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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。
The murine cells were immunized with coat protein (CP) of prokaryotic latent virus (SPLV) as antigen and two hybridoma cell lines stably secreting SPLV CP were screened by cell fusion and subcloning (5B11- 2 and 5G8-2), and monoclonal antibodies were prepared ascites. The results of indirect ELISA showed that the titer of the monoclonal antibody against 5B11-2 and 5G8-2 was 1: 512000 when the SPLV CP was coated on the enzyme-linked immunosorbent assay (ELISA), and the enzyme-linked immunosorbent assay (ELISA) The titer of the cloned antibody was 1: 400. Antibody type and subclass identification results showed that the two monoclonal antibodies were IgG1, κ light chain. Western blot analysis showed that both McAbs reacted specifically with SPLV CP and sweet potato leaf juice infected with SPLV. Indirect antigen-coated ELISA (ACP-ELISA) was used to detect SPLV by monoclonal antibody. The virus was still detected at a 1: Serological and RT-PCR results showed that the prepared monoclonal antibodies could be used to detect sweet potato samples in the field.