Purpose: The aim of the present investigation is to quantitatively evaluate the influence of Nd:YAG laser treatment on the hybrid layer on the surface of dentin after ceramic inlay luting. Materials and Methods: Twenty-eight extracted human molars were used for this study. An occlusal cavity for an inlay was prepared in each tooth and a ceramic inlay was created. In 14 teeth (Group A), Nd:YAG radiation was applied. In the control group (Group B, n = 14), no laser treatment was performed. The ceramic inlays were luted to all teeth in groups A and B using a dental adhesive resin cement. The restored teeth were then placed in 30% Hcl to dissolve dental tissues. The luted surface of the inlays was examined under SEM. The hybrid layer was classified according to a 4-point scale depending on the density and the length of the resin tags. Data were statistically evaluated using the t test. Results: Group A showed very few, short resin tags. The hybrid layer was well formed in Group B (control). The difference between the experimental groups is statistically significant (p < 0.05). Conclusion: This in vitro study demonstrates that the formulation of the hybrid layer may be affected by Nd:YAG laser conditioning when it is used as a desensitizing treatment before luting ceramic inlays.