Jungkind D, Direnzo S, Beavis K G, Silverman N S
Department of Pathology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
J Clin Microbiol. 1996 Nov;34(11):2778-83. doi: 10.1128/jcm.34.11.2778-2783.1996.
We evaluated the COBAS AMPLICOR (CA) PCR system (Roche Diagnostic Systems) designed for automated PCR amplification and detection of nucleic acids from infectious agents in clinical samples. The Roche AMPLICOR microwell plate (MWP) PCR was the reference method. CA amplifies target nucleic acid, captures the biotinylated amplification products by using magnetic particles coated with specific oligonucleotide probes, and detects the bound products colorimetrically. For Mycobacterium tuberculosis, the correlation of the results of CA tests with those of MWP tests was 100% with 230 samples, including 20 culture-positive samples. For hepatitis C virus, the correlation was 100% with 214 samples, including 60 positive samples. MultiPlex CA analysis of 199 cervical specimens for Chlamydia trachomatis, Neisseria gonorrhoeae, and the internal control gave 100% concordance. These samples included 19 C. trachomatis and 3 N. gonorrhoeae culture-positive samples. Overall, the agreement between PCR methods for all 842 comparisons was 100%. Compared with culture, the sensitivities of the assays for C. trachomatis and M tuberculosis were > or = 95%. After spiking alternating amplification tubes in the CA system with 10(14) copies of the Chlamydia amplicon per ml, we were unable to demonstrate any carryover cross-contamination of negative samples. Using the criteria of the College of American Pathologists workload recording method, we found that the total hands-on time to produce CA PCR results was 4.4, 7.9, and 3.3 min for M. tuberculosis, hepatis C virus, and the MultiPlexed assay for chlamydia plus gonorrhea and an internal control, respectively. The CA system brings true PCR automation to laboratories. In addition to the accuracy of automated results, the CA system provides labor savings, provides containment of the amplification and detection components of PCR, and supports both MultiPlex amplification and sequential algorithm (ReFlex) detection of analytes.
我们评估了COBAS AMPLICOR（CA）PCR系统（罗氏诊断系统公司），该系统设计用于临床样本中感染因子核酸的自动化PCR扩增和检测。罗氏AMPLICOR微孔板（MWP）PCR为参考方法。CA系统通过特异性寡核苷酸探针包被的磁珠捕获生物素化的扩增产物，并通过比色法检测结合产物。对于结核分枝杆菌，230份样本（包括20份培养阳性样本）的CA检测结果与MWP检测结果的相关性为100%。对于丙型肝炎病毒，214份样本（包括60份阳性样本）的相关性为100%。对199份宫颈标本进行沙眼衣原体、淋病奈瑟菌及内部对照的多重CA分析，结果一致性为100%。这些样本包括19份沙眼衣原体和3份淋病奈瑟菌培养阳性样本。总体而言，所有842次比较中PCR方法之间的一致性为100%。与培养法相比，沙眼衣原体和结核分枝杆菌检测的灵敏度≥95%。在CA系统的交替扩增管中加入每毫升10(14)拷贝的沙眼衣原体扩增子后，我们未能证明阴性样本存在任何残留交叉污染。按照美国病理学家学会工作量记录方法的标准，我们发现结核分枝杆菌、丙型肝炎病毒以及沙眼衣原体、淋病奈瑟菌和内部对照的多重检测产生CA PCR结果的总实际操作时间分别为4.4分钟、7.9分钟和3.3分钟。CA系统为实验室带来了真正的PCR自动化。除了自动化结果的准确性外，CA系统还节省人力，将PCR的扩增和检测组件封闭起来，并支持多重扩增和分析物的顺序算法（ReFlex）检测。
