Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Oral cancer is a public health problem with high prevalence in the population. Local tumor control is best achieved by complete surgical resection with adequate margins. A disease-free surgical margin correlates with a lower rate of local recurrence and a higher rate of disease-free survival. Fluorescence spectroscopy is a noninvasive diagnostic tool that can aid in real-time cancer detection. The technique, which evaluates the biochemical composition and structure of tissue fluorescence, is relatively simple, fast and, accurate. Objectives: This study aimed to compare oral squamous cell carcinoma lesions to surgical margins and the mucosa of healthy volunteers by fluorescence spectroscopy. Materials and methods: The sample consisted of 56 individuals, 28 with oral squamous cell carcinoma and 28 healthy volunteers with normal oral mucosa. Thirty six cases (64.3%) were male and the mean age was 60.9 years old. The spectra were classified and compared to histopathology to determine fluorescence efficiency for diagnostic discrimination of tumors. Results: In the analysis of the other cases we observed discrimination between normal mucosa, injury and margins. At two-year follow up, three individuals had local recurrence, and in two cases investigation fluorescence in the corresponding area showed qualitative differences in spectra between the recurrence area and the area without recurrence at the same anatomical site in the same patient. Conclusion: In situ analysis of oral mucosa showed the potential of fluorescence spectroscopy as a diagnostic tool that can aid in discrimination of altered mucosa and normal mucosa. (c) 2014 Elsevier Ltd. All rights reserved.506593599Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)ALNF scholarship [140493/2011-5]INCITOFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [98/14270-8, 07/57126-5, 11/10802-1]CNPq [477439/2007-1]ALNF scholarship [140493/2011-5]