A rapid magnetic particle-based enzyme immunoassay for human cytomegalovirus glycoprotein B quantification
A rapid magnetic particle-based enzyme immunoassay for human cytomegalovirus glycoprotein B quantification
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DOI：
10.1016/j.jpba.2018.04.019
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发表时间：
2018-07-15
影响因子：
3.4
通讯作者：
Castillo, Juan R.
中科院分区：
文献类型：
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作者：
Pires, F.;Julia Arcos-Martinez, M.;Castillo, Juan R.
人巨细胞病毒（HCMV）是一种疱疹病毒，可引起严重感染。然而，现有的诊断方法的主要缺点是需要长时间进行。本文提出了一种简单的磁性颗粒酶免疫分析法（mpEIA），用于尿液中人巨细胞病毒糖蛋白B（gB-HCMV）的定量检测。免疫传感器方案基于夹在初级单克隆抗体之间的分析物蛋白gB-HCMV，（MBsPrG-mAbl），和用辣根过氧化物酶（Ab 2-HRP）标记的第二抗gB-HCMV抗体以允许分光光度检测。在90 - 700 pg/mL（-1）浓度范围内，gB-HCMV浓度与450 nm处的吸光度信号呈线性关系。尿液中gB-HCMV的计算检测限为90 +/- 2 pg/mL（-1），RSD约为6.7%。该免疫传感器对疱疹病毒科的其他病毒，即水痘带状疱疹病毒和爱泼斯坦巴尔病毒表现出良好的选择性。在0.30-0.50 ng/mL（-1）gB-HCMV浓度水平下，加标人尿液样本的回收率范围为91%-105%。根据欧洲药品管理局（EMEA-2014）的指导原则，对拟定的mpEIA方法进行了验证，可以快速、成功和轻松地定量尿液样本中的gB-HCMV。(C)2018爱思唯尔B. V.保留所有权利。
Human cytomegalovirus (HCMV) is a herpes virus that can cause severe infections. Still, the available methods for its diagnostic have the main disadvantage of requiring long time to be performed. In this work, a simple magnetic particle-based enzyme immunoassay (mpEIA) for the quantification of glycoprotein B of Human cytomegalovirus (gB-HCMV) in urine samples is proposed. The immunosensor scheme is based on the analyte protein gB-HCMV sandwiched between a primary monoclonal antibody, (MBsPrG-mAbl), and a secondary anti-gB-HCMV antibody labelled with Horseradish peroxidase (Ab2-HRP) to allow spectrophotometric detection.The mpEIA analytical performance was tested in urine samples, showing a linear dependence between gB-HCMV concentration and the absorbance signal at 450 nm in a range of concentrations from 90 to 700 pg mL(-1). The calculated detection limits for gB-HCMV were 90 +/- 2 pg mL(-1) and the RSD was about 6.7% in urine samples. The immunosensor showed good selectivity against other viruses from Herpesviridae family, namely varicella zoster and Epstein Barr viruses. The recoveries of spiked human urine samples at 0.30-0.50 ng mL(-1) concentration levels of gB-HCMV ranged between 91 to 105%. The proposed mpEIA method was validated following the guidelines of the European Medicines Agency (EMEA-2014), and allows rapid, successful and easy quantification of gB-HCMV in urine samples. (C) 2018 Elsevier B.V. All rights reserved.