Nrf2 polyclonal antibody (rabbit anti human, 1:200 diluted, ImmunoWay Biotechnology Company) for 1 h at 37°C. Membranes were then incubated with horseradish peroxidase-conjugated antibody (goat anti rabbit, 1:5000 diluted, Boster biological engineering co., LTD, Wuhan, China; rabbit anti chicken, 1:5000 diluted, Takara, Dalian, China) for 1 h at RT. After further washing with phosphate-buffered saline, blots were incubated in commercial chemoluminescence reagents (Amersham Biosciences, USA). Band intensities were measured using Quantity One software (BioRad, Hercules, CA). The quantity of Nrf2 protein expression in nucleus was relative to Histon H1, and in cytoplasm was relative to β-actin. Nrf2 and CYP2A5/2A6 protein expressions in hepatocytes were relative to β-actin.
