Objective: The purpose of this in vitro study was to evaluate dentin bond strength to resin composite following high-speed rotary or Er,Cr:YSGG laser preparation using two different adhesive systems. Materials and Methods: One hundred and forty dentin specimens were prepared from human molars. Seventy specimens were prepared with a high-speed diamond bur and 70 of the specimens were prepared with an Er,Cr:YSGG laser. Each group was subdivided into 7 groups of 10 teeth and etched with either 37% H3PO4, laser etched at two different power levels (1.25 W and 3 W), or not etched. Nano-hybrid composite resin cylinders 4 × 2 mm in size (Tetric EvoCeram, Vivadent) were bonded to the dentin surfaces with the application of an etch-and-rinse adhesive (Exite/Vivadent), or a self-etching adhesive (AdheSe/Vivadent). After storage in distilled water and thermocycling, a shear bond strength test was performed on all specimens and the data were statistically analyzed. Results: ANOVA detected significant differences between the tested groups (p < 0.001). Duncan's multiple comparison test showed that Excite-applied specimens that were prepared with laser and laser etched (1.25 W or 3 W) had higher mean bond strengths. Both AdheSe- and Excite-applied specimens prepared with the diamond bur and laser etched (1.25 W) showed the lowest mean bond strength values (p < 0.05). Conclusion: The results suggest that dentin surfaces prepared with the Er,Cr:YSGG laser and etched with the laser (at either 1.25 or 3 W) may provide comparable or increased composite resin bond strengths, depending on the adhesive used. © 2008 Mary Ann Liebert, Inc.