Aim and objective To compare the effectiveness of erbium:yttrium–aluminum–garnet laser, GentleWave irradiation, photodynamic therapy (PDT), and sodium hypochlorite in smear layer removal and dentin permeability with a scanning electron microscope (SEM). Materials and methods Seventy-five recently extracted single-rooted teeth (maxillary second premolars) were randomly divided into 5 groups of 15 each. Group I teeth was the control group in which conventional root canal preparation (RCP) [17% ethylenediaminetetraacetic acid (EDTA)] was done without laser irradiation, group II teeth underwent RCP and GentleWave™ treatment, group III teeth were subjected to Er:YAG laser irradiation, group IV uses low-level 660 nm (PDT), and group V samples were irrigated with 5.25% NaOCl. All samples were viewed under the SEM. Images at the coronal, middle, and apical part of the root canal were obtained at ×1000. A scoring system for smear layer removal and debris removal scoring was used for analysis. Results Smear layer removal was significantly higher at different points (coronal, middle, and apical area) in group I, followed by V, IV, II, and group III in declining order (p < 0.05). Intercomparison between the groups at different points indicates a significant difference in smear layer removal score between group I and group V at coronal, middle, and apical third. The result was not significant at coronal third and middle third, between group I and V, II and III, II and IV. The result was not significant at apical third between I and V, II and III, and II and IV (p < 0.05). Ethylenediaminetetraacetic acid and sodium hypochlorite are effective in smear layer removal followed by the Er-YAG laser technique. Conclusion Ethylenediaminetetraacetic acid and NaOCl are effective in smear layer removal. Er:YAG laser-activated RCP was comparatively efficient in cleaning the smear layer and opening dentinal tubules. Clinical significance Er:YAG laser-activated RCP was comparatively efficient in cleaning the smear layer and it can be used for effective removal of smear layer for clinical usage. How to cite this article Dash S, Ismail PMS, Singh J, et al. Assessment of Effectiveness of Erbium:Yttrium–Aluminum–Garnet Laser, GentleWave Irradiation, Photodynamic Therapy, and Sodium Hypochlorite in Smear Layer Removal. J Contemp Dent Pract 2020;21(11):1266–1269.