Aim:The aim of this study is to evaluate the effect of ultrasonic activation on the penetration depth of AH Plus, Acroseal, and EndoREZ sealers into dentinal tubules using Confocal laser microscopy.Methods:A total of 60 single-rooted teeth were prepared and divided into three groups on the basis of the type of sealer used (G1: AH Plus, G2: Acroseal and G3:EndoREZ). Each group was further subdivided into two equal subgroups on the basis of ultrasonic activation of sealers and nonactivation. Teeth were obturated and coronally sealed with GIC. Horizontal sections at 2 mm, 4 mm, and 6 mm from the apex were obtained, and the depth of sealer penetration into the dentinal tubules was measured using confocal laser scanning microscopy. Statistical analysis was performed using Students unpaired t-test, ANOVA, and Tukey's multiple comparison test.Results:Within the three groups, ultrasonically activated subgroups showed significantly (P < 0.05) higher depth of sealer penetration. Among the groups, Group 1 showed significantly (P < 0.05) higher depth of sealer penetration than Group 2 and Group 3 while the difference between Group 2 and Group 3 was not significant (P > 0.05).Conclusion:The use of ultrasonic activation with AH Plus sealer showed better results.