Cleaved caspase 9 asp330

RPPA was performed as previously published by us [16 (link)] (more details in the Supplementary Data). The following antibodies were used: Parp-1#sc-7150 (Santa Cruz, TX, USA). Cleaved caspase-9 (Asp315) #9505, cleaved caspase-9 (Asp330) #9501, cleaved caspase-7 (Asp198) #9491 and XIAP#2042 (Cell Signaling, MA, USA).

Western blot assay was performed as described previously [32 (link)]. Briefly, samples were loaded (20 µg) and electrophoresis was carried out at 60 V throughout stacking gel and at 120 V afterwards. Transfer to a polyvinylidene difluoride (PVDF) membrane was performed at 80 V for 2 h followed by blocking with 5% skimmed milk for 1 h. The membrane was incubated with following primary antibodies overnight in 4 °C: cleaved caspase-3 (Asp175) (#9661), cleaved caspase-9 (Asp330) (#9501), poly (ADP-ribose) polymerase (PARP) (#9542), SQSTM1/p62 (#5114), microtubule-associated protein 1A/1B-light chain 3 (LC3B) (#2775), p53 (#2527), Akt (#9272), p-Akt (#9271), ribosomal protein P70S6 kinase beta-1 (P70S6K), p-p70S6K and anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (#2118), all purchased from Cell Signaling technology (Danvers, MA, USA). Then, the membrane was incubated with appropriate secondary antibodies (1:1000) for 1 h, and enhanced chemiluminescence (ECL) solution was used to detect protein bands.