简套式酶免RT-PCR检测患者样本中SARS冠状病毒

为建立一种快速敏感的SARS冠状病毒核酸酶免PCR检测方法,根据SARS冠状病毒基因序列,自行设计引物、杂交探针,优化反应体系和探针共价交联,采用简套式PCR扩增SARS-CoV的cDNA.其产物通过包被特异性探针的微孔板捕获杂交、比色检测,以构建的假病毒颗粒作为内对照,监控操作的全过程,同时,与现有的可靠SARS检测方法进行比较.通过对几种方法的临床检测比较,说明简套式PCR检测方法是SARS临床早期诊断快速、敏感、重复性好的有效方法.

A Simplified Method of Nested RT-PCR ELISA for the Detection of SARS-CoV

Various methods are available for the detection of SARS-CoV. Real time RT-PCR is the most widely used method; however, its application has been critically restricted in less developed areas due to the high cost of equipment and reagents. The objective of this study is to develop a detection system with sufficient sensitivity for the early detection of SARS-CoV, which will be significantly less expensive. A simplified nested PCR was used to amplify SARS-CoV cDNA. The product was captured by spe cifie probes pre-coated on microplates before colorimetric detection. Pseudo viral particles developed by our lab were applied as an internal control to monitor the entire detection process. This method was tested with clinical samples, and compared with other assays that are already available for SARS detection. Tests with clinical specimens and comparison with several other assays showed that the system developed in our lab was specilic, highly sensitive and satisfactorily reproducible for SARS-CoV. This method, essentially a nucleotide acid based detection assay, is a highly sensitive and cost saving method for the early detection of SAR-CoV.