After 48 h in the incubation buffer at 37 °C, proteolytic activity was registered under long-wave UV light scanner and quantified by using Image J software. 
Image J
Endogenous Enzymatic Activity in Dentin Treated with a Chitosan Primer.The aim of this study was to evaluate the effect of different concentrations of chitosan polymer on dentinal enzymatic activity by means of gelatin and in situ zymography.  Dentin powder aliquots were demineralized with phosphoric acid and treated with three different concentrations of lyophilized chitosan polymer (1, 0.5 and 0.1 wt%) dissolved in distilled water.  Dentin proteins were extracted from each experimental group and electrophoresed under non-reducing conditions in 10% SDS-PAGE containing fluorescein-labeled gelatin.  After 48 h in the incubation buffer at 37 °C, proteolytic activity was registered under long-wave UV light scanner and quantified by using Image J software.  Furthermore, additional teeth (n = 4) were prepared for the in situ zymographic analysis in unrestored as well as restored dentin pretreated with the same chitosan primers.  The registered enzymatic activity was directly proportional to the chitosan concentration and higher in the restored dentin groups (p < 0.05), except for the 0.1% chitosan primer.  Chitosan 0.1% only showed faint expression of enzymatic activity compared to 1% and 0.5% concentrations.  Chitosan 0.1% dissolved in water can produce significant reduction in MMPs activity and could possibly contribute to bond strength preservation over time.
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