Figure 2. .

Carcinine prevents 4-HNE modification of retinal proteins. (A) Carcinine (Car, 2.5 μg) or retinal proteins (Prot, 2.5 μg) were incubated with 0 (−) or 0.64 mM (+) 4-HNE for 16 hours at room temperature before dot-blot analysis. Protein loading is verified by staining the membrane with Ponceau red (lower blot). Signal is detected with anti-HNE antibody coupled with HRP (upper blot). Only retinal proteins modified by 4-HNE were detected by this method. (B) Representative dot-blot showing time-course of adduct formation between 4-HNE and retinal proteins. Ponceau staining (lower blot) and 4-HNE-protein adduct detected using anti-HNE antibody (upper blot) are shown. This experiment was repeated three times with similar results. (C) Representative dot-blot showing dose-response inhibition of adduct formation between 4-HNE and retinal proteins by carcinine. Results from three experiments, each done in triplicate, were quantified and plotted as percentage of adduct formed after normalizing to protein amount. Error bars correspond to SEM.