Conditioning of the enamel surface is a common strategy to improve the binding of restorative materials. Er:YAG is used broadly in clinics. However, whether it is beneficial to the binding of restorative materials is controversial. In the current report, we examined the effects of Er:YAG laser in improving the binding of the glass ionomer cement (GIC) to enamel surfaces. Briefly, 77 human premolar and molar teeth free of visible caries were used from the study and treated with different methods, including regular abrasion with diamond saw (Bur), 10 polyacrylic acid, 37% phosphoric acid, and/or Er:YAG laser. The shear bond strength (SBS) between GIC and enamel surfaces were measured; the patterns of the junction between GIC and enamel were observed by scanning electron microscopy (SEM); failure patterns were analyzed with stereomicroscope to determine the adhesive and cohesive patterns of the fracture. The results showed that the treatment of Er:YAG laser resulted in a higher SBS values than that of bur. The use of 10% polyacrylic acid could improve the GIC bonding to the bur-prepared enamel, but not for laser-prepared enamel surface. However, the treatment with 37% phosphoric acid increased the SBS dramatically both in bur-prepared and laser-prepared groups. The failure mold analysis and SEM observation demonstrated a cohesive failure within the cement. In conclusion, the treatment of Er:YAG laser was beneficial for the adhesion of GIC to enamel.