Journal
JOURNAL OF VIROLOGICAL METHODS
Volume 204, Issue -, Pages 73-80Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jviromet.2014.04.010
Keywords
HPV; HPV genotyping; Multiplex PCR; Luminex; Cervical cancer
Funding
- Medical & Biological Laboratories (MBL), Tokyo, Japan
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Regional differences in human papillomavirus (HPV) genotypes and the presence of mixed HPV infections may affect adversely the efficacy of the HPV vaccine. Therefore, a simple and high-throughput HPV genotyping system is required. Recently, a novel HPV genotyping kit (the MebgenTm HPV kit) was developed. This kit uses multiplex PCR and Luminex xMAPTm technology to detect 13 types of high-risk HPVs and an internal control in a 96-well format. In the present study, the analytical performance of the kit was examined using HPV plasmid DNA. All 13 types of HPVs were detected with a minimum detection sensitivity of 250 copies/test, and highly specific signals were observed. HPV 16 plasmid was detected in samples containing mixtures with other HPV-type plasmids in ratios ranging from 1:1 to 1:1000. No cross reactivity was observed with DNA from 27 types of other infectious microbes. A clinical evaluation was carried out using cervical samples from 356 patients with persistent abnormal smears diagnosed at mass public health screenings for cervical cancer. The samples were preserved in TacasTm medium until analysis. HPV was detected in 162 (45.5%) samples including 110 (67.9%) with single infections and 52 (32.1%) with multiple infections. The type distribution of the 13 high-risk HPVs was as follows: 28.4% HPV 16, 11.7% HPV 18, 6.8% HPV 31,3.1% HPV 33,3.7% HPV 35,9.3% HPV 39, 1.9% HPV 45, 8.6% HPV 51, 37.0% HPV 52, 9.3% HPV 56, 16.7% HPV 58, 3.7% HPV 59, and 1.9% HPV 68. To evaluate sample stability over time, changes in the detection of HPV DNA derived from HeLa and SiHa cells were measured in 3 types of liquid-based cytology media. HPV DNA was detected in Tacas and ThinprepTm samples after storage at 4 C or 30 C for 4 weeks and within 1 week of collection in SurepathTM samples. These results suggest that this newly developed HPV genotyping kit is suitable for use in both clinical applications and large-scale epidemiological studies. (C) 2014 The Authors. Published by Elsevier B.V.
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