Cells growing in monolayers were lysed using Cell Extraction Buffer (Life
Technologies) supplemented with complete protease inhibitors and PhosSTOP phosphatase
inhibitor cocktail tablets (Roche). Cell lysates were cleared by centrifugation, protein
concentrations were determined by DC Protein Assay (BioRad), and denatured lysates were
run on 4–12% Bis-Tris gradient gels (Invitrogen). Gels were transferred to
nitrocellulose membranes (BioRad) before being immunoblotted with indicated antibodies.
Cleaved-PARP antibody was obtained from Cell Signaling Technology. RAC1 activation assays
were performed as previously described according to the manufacturer's protocol (Cell
Biolabs) (10 (link)).
Technologies) supplemented with complete protease inhibitors and PhosSTOP phosphatase
inhibitor cocktail tablets (Roche). Cell lysates were cleared by centrifugation, protein
concentrations were determined by DC Protein Assay (BioRad), and denatured lysates were
run on 4–12% Bis-Tris gradient gels (Invitrogen). Gels were transferred to
nitrocellulose membranes (BioRad) before being immunoblotted with indicated antibodies.
Cleaved-PARP antibody was obtained from Cell Signaling Technology. RAC1 activation assays
were performed as previously described according to the manufacturer's protocol (Cell
Biolabs) (10 (link)).
