摘要
以原核表达的甘薯潜隐病毒(SPLV)的外壳蛋白(CP)为抗原免疫小鼠,经过细胞融合和亚克隆,筛选出2株稳定分泌抗SPLV CP的单克隆抗体杂交瘤细胞株(5B11-2和5G8-2),并分别制备了单克隆抗体腹水。间接ELISA结果表明,用SPLV CP包被酶联板,5B11-2和5G8-2单克隆抗体的效价均为1∶512 000;用感染SPLV的甘薯叶片汁液包被酶联板,2株单克隆抗体的效价均为1∶6 400。抗体类型及亚类鉴定结果表明,2株单克隆抗体均为IgG1、κ轻链。Western blot分析表明,2株单抗均能与SPLV CP和感染SPLV的甘薯叶片汁液有特异性反应。利用单克隆抗体建立的间接抗原包被ELISA(ACP-ELISA)检测SPLV方法,病叶1∶3 840倍稀释仍能检测到病毒。血清学和RT-PCR检测结果表明,制备的单克隆抗体可用于田间甘薯样品的检测。
In order to prepare the monoclonal antibodies（MAbs） against Sweet potato latent virus（SPLV）coat protein（CP）,spleen cells were isolated from BALB/c mouse immunized by the SPLV CP followed by fusing with mouse myeloma cells（SP2/0） and subcloning. Two hybridoma cell lines（5B11-2 and 5G8-2） stably secreting MAbs against SPLV CP were obtained and the titres of both MAbs were 1：512 000 and 1：6 400 when using the SPLV CP and diseased leaves,respectively,as coating antigen for the indirect ELISA assay.Isotypes and subclasses analysis of 5B11-2 and 5G8-2 indicated that both of them belong to IgG1,κ light chain.Western blot analysis further confirmed that both MAbs could specifically react with SPLV CP and sweet potato leaf extracts containing SPLV. ACP-ELISA could successfully detect virus in plant sap diluted by 3 840 fold.The consistency between serological and RT-PCR results showed that the MAbs produced in our study could be used for detecting SPLV in the field samples of sweet potato.
出处
《植物病理学报》
CAS
CSCD
北大核心
2017年第2期240-245,共6页
Acta Phytopathologica Sinica
基金
国家甘薯产业技术体系建设项目资助(CARS-11-B-07)
