Background: Due to the anatomical complexity of the root canal system, irrigation plays an essential role in endodontics. This in vitro study was sought to compare the removal of the smear layer (RSL) promoted by conventional irrigation (CI), passive ultrasonic irrigation (PUI) and continuous ultrasonic irrigation (CUI) with 17% EDTA, by using scanning electron microscopy (SEM). Material and Methods: Forty single-rooted human mandibular canines were instrumented and randomly assigned to four groups (n=10), according to the irrigation protocol aiming to the RSL: CG (control group) —conventional irrigation with distilled water; CI— conventional irrigation with 17% EDTA; PUI —passive ultrasonic irrigation with 17% EDTA; CUI— continuous ultrasonic irrigation with 17% EDTA. Hemisections from each sample were obtained, and images of each root canal third (cervical, middle and apical) were captured at 1000 X magnification by SEM. Three previously calibrated and blinded evaluators classified the RSL, according to the criteria proposed by Torabinejad et al.: small or no smear layer (all dentinal tubules were clean and open); 2 = moderate smear layer (no smear layer on the surface of root canal, but dentinal tubules contained debris); 3 = dense smear layer (covering practically all dentinal tubules entrances). Statistical analysis was performed by Kruskal-Wallis and Bonferroni tests (p<0.05). Results: Overall, CUI and cervical thirds showed better RSL rates, compared with the other methods and thirds, respectively (p<0.05). More specifically, the cervical thirds showed better results in the CG, CI and PUI groups (p<0.05), whereas the cervical and middle thirds were not significantly different in the CUI group. Conclusion: CUI was the most effective method for the RSL.