To explore the effect of combination of pulsed CO2 laser irradiation and diammine silver fluoride treatment on the ultrastructure of dentine. Each extracted molars was made into four identical dentinal specimens, which were randomly divided into 4 groups according to the treatment methods: A, control group; B, pulsed CO2 laser irradiation; C, coated with 38%Ag(NH3)2F; and D, laser irradiation plus fluoride. Scanning electron microscope (SEM) analysis was performed. As shown by SEM, group A showed clear dentinal tubuli, group B showed melted surface without dentinal tubule openings, group C showed thicker melted layer without dentinal tubule openings, and group D showed the thickest melted and recrystallized layer on dentine surface. Both pulsed CO2 laser irradiation and 38%Ag(NH3)2F treatment can seal dentinal tubule. The combination of these two techniques can form a synergistic effect.