The performance of the HPV16 real-time PCR integration assay

ObjectivesIntegration of high risk human papillomavirus (HPV) into the host genome leads to viral oncogene deregulation predisposing to neoplastic progression. Integration can be detected from pap smear or biopsy and use as marker of progressive disease.Design and methodsWe have previously developed a highly sensitive real-time PCR method to determine HPV integration frequency and viral load of HPV16 in clinical samples. The test is accurate and sensitive detecting approx. 50 copies of integrated HPV in the sample.ResultsWe found that a tenfold excess of episomal form to integrated form interferes with the test, regardless the amount of viral DNA. The same was true with background DNA more than 1500 ng in reaction.ConclusionsOverall, this method is reproducible and suitable for high-throughput screening of clinical samples, but excess episomal copies might mask the integrated form.