农业生物技术学报2021，Vol.29Issue(10)：2051-2060,10.DOI:10.3969/j.issn.1674-7968.2021.10.019
抗猪伪狂犬病毒gE蛋白纳米抗体的表达及功能验证
Expression and Functional Verification of Nano-antibody Against Porcine Pseudorabies virus gE Protein
摘要
Abstract
The outbreak of pseudorabies poses a huge challenge to its prevention and control. The establishment of a new diagnostic method with high specificity and sensitivity and the acquisition of antibodies with virus neutralizing effects are essential for the prevention, control and treatment of the disease. The purpose of this study was to express a soluble nano-antibody against the envelope glycoprotein E (gE) protein of porcine (Sus scrofa) Pseudorabies virus (PRV) in prokaryotic cells and to explore the application of this antibody in the diagnosis and treatment of porcine pseudorabies. In this study, the gene sequence of anti-porcine pseudorabies gE protein antibody was designed and synthesized, which was named PRVgENb, and PRVgENb-pET21b vector was constructed. The fusion protein was expressed in Escherichia coli. After purification on a Ni2+ affinity chromatography column, the single nano-antibody PRVgENb was obtained, which was labeled with HRP for direct ELISA and PRV virus neutralization test. The results showed that the fusion protein PRVgENb had high purity after prokaryotic expression and purification. Direct ELISA results showed that under a certain concentration of antigen, a low concentration of nano-antibody PRVgENb bound to the specific antigen with high activity. While a certain concentration of antibody could sensitively recognize low to high concentrations of specific antigen. The results of the virus neutralization test showed that the nano-antibody PRVgENb inhibited the replication of PRV and had a certain neutralization activity. This experiment established a method to stably obtain the nanobody PRVgENb, and initially explored its use in direct ELISA, and proved that it can inhibit the replication of PRV. Nanobody PRVgENb lays an experimental foundation for the diagnosis, treatment and follow-up research of PRV.关键词
猪伪狂犬病毒/gE蛋白/纳米抗体/直接ELISA/中和抗体Key words
Porcine pseudorabies virus/gE protein/Nano-antibody/Direct ELISA/Neutralizing antibody分类
农业科技引用本文复制引用
郭玉堃,马英先,明胜利,郭瑞珍,杨国宇,郭豫杰..抗猪伪狂犬病毒gE蛋白纳米抗体的表达及功能验证[J].农业生物技术学报,2021,29(10):2051-2060,10.基金项目
国家重点研发计划项目(2017YFD0501100) （2017YFD0501100）
