Apoptosis Pathway Protein Analysis

Primary antibodies against p53, Phospho-Akt (S473), Total-Akt, PUMA, Phospho-FoxO3a (S253), Total-FoxO3a, Phospho-p65 (S536), Total-p65, Bax, Noxa, Bid, Bim, Bcl-2, Bcl-XL, Mcl-1, Cox IV, Cleaved-Caspase3, β-actin LC3, Total-MLKL, and Phospho-MLKL (Ser358) were purchased from Cell Signaling Technology. Lipofectamine™ Reagent was purchased from Invitrogen. HRP-conjugated anti-rabbit and/or anti-mouse secondary antibodies and ECL-plus kit were from GE Healthcare. Ipatasertib and 5-FU were purchased from APP Pharmaceuticals. afuresertib and perifosine were purchased from Selleck Chemicals(Houston, TX). Regofenib and cisplatin were purchased from Axon Medchem. Other chemicals were mainly from Sigma. CCK-8 kit was from 7 Sea Biotech (Shanghai, China). The plasmid of expressing PUMA was kindly supplied by Jian Yu, PhD^{9 (link)}.