The aim of the present in vitro study was to evaluate the effect of chlorhexidine, applied before a self-etching adhesive system (Clearfil SE Bond) on microtensile bond strength to superficial and deep dentin (DD), immediately and after six months of water storage (WS). Forty dentin specimens were divided into two groups according to dentin depth: superficial and deep. The specimens were then divided according to the solution to be applied (n=10): CLX: 2% chlorhexidine (passively applied for 60s) and NT (no treatment). A self-etching adhesive system was applied according to the manufactures instructions, with composite restorative placed on the dentin surface. After 24h, dentin-resin blocks were sectioned into beam-shaped specimens that were submitted to microtensile bond strength testing either immediately or after six months of WS. Data were submitted to three-way ANOVA (alpha=0.05). Bond strength values for the deep dentin group were significantly lower than those observed for superficial dentin (SD) (p=0.002), whether chlorhexidine solution had been applied or not. There was no statistical difference in bond strength for specimens tested after 24h and 6months of WS. The application of chlorhexidine did not affect immediate and long-term bond strength to dentin. Bond strength in deep dentin was lower than in SD.