Hyperplastic fibro-epithelial lesions are the most common tumor-like swellings in the mouth. The neodymium yttrium aluminium garnet (Nd:YAG) laser appears to be useful for the surgical treatment of these lesions. Some controversies of laser surgery concern the accuracy of pathological diagnosis as well as the control of thermal damage on the target tissue. The aim of this study was to establish if the thermal changes induced by the Nd:YAG laser may affect the histopathological diagnosis and the evaluation of the resection margins. Furthermore, we compared the histological features of oral benign fibro-epithelial lesions excised through Nd:YAG laser and traditional scalpel. Twenty-six benign fibro-epithelial oral lesions from 26 patients, localized in the same oral subsites (cheek and buccal mucosa), were collected at the Unit of Oral Pathology and Oral Laser-assisted Surgery of the Academic Hospital of the University of Parma, Italy. Specimens were subclassified into three groups according to the tool used for the surgical excision. Group 1 included six specimens excised through Nd:YAG laser with an output power of 3.5 W and a frequency of 60 Hz (power density 488,281 W/cm2); Group 2 included nine specimens excised through Nd:YAG laser with an output power of 5 W and a frequency of 30 Hz; Group 3 included 11 specimens excised through a Bard-Parker scalpel blade no. 15c. Epithelial changes, connective tissue modifications, presence of vascular modifications, incision morphology and the overall width of tissue modification were evaluated. Differences between specimens removed with two different parameters of Nd:YAG laser were not significant with regard to stromal changes (p=0.4828) and vascular stasis (p=0.2104). Analysis of regularity of incision revealed a difference which was not statistically significant (p=1.000) between group 1 and group 2. Epithelial and stromal changes were significantly more frequent in specimens with a mean size less than 7 mm (p<0.0001). Nd:YAG laser induced serious thermal effects in small specimens (mean size less than 7 mm) independently from the frequency and power employed. The quality of incision was better and the width of overall tissue injuries was less in the specimens obtained with higher frequency and lower power (group 1: Nd:YAG laser at 3.5 W and 60 Hz).