期刊
PLOS ONE
卷 16, 期 4, 页码 -出版社
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0250516
关键词
-
资金
- Federal funds from the Department of Health and Human Services
- Office of the Assistant Secretary for Preparedness and Response
- Biomedical Advanced Research and Development Authority [HHSO100201600015C]
The development of a high-throughput plate-based Zika virus reporter virus particle assay allows for quantitative measurement of virus neutralizing antibody titers in serum samples and shows high levels of precision and reproducibility. The assay uses infective, non-replicating particles encoding Zika virus surface proteins and capsid, and a reporter gene, and demonstrate close correlation with results from a plaque reduction neutralization test.
Zika virus is a Flavivirus, transmitted via Aedes mosquitos, that causes a range of symptoms including Zika congenital syndrome. Zika has posed a challenging situation for health, public and economic sectors of affected countries. To quantitate Zika virus neutralizing antibody titers in serum samples, we developed a high throughput plate based Zika virus reporter virus particle (RVP) assay that uses an infective, non-replicating particle encoding Zika virus surface proteins and capsid (CprME) and a reporter gene (Renilla luciferase). This is the first characterization of a Zika virus RVP assay in 384-well format using a Dengue replicon Renilla reporter construct. Serially diluted test sera were incubated with RVPs, followed by incubation with Vero cells. RVPs that have not been neutralized by antibodies in the test sera entered the cells and expressed Renilla luciferase. Quantitative measurements of neutralizing activity were determined using a plate-based assay and commercially available substrate. The principle of limiting the infection to a single round increases the precision of the assay measurements. RVP log(10)EC(50) titers correlated closely with titers determined using a plaque reduction neutralization test (PRNT) (R-2>95%). The plate-based Zika virus RVP assay also demonstrated high levels of precision, reproducibility and throughput. The assay employs identical reagents for human, rhesus macaque and mouse serum matrices. Spiking studies indicated that the assay performs equally well in different species, producing comparable titers irrespective of the serum species. The assay is conducted in 384-well plates and can be automated to simultaneously achieve high throughput and high reproducibility.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据