Arsenite and UVA activate NADPH oxidase. HaCaT cells were incubated with 3 μM arsenite (■) or 10kJ/m2 UVA (▲) for the indicated times. NADPH oxidase activity was assessed over time as described in “Experimental Procedures” (A). Total protein was collected at the indicated times and 20 μg resolved via a 8-12% SDS-PAGE gradient gel and the indicated proteins were analyzed, transferred to nitrocellulose membrane, immunoreacted with the appropriate antibodies and detected by ECL. Panel (B) shows results following arsenite treatment and UVA results are shown in (C). Equal protein loading was confirmed by reprobing original membranes with β-tubulin. Each point and blot is representative of at least 3 independent experiments and error bars indicate ± S.D. All p values were <0.05 versus the untreated control. * = significantly different from untreated controls.