Aim: The present study aimed to evaluate the cleaning efficacy of Brush File as an irrigation activation tool compared to Passive Ultrasonic Irrigation and conventional syringe and needle irrigation without activation in terms of smear layer and debris removal in single-rooted teeth. Subjects and methods: 36 single-rooted premolars with single root-canal system were used, Root apices were sealed to simulate the in-vivo conditions. Instrumentation was done using Wave-one gold reciprocating system, then teeth were assigned randomly into three groups (n=12) according to the final irrigation protocol, group (1): Brush File (BF), group (2): Passive ultrasonic irrigation (PUI), group (3): Conventional irrigation with no activation (CI). Longitudinal splitting of teeth for Scanning Electron Microscope (SEM) imaging, Parente's scoring system was used for evaluation. Data were analyzed using Kruskal-Wallis and Mann-Whitney tests, with significance level set at (p < 0.05). Results: The inter-group comparison showed that both BF and PUI groups removed smear layer and debris better than CI group at all root-canal thirds, with no significant difference between them (p > 0.05). For the intra-group comparison, no significant difference between the three root thirds in both BF group and CI group (P > 0.05). Regarding PUI, apical third showed the highest smear layer scores (P = 0.013), while the coronal third showed higher debris scores than middle third (P = 0.05). Conclusion: No activation protocol completely removed smear layer and debris, Brush file showed great cleaning efficacy for both smear layer and debris at all root-canal thirds that were comparable to PUI.