Increasing the accuracy of self-sampling methods to detect oncogenic human papillomavirus (HPV) infection would contribute to the wider application of these approaches. In this study, 120 women were tested for HPV-16 by conventional and quantitative real-time PCR (QRT-PCR) in cervical and self-sampled vaginal and urine specimens. QRT-PCR had a higher detection rate, and the HPV viral load in all three sampling sites correlated with the severity of disease, as determined by histology. The vaginal and urine viral loads correlated with HPV-16 positivity according to both conventional and QRT-PCR, and were proportional to the cervical viral load.