NRF2 Antibody / Nuclear factor erythroid 2-related factor 2 / NFE2L2 (FY13169)

Flow Cytometry analysis of human RT4 cells using anti-NRF2 antibody. Overlay histogram showing RT4 cells stained with (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-NRF2 antibody (1 ug/million cells) for 30 min at 20oC. DyLight 488 conjugated goat anti-rabbit IgG (5-10 ug/million cells) was used as secondary antibody for 30 minutes at 20oC. Isotype control antibody (Green line) was rabbit IgG (1 ug/million cells) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Western blot analysis of NRF2/NFE2L2 using anti-NRF2 antibody. Lane 1: human U20S whole cell lysates, Lane 2: human U20S(+) whole cell lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-NRF2 antibody at 0.5 ug/ml overnight at 4oC, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. NRF2/NFE2L2 antibody detects a doublet at ~90-100 kDa in U2OS cells following MG132 treatment. Although the calculated mass is ~68 kDa, NRF2 typically migrates slower due to phosphorylation and ubiquitination. Proteasome inhibition (MG132) stabilizes NRF2, increasing signal intensity and revealing multiple modified species.