Parp clone c2 10

After treatment, cells were washed twice with ice-cold PBS containing 10% fetal bovine serum, centrifuged at 1000 r.p.m. for 5 min, and lysed in 50 μl of ice-cold Cell Lytic (Sigma) supplemented with protease (Roche Diagnostics Corporation, Indianapolis, IN, USA) and phosphatase (Sigma) inhibitors. Protein concentrations were determined by the BSA assay (Invitrogen) and 50 μg of protein electrophoresed by SDS–PAGE (Invitrogen). Separated proteins were transferred to nitrocellulose membranes utilizing an iBlot (Invitrogen) device. Blots were probed with MCL-1 (clone S-19; Santa Cruz Biotechnology, La Jolla, CA, USA), PARP (clone C2-10) and BCL-2 (Clone 7; both BD Biosciences, CA, USA), caspase-3 (clone 31A1067, Abcam, Cambridge, UK) or β-actin (Sigma) antibodies followed by IRDye 680/800CW-conjugated secondary antibodies (LICOR Biosciences, NE, USA). Proteins were visualized using the Odyssey infrared imaging system (LICOR Biosciences) and were not further manipulated with imaging software.

Flow cytometry, qRT-PCR and Western Blot were performed according to standard methods as described earlier (22 (link)). Western blot was performed by using antibodies against alpha-Tubulin (clone B-5-1-2, Sigma-Aldrich), GAPDH (clone 6C5, Santa Cruz Biotechnology), Caveolin-1 (N-20 polyclonal igG, Santa Cruz Biotechnology), PARP (clone C2-10, BD Pharmingen), Lamin A/C (clone mab636, ThermoFisher Scientific).