Anti ho 1

Spinal cord tissues or cells were collected for WB assay as we described previously [22 (link)]. The following antibodies were used: anti-cleaved caspase 3 (1 : 1000, Affinity, USA), anti-Bax (1 : 1000, Affinity, USA), anti-Bcl-2 (1 : 1000, Affinity, USA), anti-PGC-α (1 : 1000, CST, USA), anti-NRF2 (1 : 1000, CST, USA), anti-p-AMPK (1 : 1000, CST, USA), anti-AMPK (1 : 1000, CST, USA), anti-GLUT4 (1 : 1000, Affinity, USA), anti-GLUT3 (1 : 1000, Affinity, USA), anti-HO-1 (1 : 1000, Affinity, USA), anti-NQO1 (1 : 1000, Affinity, USA), anti-β-actin (1 : 1000, Proteintech, USA), HRP-conjugated AffiniPure Goat Anti-Mouse IgG (1 : 10000, Proteintech, USA), and HRP-conjugated AffiniPure Goat Anti-Rabbit IgG (1 : 10000, Proteintech, USA).

Cells were seeded in a 24-well plate and processed accordingly. All cells were washed 5 times with PBS for 3 minutes and fixed in 4% PFA for 20 minutes. After washed by PBS, cells were incubated for 15 minutes in 0.1% Triton X-100 and then cultured for 2 h with 5% goat serum. Then, cells were incubated with the primary antibody at 4°C overnight. Next, the cells were incubated with secondary antibody for 2 hours at room temperature. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI) (Invitrogen, USA) solution for 15 min. The cells were then imaged with a fluorescent microscope (Olympus, Tokyo, Japan), and the results were dealt with by ImageJ. The following antibodies were used: anti-GLUT4 (1 : 200, Affinity, USA), anti-GLUT3 (1 : 200, Affinity, USA), anti-HO-1 (1 : 200, Affinity, USA), anti-NQO1 (1 : 200, Affinity, USA), anti-NRF2 (1 : 200, Affinity, USA), anti-NeuN (1 : 1000, Abcam), TRITC phalloidin (1 : 200, Solarbio, Beijing, China), and Alexa Fluor 488/568 goat anti-mouse IgG and Alexa Fluor 568 goat anti-rabbit IgG (1 : 1000, Thermo Fisher Scientific).