Horseradish peroxidase labeled goat anti rabbit immunoglobulin g igg h l

ZPP 50 mg/vial was obtained from Sigma-Aldrich Co., St Louis, MO, USA. Bicinchoninic acid (BCA) protein assay kit, mouse anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) monoclonal antibody and horseradish peroxidase labeled goat anti-rabbit immunoglobulin G (IgG) (H + L) were purchased from Beyotime Institute of Biotechnology, Haimen, Jiangsu, People’s Republic of China. Rabbit anti-Bcl-2, Bax, NF-κB p65, p-NF-κB p65 and IκBα, p-IκBα monoclonal antibodies were from Cell Signaling Technology, Beverly, MA, USA. Rabbit anti-HO-1 polyclonal antibody were from Abcam Corporation, Cambridge, MA, USA. TRIzol assay kit and one step reverse transcription polymerase chain reaction (RT-PCR) kit were from Thermo Fisher Scientific, Waltham, MA, USA. Caspase 3, Caspase 8, Caspase 9 activity detection kit were from Nanjing KeyGen Biotechnology Corporation Limited, Nanjing, People’s Republic of China. TNF-α, IL-1β, IL-6 and IL-8 ELISA assay kit were from R&D System, Minneapolis, MN, USA. TUNEL assay kit were from Wuhan Boster Biotechnology Corporation Limited, Wuhan, People’s Republic of China. Electrophoresis, electrophoresis transfer and gel imaging system were from Bio-Rad Laboratories, Hercules, CA, USA. Microplate reader were from TECAN Trading AG, Männedorf, Switzerland.

Proteins extracted from cells cultured under different oxygen conditions were subjected to a western blot test using standard procedures. The primary antibodies are listed in Table 2. They are HIF1-α Rabbit pAb (ABclonal Cat# A11945, RRID:AB_2758885), HO-1 Rabbit pAb (ABclonal Cat# A19062, RRID:AB_2862555), p-gp Rabbit pAb (ABclonal Cat# A19093, RRID:AB_2862585), GAPDH Rabbit pAb (ABclonal Cat# A19056, RRID:AB_2862549), MRP1 Rabbit pAb (Abcam Cat# ab260038, RRID:AB_2889834). The secondary antibodies horseradish peroxidase-labeled goat anti-rabbit immunoglobulin G (IgG) (H + L) (Beyotime Cat# A0208, RRID:AB_2892644) was purchased from Beyotime (Shanghai, China). Protein bands were detected by Western Blotting Substrate (ECL). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an internal reference.