Western blotting was performed as previously described4 (link) with slight modifications: unless otherwise noted, protein lysates from tumor cells were extracted using 1X Cell Lysis Buffer (CST 9803) supplemented with PhosSTOP phosphatase inhibitors (Sigma) and cOmplete™, Mini EDTA-free Protease Inhibitor Cocktail (Sigma), and samples were normalized by total protein concentration measured by the Bio-Rad Protein Assay Dye Reagent (Bio-Rad 5000006) or equal numbers of nuclei, as indicated.
Primary antibodies used include cleaved-PARP (CST 9541), Bcl-xL (Abcam ab32370), RB (CST 9309), Lamin A/C (CST 4777), JunB (CST 3753), c-Jun (CST 9165), Fra-2 (CST 19967) and vinculin (Sigma V9131). Primary antibodies were used at 1:1,000, except vinculin, which was used at 1:5,000. Secondary antibodies used include Quick Western Kit IRDye 680RD (Licor 926–68100) used at 1:1000, Mouse IgG (H&L) Antibody DyLight™ 800 Conjugated (Rockland 610–145-002) used at 1:5,000, and Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 680 (Invitrogen A21109) used at 1:3,000. Secondary antibodies with different lot numbers were used and consistent results were observed. Western blot images were acquired on the Odyssey CLx Imaging System (LI-COR Biosciences) and processed using ImageStudio Lite v5.2.5 software.
Primary antibodies used include cleaved-PARP (CST 9541), Bcl-xL (Abcam ab32370), RB (CST 9309), Lamin A/C (CST 4777), JunB (CST 3753), c-Jun (CST 9165), Fra-2 (CST 19967) and vinculin (Sigma V9131). Primary antibodies were used at 1:1,000, except vinculin, which was used at 1:5,000. Secondary antibodies used include Quick Western Kit IRDye 680RD (Licor 926–68100) used at 1:1000, Mouse IgG (H&L) Antibody DyLight™ 800 Conjugated (Rockland 610–145-002) used at 1:5,000, and Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 680 (Invitrogen A21109) used at 1:3,000. Secondary antibodies with different lot numbers were used and consistent results were observed. Western blot images were acquired on the Odyssey CLx Imaging System (LI-COR Biosciences) and processed using ImageStudio Lite v5.2.5 software.
