Anti gapdh 60004 1g

Immunoblots were performed based on methods described in a previous paper [13 (link)]. The following antibodies were used: anti-NRF2 (GTX103322, GeneTex, Irvine, CA, USA), anti-GAPDH (60004-1g, Proteintech, Rosemont, IL, USA), anti-IκB alpha (12045-R116, Sino Biological, BDA, Beijing, China), anti-phospho- IκB alpha (#9246, Cell Signaling Technology, MA, USA), anti-alpha Tubulin (11224-1-AP, Proteintech, Rosemont, IL, USA), and anti-Lamin B1 (66095-1-Ig, Proteintech, Rosemont, IL, USA). Nuclear and cytoplasmic proteins were collected using EPIXTRACT Nuclear Protein Isolation Kit according to the manufacturer’s instructions.

Cells were washed with PBS and harvested with RIPA lysis buffer (50 mM Tris (pH 7.5), 150 mM NaCl, 1mM EDTA, 1% NP-40, 0.1% SDS) supplemented with protease and phosphatase inhibitors (1 mM PMSF, 10 μg/mL Leupeptin, 50 μg/mL TLCK, 50 μg/mL TPCK, 1 μg/mL Aprotinin, 1 mM NaF, 5 mM NaPPi, and 10 mM Na₃VO₄). The cell lysates were cleared via 15 min spins at 13,000 rpm, 4 °C. Protein concentrations were determined by Bio-Rad Protein Assay (Bio-Rad, Hercules, CA, USA). The following antibodies were used: anti-NRF2 (GTX103322, GeneTex, Irvine, CA, USA), anti-GAPDH (60004-1g, Proteintech, Rosemont, IL, USA), anti-KEAP1 (10503-2-AP, Proteintech, Rosemont, IL, USA), anti-lamin B1 (66095-1-Ig, Proteintech, Rosemont, IL, USA).