应用猪圆环病毒2型缺失NLS的Cap蛋白建立一种ELISA检测方法  

Development of an ELISA based on the NLS-deleted capsid protein of porcine circovirus type 2

作　　者：张春雷[1] 陈春丽[2] 王强[3] 叶昱[1] 张杰[1] 谢康尚 靳立明[1] 廖明[1] 樊惠英[1] 

机构地区：[1]华南农业大学农业部兽用疫苗创制重点实验室兽医学院,广东广州510642 [2]广东大华农动物保健品股份有限公司,广东新兴527400 [3]广东华农温氏畜牧股份有限公司,广东新兴527400

出　　处：《中国兽医杂志》2014年第3期14-17,共4页Chinese Journal of Veterinary Medicine

基　　金：国家高技术研究发展计划(863计划;2011AA10A209);国家重点基础研究发展计划(973计划;2011CB504701);广州市"珠江科技新星专项"项目(2012J2200086);公益性行业(农业)科研专项经费项目(201303034)

摘　　要：本试验通过诱导重组质粒pCold-SUMO-dCap在大肠杆菌中高效表达,获得了可溶性表达的SUMO-dCap融合蛋白,以SUMO-dCap融合蛋白作为包被抗原,建立了一种检测猪圆环病毒2型抗体的ELISA方法.用该方法与商品化试剂盒对267份猪血清进行平行检测,两种方法的符合率为90.26％,该方法的特异性和灵敏性分别为93.33％、97.16％;与猪蓝耳病病毒、猪伪狂犬病病毒、猪乙型脑炎病毒、猪口蹄疫病毒和猪瘟病毒阳性参考血清无交叉反应;批内和批间变异系数分别为1.25％～6.37％和6.68％～13.58％.研究结果表明,本试验建立的ELISA方法特异性强、敏感性好、重复性高,为临床上PCV2血清抗体检测和PCV2流行病学调查提供了一种简便的血清学诊断方法,为商品化试剂盒开发奠定了良好的基础.An indirect ELISA was developed for detecting antibodies against porcine circovirus type 2 （PCV2） based on a recombinant fusion protein. The antigen protein, which was SUMO-fused capsid protein of PCV2 without nuclear localization signal （NLS）, named as SUMO-dCap, was produced by recombinant plasmid pCold-SUMO-dCap in Escherichia coli （E.coli）. To validate the assay, 267 serum samples from pigs were examined by the developed method and it was compared with commercial ELISA kit. The coincidence rate, sensitivity and specificity of the established ELISA method were 90.26%, 93.33% and 97.16%, respectively. The assay showed no cross-reactivity with the reference sera of PRRS ,PRV,PJEV,FMBV and CSFV. Repeatability tests showed that the coefficient of variation of serum samples within and among runs were 1.25% -6.37% and 6.68% - 13.58% respectively. The results suggest that this ELISA is simple to perform, and is a specific, sensitive and convenient method for routine serological diagnosis, epidemiological survey of PCV 2-associated diseases and evaluation of serum neutralizing antibodies against PCV 2. It may serves as a PCV2 ELISA diagnostic kit.

关 键 词：猪圆环病毒2型 CAP蛋白 间接ELISA 

分 类 号：S852.651[农业科学—基础兽医学]