Department of Laboratory Medicine, University of Ulsan College of Medicine and Asan Medical Center, Asanbyeongwon-gil 86, Songpa-gu, Seoul 138-736, South Korea.
J Virol Methods. 2009 Dec;162(1-2):208-12. doi: 10.1016/j.jviromet.2009.08.007. Epub 2009 Aug 25.
The objective of this study was to compare the performance of two automated nucleic acid extraction systems. Specifically, the NucliSens easyMAG system (bioMerieux, Marcy l'Etoile, France), which incorporates magnetic bead technology, was compared with the BioRobot MDx system (Qiagen GmbH, Hilden, Germany), which uses a silica membrane-based method of nucleic acid extraction. Nucleic acids from the BK virus (BKV DNA) were extracted from 98 plasma and 57 urine specimens using the Real-Q BKV quantitation kit (Biosewoom, Seoul, Korea). Failed PCR was defined as negative BKV DNA results having more than 36 threshold cycles of the internal control by the manufacturer's instruction. The PCR failure rate of nucleic acids isolated from plasma samples using the MDx system was similar to that of plasma samples processed using the easyMAG system (2.0% and 3.1%, respectively). The PCR failure rate of nucleic acids isolated from urine samples using the MDx system was higher than that of urine samples processed using the easyMAG system (33.3% and 12.5%, respectively). These data suggest that the PCR inhibitors present in urine specimens are removed more efficiently by the easyMAG system. Among amplified specimens, the discordant results obtained from the two systems revealed that the BKV DNA load ranged from 2.3 log10 copies/mL to 4.6 log10 copies/mL. Of the 25 urine specimens that yielded BKV DNA by both extraction systems, 15 specimens (60.0%) yielded higher BKV DNA loads by the easyMAG system, indicating that the easyMAG system extracted nucleic acid more efficiently than did the MDx system. In conclusion, the easyMAG method outperformed the MDx method when used to extract BKV DNA from urine samples. Magnetic bead-based extraction methods such as the easyMAG system are therefore preferable for the quantitation of viral DNA in urine.
本研究旨在比较两种自动化核酸提取系统的性能。具体而言，比较了采用磁珠技术的 NucliSens easyMAG 系统（bioMerieux，Marcy l'Etoile，法国）与采用基于硅胶膜的核酸提取方法的 BioRobot MDx 系统（Qiagen GmbH，Hilden，德国）。使用来自韩国 Biosewoom 的 Real-Q BKV 定量试剂盒（Biosewoom，首尔）从 98 份血浆和 57 份尿液标本中提取 BK 病毒（BKV DNA）的核酸。根据制造商的说明，将 PCR 失败定义为 BKV DNA 结果的内部对照超过 36 个阈值循环的阴性。使用 MDx 系统从血浆样本中分离的核酸的 PCR 失败率与使用 easyMAG 系统处理的血浆样本相似（分别为 2.0%和 3.1%）。使用 MDx 系统从尿液样本中分离的核酸的 PCR 失败率高于使用 easyMAG 系统处理的尿液样本（分别为 33.3%和 12.5%）。这些数据表明，尿液标本中存在的 PCR 抑制剂通过 easyMAG 系统更有效地去除。在扩增的标本中，两种系统获得的不一致结果表明，BKV DNA 载量范围为 2.3 log10 拷贝/mL 至 4.6 log10 拷贝/mL。在两种提取系统均产生 BKV DNA 的 25 份尿液标本中，15 份标本（60.0%）通过 easyMAG 系统获得更高的 BKV DNA 载量，表明 easyMAG 系统比 MDx 系统更有效地提取核酸。总之，当从尿液样本中提取 BKV DNA 时，easyMAG 方法优于 MDx 方法。因此，对于尿液中病毒 DNA 的定量，基于磁珠的提取方法，如 easyMAG 系统，是首选方法。
