Anti gclm 14241 1 ap

Anti-Nrf2 (ab62352), anti-Ubiquitin (ab134952), anti-IL-1β (ab45692), anti–NF–κB p65 (ab16502), anti-SOD1 (ab13498), anti-GPx1 (ab108427) and anti-catalase (ab76110) antibodies were purchased from Abcam Technology (Abcam Technology, England). Anti-phospho–NF–κB p65 antibodies (CST #3031), anti-phospho-IκBα (CST #2859) and anti-phospho-IKKβ (CST #2694) antibodies were purchased from Cell Signalling Technology (Cell Signalling Technology, USA). Anti–HO–1 (SC-136960) and anti-NQO1 (SC-271116) antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Anti-β-actin (60008-1-lg), anti-Histone-H3 (17168-1-AP) and anti-GCLM (14241-1-AP) antibodies were purchased from Proteintech Group (Proteintech Group, USA). Isolation of cell fractions and Western blotting were performed as detailed previously [36 (link)].

Anti-Nrf2 (ab62352), anti-HO-1 (ab52947), and anti-NLRP3 (ab263899) antibodies were purchased from Abcam (Abcam Technology, England). Anti- Caspase 1 p20 (PA5-99390) antibody was purchased from Invitrogen (Thermo Fisher Scientific, Waltham, MA, USA). Anti-α-Tubulin (66031-1-Ig), anti-NQO1(67240-1-Ig), and anti-GCLM (14241-1-AP) antibodies were purchased from Proteintech (Proteintech Group, Rosemont, IL, USA). NCM460 cells were incubated with indicated compounds for the specified time interval, and cells were harvested and lysed using RIPA lysis buffer with Roche protease inhibitor complete cocktail. The solution was centrifuged for 10 min at 4 °C, and the total protein concentrations were determined by BCA protein assay. Individual cell lysates were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (12% gel, SDS-PAGE), and the separated proteins were transferred to PVDF membranes by wet transfer. The membrane was blocked in 5% fat-free milk, followed by incubation with a primary antibody overnight at 4 °C and a horseradish peroxidase (HRP)-conjugated secondary antibody for 1 h at room temperature. The membrane was imaged by Tanon 5200 Multi Imaging Workstation.