Anti ndufs3

Protein samples were lysed in 5 × SDS sample buffer containing beta-mercaptoethanol, boiled at 95°C for 10 min and loaded into each well for SDS-PAGE. Samples were then transferred to PVDF membrane (Thermo) and immunoblotted using anti-NDUFA9 (Invitrogen), anti-NDUFS3 (Invitrogen), anti-NDUFA13 (Invitrogen), anti-ATP5A (Invitrogen), anti-SDHA (CST), anti-VDAC (CST), anti-HSP60 (CST), anti-PHB1 (CST), anti-PDH (CST), anti-GAPDH (Sigma), anti-GRP78 (SCBT), anti-ABCA1 (SCBT), anti-FLOT1 (CST), anti-STAT3 (CST), anti-STAT1 (SCBT), anti-AMPK (CST), anti-ERK1/2 (CST), anti-p38 (CST), anti-SYP (Abcam), anti-ACSL4 (Abcam), all diluted in 5% BSA:TBST at 1:1000, followed by appropriate HRP-conjugated secondary antibodies (Thermo) incubation (diluted in 5% non-fat milk:TBST at 1:10,000), and developed using the SuperSignal^TM West Femto Maximum Sensitivity Substrate (Thermo).

Cells were lysed with RIPA buffer (50 mM Tris-HCl pH7.4, 150 mM NaCl, 1% NP-40, 0.25% Na-deoxycholate and 1 mM EDTA), centrifuged at 20000 g for 10 min and supernatant were collected. Protein concentrations were measured and 20 μg of total protein were loaded into each well in SDS-PAGE. Samples were then transferred to PVDF membrane(Thermo) and immunoblotted with anti-NDUFS3 (Invitrogen), anti-NDUFA13(Invitrogen), anti-SDHA(CST), anti-beta-actin (Santa Cruz), anti-GAPDH (Sigma), anti-STAT3 (CST), anti-pAMPK (CST), AMPK (CST), anti-pS-AKT(CST), anti-AKT(CST), anti-CASP3(CST) anti-p-p38(CST), anti-p38(CST), anti-mTOR(CST), anti-pS-STAT3(CST), anti-pY-STAT3(CST), anti-LC3(CST), anti-GFAP(SCBT), anti-ALDH1L1(Abcam) followed by HRP-conjugated secondary antibody (Thermo) incubation and the SuperSignal West Femto Maximum Sensitivity Substrate (Thermo) was used to detect signal.