用于直接从血液中进行埃博拉病毒即时检测的分子诊断现场试验
Molecular Diagnostic Field Test for Point-of-Care Detection of Ebola Virus Directly From Blood.
作者信息
Benzine Jason W, Brown Kerry M, Agans Krystle N, Godiska Ronald, Mire Chad E, Gowda Krishne, Converse Brandon, Geisbert Thomas W, Mead David A, Chander Yogesh
机构信息
Lucigen Corp., Middleton, Wisconsin.
Department of Microbiology and Immunology Galveston National Laboratory, University of Texas Medical Branch at Galveston.
出版信息
J Infect Dis. 2016 Oct 15;214(suppl 3):S234-S242. doi: 10.1093/infdis/jiw330. Epub 2016 Sep 16.
Abstract
A molecular diagnostic method for robust detection of Ebola virus (EBOV) at the point of care (POC) directly from blood samples is described. This assay is based on reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) of the glycoprotein gene of EBOV. Complete reaction formulations were lyophilized in 0.2-mL polymerase chain reaction tubes. RT-LAMP reactions were performed on a battery-operated isothermal instrument. Limit of detection of this RT-LAMP assay was 2.8 × 10 plaque-forming units (PFU)/test and 1 × 10 PFU/test within 40 minutes for EBOV-Kikwit and EBOV-Makona, respectively. This assay was found to be specific for the detection of EBOV, as no nonspecific amplification was detected in blood samples spiked with closely related viruses and other pathogens. These results showed that this diagnostic test can be used at the point of care for rapid and specific detection of EBOV directly from blood with high sensitivity within 40 minutes.
摘要
本文描述了一种分子诊断方法，可直接从血样中在护理点（POC）对埃博拉病毒（EBOV）进行可靠检测。该检测方法基于对EBOV糖蛋白基因的逆转录环介导等温扩增（RT-LAMP）。完整的反应配方冻干在0.2 mL聚合酶链反应管中。RT-LAMP反应在电池供电的等温仪器上进行。对于EBOV-Kikwit和EBOV-Makona，该RT-LAMP检测的检测限分别为2.8×10噬斑形成单位（PFU）/测试和40分钟内1×10 PFU/测试。该检测方法被发现对EBOV的检测具有特异性，因为在添加了密切相关病毒和其他病原体的血样中未检测到非特异性扩增。这些结果表明，该诊断测试可在护理点用于在40分钟内直接从血液中快速、特异性且高灵敏度地检测EBOV。
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