A rapid and sensitive detection of HPV by combined assay of PCR and fluorescence DNA chip

A human papilloma virus (HPV) test can be used to identify women that are at risk for cervical cancer. Hybrid Capture (HC2) and PCR are the current gold standard assays and are used globally to detect HPV infection. We present a combined assay system that uses PCR and fluorescence (FL) DNA chip for simple, rapid, and sensitive HPV DNA detection. In the developed assay system, PCR-amplified DNA produced by biotin-labeled consensus GP5(+)/GP6(+) primers was denatured, neutralized, mixed with FL-streptavidin detector solution and then the mixture was loaded onto the HPV DNA chip for DNA hybridization. The DNA chip was inserted into the laser FL scanner and the FL signal was measured. MgCl2 of PCR mixture completely nullified FL signal in DNA chip assay but addition of 50 mM EDTA to detector solution recovered FL signal. When PCR product of HPV 16 or 18 infected patient's sample was subjected to DNA chip assay, FL signal was observed only from its corresponding HPV type DNA chip, demonstrating the HPV assay system differentiate different HPV genotypes. In rapidity and sensitivity test, the FL signal was illuminated from a PCR product produced by 5 cycles, which did not resolve as a DNA band in gel electrophoresis, suggesting that the combined assay system is more sensitive than an HPV PCR assay. Additionally, this test can be completed within 30 minutes, which will allow patients to be evaluated for infection.