Purpose: The aim of this study was to evaluate the effectiveness of previous dental substrate treatments on the bond strength of two self-etching adhesives. Materials and Methods: One hundred bovine incisors were sectioned at the cementoenamel junction. The labial tooth surface was ground to obtain flat superficial dentin. Teeth were randomly divided into two groups (n = 50): Clearfil Protect Bond and One Up Bond F. The specimens of each group were subdivided into five groups, according the dental substrate treatment, and then the adhesive system was applied as follows (n = 10): C: control, without previous treatment; TA: 37% phosphoric acid etching on enamel and dentin for 15 s; PA: 37% phosphoric acid etching only on enamel for 15 s; SB: Al2O3 sandblasting on dental surface; EDTA: enamel and dentin conditioning with 24% EDTA gel, pH 7.0. Resin composite blocks were fixed to the dental surfaces with dual-polymerized resin cement. Samples were sectioned in the mesiodistal direction and were trimmed to reduce the cross-sectional area to approximately 1.2 mm2. They were tested at a crosshead speed of 0.5 mm/min on a testing machine. Microtensile bond strength values from enamel, superficial dentin, and deep dentin were compared. Mean microtensile bond strength values were expressed in MPa and were analyzed by two-way ANOVA (2 x 3 x 5) and Tukey's tests. Results: A significant difference was observed for interaction (p = 0.00001) among the factors. The highest mean bond strength to enamel was found with previous phosphoric acid etching, and to dentin when EDTA was previously applied. Conclusion: The most indicated previous treatment for increasing the bond strength to enamel and dentin simultaneously is EDTA for both self-etching adhesive systems used. Keywords: dentin bonding, microtensile test, resin-dentin bond strength