Objective: The objective of the current study was to evaluate the laser agitation of nanoparticle irrigants on root canal cleanliness.  Materials and methods: A total of 60 sounds freshly-extracted maxillary central incisor teeth were collected to be used in this study. The selected roots were divided into two main groups, according to irrigant solution used, thirsty roots each. Each main group was subdivided into three subgroups, according to the agitation of the irrigant solution and the time of agitation used, ten roots each (n=10). All roots were prepared using Protaper rotary file system. In group (I) 30 of the selected roots were irrigated with 5ml 17% EDTA  solution, 10 roots were not activated,10  roots were activated by laser for 10seconds and 10 roots were activated by  laser for 20seconds. All roots were then irrigated with 5ml 5.25% NaOCl followed by 5ml saline as a final rinse. In group (II) 30 of the selected roots were irrigated with 5ml 0.2% Chitosan nanoparticle irrigant solution, 10 roots were not activated, 10 roots were activated by laser for 10 seconds, and 10 roots were activated by laser for 20 seconds. Then, these roots were irrigated with 5ml 5.25% NaOCl followed by 5ml saline as a final rinse. Specimens were scanned using a scanning electron microscope and statistical analysis was performed using Graph-Pad Instat statistics software. Results: In removal of debris and smear layer the number of open dentinal tubules was higher in group (I) than in group (II) and statistically significant differences were found between both  groups in the cervical, middle and apical thirds of the root canals (P < .05). Conclusions: Laser agitation is an effective technique in debris and smear layer removal when used either with 17% EDTA or 0.2% Chitosan nanoparticle irrigant solutions, with better results when used with 17% EDTA.