone H4 (upper panel) was performed by loading 10 µg of acid extracted HeLa nuclear lysate extracted from cells not treated (1), treated with sodium butyrate (2) and treated with anacardic acid (3) in reducing sample buffer (Prod # 39000) and Page Ruler Plus Protein Ladder (Prod # 26619) onto a Novex 4-20% Tris-Glycine polyacrylamide gel (Prod # WT4201BX10). Proteins were transferred to nitrocellulose membrane (Prod # 88018) with Transfer Buffer (Prod # 84731) using the G2 Fast Blotter (Prod # 62288). Membrane was blocked in StartingBlock T20 (Prod # 37543) for 30 min at room temperature. Pan-acetyl Histone H4 was detected at approximately 11 kDa using a pan-acetyl Histone H4 monoclonal antibody (Prod # MA3-066) at a dilution of 1:2000 in StartingBlock T20 overnight at at 4°C on a rocking platform, followed by a goat anti-mouse superclonal IgG-HRP secondary antibody (Prod # A28177) at a dilution of 1:5000 for one hour. Chemiluminescent detection performed using SuperSignal West Pico (Prod # 34078) and myECL Imager (Prod # 62236). A loading control (lower panel) was performed by stripping the blot in Restore™ PLUS (Prod #46430) and reprobed with Histone H3 Abfinity™ oligoclonal antibody (Prod # 711055) at a dilution of 1:2000 in StartingBlock T20 overnight at at 4°C on a rocking platform, followed by a goat anti-rabbit superclonal IgG-HRP secondary antibody (Prod # A27036) at a dilution of 1:5000 for one hour.
