Dunn S Terence, Allen Richard A, Wang Sophia, Walker Joan, Schiffman Mark
Department of Pathology, The University of Oklahoma Health Sciences Center, Oklahoma City, OK 73190, USA.
J Virol Methods. 2007 Jul;143(1):45-54. doi: 10.1016/j.jviromet.2007.02.006. Epub 2007 Mar 30.
Testing for the group of approximately 15 carcinogenic human papillomavirus (HPV) genotypes is an important adjunct to cytology. Because carcinogenic strengths of HPV types differ greatly, assays that permit identification of individual HPV genotypes are being introduced. Most HPV genotyping systems proposed for clinical use are PCR-based, depending heavily for validity on careful attention to numerous details. One understudied detail is the effect of different sample preparation methods including DNA extraction. This study examines the influence of DNA extraction on performance of a new PCR-based genotyping kit, the Roche LINEAR ARRAY HPV assay. When volume of sample extracted, DNA extraction methods and/or amount of DNA tested were varied, the HPV type results were reproducible for strong viral bands but not weak ones. Moreover, although the experiments were not comprehensive, they showed that the manufacturer-approved DNA extraction method might not be the best method for use in this assay. Because different "front end" protocols introduce variability into genotyping results, the authors urge laboratories not to vary methods for this assay without due consideration. The results suggest that companies carefully optimize DNA extraction methods prior to commercial introduction of their PCR-based genotyping assays destined for widespread clinical use.
对大约15种致癌性人乳头瘤病毒（HPV）基因型进行检测是细胞学检查的一项重要辅助手段。由于HPV各型的致癌强度差异很大，能够识别各HPV基因型的检测方法正在被引入。大多数提议用于临床的HPV基因分型系统都是基于聚合酶链反应（PCR）的，其有效性在很大程度上依赖于对众多细节的密切关注。一个研究较少的细节是包括DNA提取在内的不同样本制备方法的影响。本研究考察了DNA提取对一种新的基于PCR的基因分型试剂盒——罗氏线性阵列HPV检测法性能的影响。当提取的样本体积、DNA提取方法和/或检测的DNA量发生变化时，对于强病毒条带，HPV型别结果具有可重复性，但对于弱条带则不然。此外，尽管实验并不全面，但结果表明，厂家认可的DNA提取方法可能并非该检测法的最佳使用方法。由于不同的“前端”方案会给基因分型结果带来变异性，作者敦促实验室在进行该检测时，未经充分考虑不要改变方法。结果表明，各公司在将其用于广泛临床应用的基于PCR的基因分型检测法商业化推出之前，应仔细优化DNA提取方法。
