A sensitive and specific antigen detection assay for Middle East respiratory syndrome coronavirus
A sensitive and specific antigen detection assay for Middle East respiratory syndrome coronavirus
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DOI：
10.1038/emi.2015.26
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发表时间：
2015-04-22
影响因子：
13.2
通讯作者：
Woo, Patrick C. Y.
中科院分区：
文献类型：
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作者：
Chen, Yixin;Chan, Kwok-Hung;Woo, Patrick C. Y.
自2012年出现以来，已报告了900多例实验室确诊的中东呼吸综合征（MERS）病例，死亡率超过30%。然而，没有用于商业用途的抗原检测测定可用于诊断。本研究克隆了中东呼吸综合征冠状病毒（MERS-CoV）核衣壳蛋白（NP）基因全长，并在大肠杆菌中进行了表达。开发了使用产生的两种MERS-CoV-NP特异性单克隆抗体（MAbs）的MERS-CoV NP捕获酶联免疫吸附测定（ELISA）。使用129例各种呼吸道病毒阳性的鼻咽抽吸物（NPA）和通过添加MERS CoV系列稀释液模拟阳性NPA评价ELISA。使用0.19的截止OD，所有129个呼吸道病毒阳性的NPA显示出非常低的OD，特异性为100%。对于两种模拟的MERS-CoV阳性NPA（采用活MERS-CoV的系列稀释液），所有>= 50%组织培养感染剂量（TCID 50）/0.1 mL的样本均显示阳性结果。对于添加20和200 TCID 50/0.1 mL活MERS-CoV的另外10个NPA，均为阳性。一个高度敏感和特异性的单克隆抗体为基础的抗原捕获ELISA已开发的MERS。这种敏感和特异性的抗原捕获ELISA应该是有用的检测MERS冠状病毒在人类和单峰骆驼和在现场研究。
Since its emergence in 2012, more than 900 laboratory-confirmed cases of Middle East respiratory syndrome (MERS) have been reported with a fatality rate of more than 30%. However, no antigen detection assay for commercial use is available for diagnosis. In this study, the full-length nucleocapsid protein (NP) gene of MERS coronavirus (MERS-CoV) was cloned and expressed in Escherichia coli. A MERS-CoV NP capture enzyme-linked immunosorbent assay (ELISA) using two MERS-CoV-NP-specific monoclonal antibodies (MAbs) generated was developed. The ELISA was evaluated using 129 nasopharyngeal aspirates (NPAs) positive for various respiratory viruses and simulated positive NPAs by adding serial dilutions of MERS-CoV. Using a cutoff OD of 0.19, all 129 NPAs positive for respiratory viruses showed very low OD, with a specificity of 100%. For the two simulated MERS-CoV-positive NPAs with serial dilutions of live MERS-CoV, all samples with >= 50% tissue culture infective dose (TCID50)/0.1 mL showed positive results. For the 10 additional NPAs with 20 and 200 TCID50/0.1 mL of live MERS-CoV added, all were positive. A highly sensitive and specific MAbs-based antigen capture ELISA has been developed for MERS. This sensitive and specific antigen capture ELISA should be useful for detection of MERS-CoV in human and dromedaries and in field studies.