Development of an indirect ELISA for serological detection of reticuloendotheliosis virus using the gp90 protein expressed in Pichia pastoris

文献类型： 外文期刊

第一作者： Li, Kai

作者： Li, Kai;Gao, Honglei;Gao, Li;Qi, Xiaole;Gao, Yulong;Qin, Liting;Wang, Yongqiang;Wang, Xiaomei

作者机构：

关键词： Reticuloendotheliosis virus;gp90 protein;Indirect ELISA;Pichia pastoris

期刊名称：JOURNAL OF VIROLOGICAL METHODS （ 影响因子：2.014； 五年影响因子：2.001 ）

ISSN：

年卷期：

页码：

收录情况： SCI

摘要： The present study was undertaken to express the gp90 protein of reticuloendotheliosis virus (REV) in Pichia pastoris and evaluate its potential use as a diagnostic antigen in ELISA. The full-length gp90 gene of REV was cloned into pPIC9k vector and then integrated into the chromosome of P. pastoris for induced expression. SDS-PAGE and western blot assay demonstrated that gp90 protein was expressed and secreted into the culture medium at about 100 mg/L of culture under optimized condition. An indirect ELISA was then established by using the recombinant gp90 protein as the coating antigen. The optimal concentration of coated antigen was 0.1 mu g/well at a serum dilution of 1:200 and the optimal positive threshold value of the assay was 0.409. Cross-reactivity assay showed that this antigen was REV specific. The reproducibility experiment displayed good consistency. Furthermore, the gp90 protein based indirect ELISA showed good correlation rates of 96.3% and 97.5% with virus neutralization test and a commercially whole virus based indirect ELISA, respectively. This study demonstrates the efficacy of recombinant gp90 protein as an antigen in ELISA for seroepidemiological study of REV infection on a large scale. (C) 2011 Elsevier B.V. All rights reserved.

分类号： R37

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