MGB-based one-step multiplex real-time PCR method for rapid detection of HPV

Abstract

For full-scale analysis of Human Papillomavirus (HPV) status in humans, two minor groove binder (MGB)-based one-step multiplex real-time PCR systems were developed: one to screen 16 high-risk HPV (HR-HPV) types, and one to screen a broader spectrum of HPV types (common HPV or C-HPV). Sensitivity and specificity were evaluated using diluted reference plasmids and 20 control human DNA samples. For clinical evaluation, 510 cervical scrape samples were evaluated. The sensitivity assays revealed that the C-HPV detection system could detect 10 ~ 100 plasmid copies/reaction, while the HR-HPV detection system detected 10 ~ 500 copies. The specificity test revealed that the systems did not yield positive signals from the 20 human genomic DNA samples. Performance was tested on 510 usable clinical samples. The HR-HPV results were compared to those from the Hybrid Capture 2 (HC2) test, which assesses 13 HR-HPV types; the concordance level between the two methods was 90.8% with a kappa value of 0.813. These results showed that our novel MGB-based one-step multiplex real-time PCR method may be used for the diagnosis and mass screening of HPV in clinical and large-scale epidemiological studies.