猪瘟病毒E2蛋白夹心ELISA抗原检测方法的建立及初步应用

Establishment and preliminary application of ELISA detection method for swine fever virus E2 protein

关键词:  猪瘟病毒  E2蛋白  ELISA  检测

Key words:  Swine fever virus  E2 protein  ELISA  Test

摘要:

为了建立一种猪瘟病毒E2蛋白夹心ELISA抗原检测方法,以纯化的兔多抗作为包被用抗体,辣根过氧化物酶(HRP)标记鼠单抗作为检测用抗体,以大肠杆菌表达的猪瘟病毒E2蛋白作为标准品,建立了猪瘟病毒E2蛋白夹心ELISA抗原定量方法。结果表明,兔多抗最佳包被浓度为4 μg/mL,最佳封闭液为含1%BSA的PBS溶液,最佳反应时间为60分钟,最佳二抗稀释度为1:3200,反应时间为60分钟。建立的ELISA方法最低检出量为39 ng/ml,该方法与猪其他病原无交叉反应。建立的ELISA检测方法为猪瘟病毒及疫苗半成品检测奠定了基础,具有重要的应用价值。

Abstract:

In order to develop a swine fever virus E2 protein detection method. Rabbit polyclonal antibody was used as the coating antibody.Horseradish Peroxidase (HRP)-labeled monoclonal antibody was used as the detection antibody, and purified swine fever virus E2 protein was used as the standard. to establish The ELISA detection method was established for swine fever virus E2 protein. The results showed that the optimal coating concentration of rabbit polyclonal antibody was 4 μg/mL. The optimal blocking solution was PBS solution containing 1% BSA. The optimal reaction time was 60 minutes. The optimal secondary antibody dilution was 1:3200, and the reaction time was for 60 minutes. The minimum detectable amount of the established ELISA method is 39ng/ml, and this method has no cross-reaction with other swine pathogens. The established ELISA detection method lays the foundation for the quantification of semi-finished products of swine fever virus E2 protein subunit vaccine, and has important application value.