An Optimized High-Throughput Neutralization Assay for Hepatitis E Virus (HEV) Involving Detection of Secreted Porf2
An Optimized High-Throughput Neutralization Assay for Hepatitis E Virus (HEV) Involving Detection of Secreted Porf2
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针对戊型肝炎病毒 (HEV) 的优化高通量中和试验，涉及分泌型 Porf2 的检测
DOI：
10.3390/v11010064
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发表时间：
2019-01
期刊：
影响因子：
4.7
通讯作者：
Xia Ningshao
中科院分区：
文献类型：
--
作者：
Liu Chang;Cai Wei;Yin Xin;Tang Zimin;Wen Guiping;Ambardekar Charuta;Li Xinlei;Ying Dong;Feng Zongdi;Zheng Zizheng;Xia Ningshao
戊型肝炎病毒(HEV)是世界范围内急性肝炎的常见原因。目前用于评价HEV特异性抗体中和活性的方法包括免疫荧光焦点分析(IFAS)和实时荧光定量聚合酶链式反应(Real-time PCR)，这两种方法不敏感且操作复杂。在这里，我们开发了一种高通量中和试验，方法是使用HEV抗原酶联免疫吸附试验(ELISA)试剂盒，检测分泌的pORF2水平，该试剂盒基于高复制的HEV基因型(GT)3毒株Kernow。我们同时用传统的IFA和新的方法评估了HEV特异性抗体和免疫个体(n=15)血清的中和活性。线性回归分析表明，两种方法之间存在高度的相关性。此外，抗-HEV-Ig G水平与免疫个体血清中和滴度呈中度相关，提示GT-1免疫可预防GT-3 Kernow感染。然后用27份抗-HEV-Ig G阳性血清和23份抗-HEV-Ig G阴性血清测定了新方法的特异性和中和能力的潜在阈值。抗-HEV-Ig G阳性血清中和能力明显强于抗-HEV-Ig G阴性血清。ROC曲线分析表明，血清中和能力的潜在阈值为8.07，灵敏度为88.6%，特异度为100%。我们的结果表明，用抗原ELISA试剂盒进行中和试验可以作为HEV临床研究的有用工具。
Hepatitis E virus (HEV) is a common cause of acute hepatitis worldwide. Current methods for evaluating the neutralizing activity of HEV-specific antibodies include immunofluorescence focus assays (IFAs) and real-time PCR, which are insensitive and operationally complicated. Here, we developed a high-throughput neutralization assay by measuring secreted pORF2 levels using an HEV antigen enzyme-linked immunosorbent assay (ELISA) kit based on the highly replicating HEV genotype (gt) 3 strain Kernow. We evaluated the neutralizing activity of HEV-specific antibodies and the sera of vaccinated individuals (n = 15) by traditional IFA and the novel assay simultaneously. A linear regression analysis shows that there is a high degree of correlation between the two assays. Furthermore, the anti-HEV IgG levels exhibited moderate correlation with the neutralizing titers of the sera of vaccinated individuals, indicating that immunization with gt 1 can protect against gt 3 Kernow infection. We then determined specificity of the novel assay and the potential threshold of neutralizing capacity using anti-HEV IgG positive sera (n = 27) and anti-HEV IgG negative sera (n = 23). The neutralizing capacity of anti-HEV IgG positive sera was significantly stronger than that of anti-HEV IgG negative. In addition, ROC curve analysis shows that the potential threshold of neutralizing capacity of sera was 8.07, and the sensitivity and specificity of the novel assay was 88.6% and 100%, respectively. Our results suggest that the neutralization assay using the antigen ELISA kit could be a useful tool for HEV clinical research.
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影响因子：
12.7
作者：
Zhang, Feng;Li, Xiuhua;Wang, Youchun
通讯作者：
Wang, Youchun
影响因子：
64.8
作者：
Feng Z;Hensley L;McKnight KL;Hu F;Madden V;Ping L;Jeong SH;Walker C;Lanford RE;Lemon SM
通讯作者：
Lemon SM
影响因子：
9.4
作者：
Tanaka, Toshinori;Takahashi, Masaharu;Okamoto, Hiroaki
通讯作者：
Okamoto, Hiroaki
影响因子：
5.4
作者：
Yin, Xin;Ambardekar, Charuta;Feng, Zongdi
通讯作者：
Feng, Zongdi
ORF2分泌形式在戊型肝炎病毒感染中的起源、抗原性和功能
DOI：
10.1073/pnas.1721345115
发表时间：
2018-05-01
影响因子：
11.1
作者：
Yin, Xin;Ying, Dong;Feng, Zongdi
通讯作者：
Feng, Zongdi