Background. p16(INK4a) seems to be an indicator of the grade of Human Papillomavirus-induced lesions and a possible predictor of the lesion evolution. There are few studies about the role of HPV test and p16(INK4a) in diagnosis of high-grade cervical lesions in South-American women. The aim of the present study was to evaluate the presence of p16(INK4a) and high-risk HPV-DNA expression in cases diagnosed as squamous intra-epithelial lesion and evaluate their role in the approach of high-grade squamous intra-epithelial lesion. Methods. p16(INK4a) and high-risk Human papillomavirus were investigated in 96 samples of the cervix ( 13 cases of high grade squamous intraepithelial lesions, 26 cases of low grade intraepithelial lesions and 57 normal tissues). The p16(INK4a) was identified by immunohistochemistry using the p16(INK4a) kit (E6H4 clone, DakoCytomation, Carpinteria, CA) and Human papillomavirus DNA was classified by hybrid capture (Digene (R)). Associations were evaluated by the KAPPA index. Results. The p16(INK4a) was detected in 92.3% of the high-grade squamous intraepithelial lesions, in 15.4% of the low-grade and in none of the normal tissues. The sensitivity, specificity, positive predictive value and negative predictive value for high-grade lesion were 92.3%, 100%, 100%, and 98.3%, respectively when considering p16(INK4a) expression, and 100%, 70.2%, 43.3% and 100%, respectively when considering high-risk HPV. Conclusions. p16(INK4a) test was better associated with high-grade intraepithelial lesion ( kappa = 0.95) than was the presence of high-risk HPV ( kappa = 0.47). Both tests could be complementary to high-grade squamous intra-epithelial lesion screening and help to define the diagnosis of the inconclusive low-grade/high-grade squamous intraepithelial lesion cases.8619498