Development of a robust, higher throughput green fluorescent protein (GFP)-based Epstein-Barr Virus (EBV) micro-neutralization assay.
J Virol Methods
; 247: 15-21, 2017 09.
Article
em En
| MEDLINE
| ID: mdl-28457783
The goal of most prophylactic vaccines is to elicit robust and effective neutralizing antibodies against the human pathogen target. The titer of neutralizing antibodies to Epstein-Barr Virus (EBV) is a useful biomarker for evaluating EBV vaccines. Here, the development and optimization of a 96-well micro-neutralization fluorescent imaging assay (FIA) using an EBV virus-encoding green fluorescent protein (GFP) to infect adherent EBV recipient cells is reported. The conditions were optimized for generating reproducible EBV-GFP virus, for maintaining viral infectivity for months, and for efficient viral infection of recipient cell culture. The utility of the EBV-GFP FIA neutralization assay was demonstrated in a mouse study of an investigational adjuvanted EBV gp350 subunit vaccine. This assay confirmed the generation of high titers of anti-EBV-neutralizing antibodies which correlated well with the established Raji cell-based flow cytometry-based EBV neutralization assay, as well as with anti-gp350 IgG titers. In naturally infected EBV+ human serum samples, a good correlation between anti-gp350 IgG ELISA titer and EBV-GFP FIA neutralization antibody titer was also observed. Taken together, these results demonstrate the establishment of a scalable high throughput EBV-GFP FIA micro-neutralization assay suitable to measure humoral EBV vaccine response in a large-scale human trial.
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MEDLINE
Assunto principal:
Testes de Neutralização
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Herpesvirus Humano 4
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Proteínas de Fluorescência Verde
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Ensaios de Triagem em Larga Escala
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Anticorpos Antivirais
Limite:
Animals
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article