禽网状内皮增生症病毒gp90基因真核表达及ELISA检测方法的建立  

Eukaryotic expression of reticuloendotheliosis virus gp90 protein and development of ELISA for detection of antibody against REV

作　　者：石星明[1] 赵妍[1] 王玫[1] 张晶[1] 魏秀英[1] 杨桂花[1] 高宏博[1] 全炎铭[1] 黄婷婷[1] 张晓艳[1] 崔红玉[1] 王云峰[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点试验室/禽传染病研究室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2010年第11期862-866,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：中央级公益性科研院所基本科研业务费(ZGKJ201003);黑龙江省杰出青年科学基金(JC200812)

摘　　要：为建立禽网状内皮增生症病毒(REV)的ELISA检测方法,本研究通过RT-PCR方法扩增gp90基因,构建杆状病毒重组质粒,转化DH10Bac后,通过转座获得重组杆粒,并将其转染Sf9昆虫细胞,SDS-PAGE及western blot分析表明,表达的重组蛋白分子量约为45 ku,具有良好的反应原性。将表达的重组蛋白纯化后包被ELISA反应板,建立检测REV抗体的rgp90-ELISA方法。特异性试验结果表明该方法具有良好的特异性;将该方法与IDEXX公司生产的试剂盒共同检测包括63份已知背景的血清在内的黑龙江省和广东省的临床血清样品共477份,符合率分别为90.5%和86.4%。该方法的建立为禽群REV抗体的监测提供了方便、快捷、准确的技术手段。An ELISA method for detecting antibodies against Avian reticuloendotheliosis virus（REV） was developed using recombinant gp90 protein and monoclonal antibody against chickens IgG.The recombinant gp90 protein was prepared by expressing the gp90 gene in a baculovirus expression system and confirmed by SDS-PAGE and western blot.The ELISA was highly specific and showed no cross-reaction with the positive sera of other avian diseases.Test on 63 positive serum samples and 477 clinic serum samples showed that this assay had a coincidence rate of 86.4% to 90.5% with IDEXX commercial kit.This ELISA method was simple to perform and suitable for large scale surveys of REV infection.

关 键 词：禽网状内皮组织增生症病毒 GP90基因 真核表达 ELISA 

分 类 号：S852.65[农业科学—基础兽医学]