Western Blotting of Protein Markers

Cell lysates were prepared using RIPA lysis buffer supplemented with protease and phosphatase inhibitors (Complete Protease Inhibitor Cocktail, Roche, Basel, Switzerland and Halt Protease and Phosphatase Inhibitor Thermo Scientific, Rockford, IL, USA) as previously described [29 (link)]. Primary antibodies used included AR-V7 (31-1109-00, RevMab Biosciences, San Francisco, CA, USA), γH2A.X (05-636, Merck Millipore, Darmstadt, Germany), and β-Actin (ab6276, Abcam, Cambridge, UK). As a secondary antibody, a horseradish peroxidase-conjugated antibody (ab6789, Abcam) was used. Signals were detected using ECL Western Blot Substrate or SuperSignal West Dura Extended Duration Substrate (Thermo Scientific, Rockford, IL, USA) on a Fusion S imaging system (Vilber Lourmat, Radolfzell, Germany).