Hybrid Capture II (HC-II) is a commercial human papillomavirus (HPV) detection test designed to detect 18 HPV types divided into high-risk and low-risk groups. We have tested 1647 scrapes from 1518 unselected women attending routine cytological screening by this assay for the detection of histologically proven high-grade lesions. The reliability of this test was also evaluated on 117 fresh cone biopsy samples. HPV DNA has been detected in 400 scrapes (24.3%), 296 containing a high-risk HPV (18.0%). All the smears evocative of high-grade lesions were positive for high-risk HPV, and high-risk HPV were detected in all the 34 cases presenting a histologically proven high-grade lesion and in 68 (97.1%) of the 70 cone biopsy samples showing a high-grade lesion or an invasive carcinoma. Thus, the sensitivity was superior to the sensitivity of cytology (85.3%). Nevertheless, the quantitative approach provided by the HC-II assay for the assessment of the viral load could not clearly distinguish among cases with or without high-grade lesions. Thus this assay is recommended for the screening of high-grade lesions on a large scale, in association with classic cytology.