The purpose of this study was to evaluate the thickness and qualitative characteristics of the hybrid layer after two cavity preparation methods, using Er:YAG laser in QSP mode and conventional carbide burs. Additionally, two different adhesive techniques were investigated using etch-and-rinse and self-etch adhesive systems. Sixty sound human third molars were used and were randomly divided into four groups (n=15). In the first two groups, large (4mm length, 3mm wide, and 3mm deep) class I cavities were prepared using Er:YAG laser (2.94m) in QSP mode, while in the other two groups, the cavities were prepared using carbide burs. After cavity preparations, two different adhesive techniques with GLUMA (R) 2 Bond (etch-and-rinse) and Clearfil Universal Bond Quick (self-etch) were applied. For the qualitative evaluation of the formed hybrid layer, photomicrographs were taken using SEM, and elemental semi-quantitative analysis was performed using EDS to confirm the extent of the hybrid layer. One-way ANOVA was applied to verify the existence of statistically significant differences, followed by Tukey test for post hoc comparisons (Bonferroni corrected), and the level of significance was set at a=0.05. The laser-treated groups exhibited higher hybrid layer thickness than bur-treated groups (p<0.001). Between the laser-treated groups, etch-and-rinse technique presented higher hybrid layer thickness than self-etch technique (p<0.001), while between the bur-treated groups, no significant differences were detected (p=0.366). Er:YAG laser cavity preparations in QSP mode may be advantageous for adhesion of composite restorations, but more data are necessary to confirm its clinical effectiveness.