Automated high throughput DNA isolation for detection of human papillomavirus in oral rinse samples

作者：Broutian Tatevik R; He Xin; Gillison Maura L^*

来源：Journal of Clinical Virology, 2011, 50(4): 270-275.

DOI：10.1016/j.jcv.2010.12.005

• Human papillomavirus
• Oral rinse samples
• HPV detection
• Qiasymphony
• Puregene DNA isolation

Background: Oral HPV infection elevates risk of oropharyngeal cancer, but its natural history is unknown. Natural history studies necessitate validation of an automated, high-throughput method for HPV genomic DNA detection in oral rinse samples (ORS).
Objectives: To compare agreement of oral HPV detection in ORS processed by a magnetic-bead based automated platform to a previous gold-standard, manual protein-precipitation method. Agreement was compared to that of repeat sampling and repeat HPV testing.
Study design: HIV-infected individuals (n = 100) provided two ORS collected 15 min apart. DNA was isolated from equal aliquots by either a protein-precipitation based (Puregene, Qiagen) or magnetic bead-based (QIAsymphony (TM) SP, Qiagen) method. HPV DNA was detected and type-specified by consensus primer PCR and reverse line blot hybridization. The kappa statistic was used to assess overall agreement (OA) and agreement on a positive test (Ps+).
Results: The DNA purification methods had very high agreement for categorizing an individual as HPV infected (OA = 0.95; Ps+ = 0.94) as well as for detection of HPV type-specific infection (OA = 0.99; Ps+ = 0.88) in ORS. Agreement for detection of HPV type-specific infection was greater than that observed with repeat oral rinse sampling (OA = 0.99, Ps+ = 0.76) but comparable to inter-assay agreement (OA = 1.00, Ps+ = 0.90).
Conclusions: HPV detection in ORS processed with a magnetic-bead based automated platform will facilitate large natural history studies of oral HPV infection necessary to evaluate the potential use of oral HPV detection in oral cancer screening.

• 出版日期2011-4