ABSTRACT Aim: The aim of this study was to evaluate the penetration depth of three different irrigant solutions incorporated with nanoparticles (NPs) using a confocal laser scanning microscope. Methods: Forty-two single-rooted teeth were used. Access cavities were prepared, and working length was determined. The samples were split into three groups at random (n = 14). Group 1 – chitosan NPs, Group 2 – zinc oxide NPs, and Group 3 – silica-doped titanium NPs. Each group was further divided into subgroup A with activation and subgroup B without activation. Each tooth received final irrigants for 1 min. Each sample is sectioned at 3 mm from the apex. A confocal laser scanning microscope was used to observe the irrigant penetration. Results: The results showed that there was a significant difference in the penetration depth in Group 2A and Group 3A. Conclusion: Activation with PATS Vario increases the penetration when compared to conventional syringe, and zinc oxide NP solution showed increased penetration when compared with other groups.