Purpose: The present study aimed to investigate the impact of a femtosecond laser and Er:YAG laser as an alternative to orthophosphoric acid etching on the shear bond strength. Methods: Thirty permanent mandibular teeth were assigned to one of three groups based on the treatment performed on their buccal/lingual surfaces. Group I, 37% orthophosphoric acid etching; Group II, Er:YAG laser; Group III, femtosecond laser. A 3 × 3 mm diameter bonding area was demarcated on the enamel. Fissure sealant material was applied to the test areas and cured for 20 s in all groups. The shear bond strength of all the specimens was determined using an Instron universal testing machine. After testing, the fracture surfaces were analyzed using a stereomicroscope, and the fracture modes of each sample were categorized according to the adhesive remnant index. Results: The results revealed no significant differences in terms of shear bond strength of the femtosecond laser (7.55 ± 0.6 MPa), Er:YAG laser (7.26 ± 0.6 MPa), and acid etching groups (6.61 ± 0.5 MPa) (p > 0.05). Conclusion: The present in vitro study showed that the bond strength of the enamel surface following treatment with femtosecond and Er:YAG laser systems alone was sufficient when compared with that after acid etching. Clinical significance It is possible to use femtosecond lasers for bonding fissure sealant to enamel surfaces as an alternative to acid etching.