Anti nrf2 ab62352

Cells were lysed in NETN buffer (20 mM Tris, at pH 8.0, 100 mM NaCl, 1 mM EDTA and 0.5% NP-40). For all samples, 1000 μg lysate was incubated with Protein G Sepharose beads and then followed by the indicated antibodies according the manufacturer’s instruction. Immunoprecipitates were resolved by immunoblotting with the indicated antibodies: Anti-Nrf2 (#ab62352) was obtained from Abcam plc (Cambridge, CB2 0AX, UK), where as Anti-HA (#H3663) was acquired from Sigma Chemical Co. (St. Louis, MO, USA).

Bafilomycin A1 (S1413) and ZVAD-FMK (S7023) were purchased from Selleckchem (Houston, TX, USA). N-acetyl-L-cysteine (S0077) was from the Beyotime Institute of Biotechnology (Shanghai, China). Sorafenib (HY-10201), Erastin (HY-15763), Ferrostatin-1 (HY-100579), Deferoxamine (HY-B0988), and Necrosulfonamide (HY100573) were obtained from MedChemExpress (MCE; Shanghai, China). Tertiary butylhydroquinone (112941) was purchased from Sigma–Aldrich (Shanghai, China). Brusatol (MB7292) was obtained from Meilunbio (Dalian, China). Carmustine (BCNU) was from Macklin (Shanghai, China). Antibodies against SLC27A5 (NBP1-89267) were brought from Novusbio (Centennial, CO, USA). Anti-NRF2 (ab62352), anti-β-actin (ab6276), and anti-4-HNE (ab46545) were obtained from Abcam (Cambridge, MA, USA). Anti-GSR (sc-133245) was purchased from Santa Cruz Biotechnology (Santa Cruz; CA, USA).

Total cell lysates of SRA-hLECs were prepared in ice-cold radioimmune precipitation buffer (RIPA buffer), and protein blot analysis was performed as described previously [37 (link),40 (link),41 (link),59 (link),60 (link)]. The membranes were probed with anti-Bmal1 (sc-365645, Santa Cruz Biotechnology, Dallas, TX, USA); anti-Clock (#5157S, Cell Signaling Technology, Danvers, MA, USA); Anti-Nrf2 (sc-365949 Santa Cruz Biotechnology); Anti-Nrf2 (ab62352, Abcam^®, Cambridge, MA, USA); Anti-Prdx6 antibody (LF-PA0011, Ab Frontier, South Korea), Anti-NQO1 (ab28947, Abcam^®, Cambridge, MA, USA), Anti-HO1 (Ab13248, Abcam^®, Cambridge, MA, USA), Anti-SOD1 (sc-515404, Santa Cruz Biotechnology, Dallas, TX, USA), Anti-SOD2 (sc-137254, Santa Cruz Biotechnology, Dallas, TX, USA), or β-actin (A2066, Sigma-Aldrich, St. Louis, MO, USA)/Anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH)(sc-365062, Santa Cruz Biotechnology, Dallas, TX, USA)/Tubulin (Abcam, Cambridge, MA, USA) as an internal control to monitor those protein expressions. After secondary antibody (sc-2354 and sc-2768, Santa Cruz Biotechnology, Dallas, TX, USA), protein bands were visualized by incubating the membrane with luminol reagent (sc-2048; Santa Cruz Biotechnology, Dallas, TX, USA), and images were recorded with a FUJIFILM-LAS-4000 luminescent image analyzer (FUJIFILM Medical Systems Inc., Hanover Park, IL, USA).