To evaluate HO-1, NQO1 and SOD expression, IEC-6 cells (2 × 104 cells/well; 96-well plates) were treated with IS (31.2–250 μM) for 24 h.
Thereafter, the cells were collected, washed with PBS, and incubated with Fixing Solution for 20 min at 4 °C and successively for 30 min at 4 °C with Fix Perm Solution. Anti-HO-1 (sc-10789, Santa Cruz Biotechnology) or anti-NQO1 (sc-376023, Santa Cruz Biotechnology) or anti-SOD (sc-30080, Santa Cruz Biotechnology) antibody were subsequently added. The secondary antibody was added, for 30 min, in Fix Solution, and cell fluorescence was evaluated by FACSscan (Becton Dickinson) and elaborated with Cell Quest software, as previously reported [60 (link)].
Thereafter, the cells were collected, washed with PBS, and incubated with Fixing Solution for 20 min at 4 °C and successively for 30 min at 4 °C with Fix Perm Solution. Anti-HO-1 (sc-10789, Santa Cruz Biotechnology) or anti-NQO1 (sc-376023, Santa Cruz Biotechnology) or anti-SOD (sc-30080, Santa Cruz Biotechnology) antibody were subsequently added. The secondary antibody was added, for 30 min, in Fix Solution, and cell fluorescence was evaluated by FACSscan (Becton Dickinson) and elaborated with Cell Quest software, as previously reported [60 (link)].
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