Aim: To compare the depth of penetration and persistence of sealer residues in obturated canals with three commercially available root canal sealers using confocal laser scanning microscopy (CLSM). Methodology: Thirty single-rooted human premolars were selected and decoronated with diamond disks to standardize the root length at 14 mm. Access cavity and working length were determined; following which biomechanical preparation was carried out using rotary files in a crown-down manner. Copious irrigation was carried out during recapitulation so as to effectively debride the canal. Subsequently, the canals were dried and obturated using lateral condensation technique with gutta-percha (GP) sticks, coated with three categories of dye-incorporated sealers, that is, AH Plus, Sealapex, and BioRoot RCS. The teeth were coronally sealed and allowed to set for 2 weeks. All samples were reattended, sectioned 6 mm from the apex, and evaluated by confocal laser scanning microscope. Results: AH Plus sealer showed the highest depth of penetration followed by BioRoot RCS and Sealapex, respectively. Conclusion: The depth of penetration of sealers plays a pivotal role in the outcome of endodontic treatment. It is virtually impossible to remove the sealer residue of the contracted dentin tubules. However, complete removal is not an essential factor in follow-up treatment in endodontics. Clinical significance: The study provides insights into selection of appropriate sealer to achieve optimal penetration and retrievability clinically.