猪源盖塔病毒Cap蛋白与E2蛋白多克隆抗体的制备及其免疫特性分析

Preparation and immune character of polyclonal antibodies of porcine Getah virus Cap and E2 Proteins

采用RT-PCR技术扩增猪源盖塔病毒(Getah virus,GETV)的衣壳蛋白(Cap)与糖蛋白(E2)基因,构建了Cap与E2蛋白的原核表达载体pET-Cap和pET-E2,转化至BL21(DE3)诱导大量表达。纯化并复性原核表达的目的蛋白,免疫BALB/c小鼠,制备了Cap蛋白和E2蛋白的多克隆抗体,间接ELISA效价达1∶105以上。Western blot和免疫荧光(IFA)结果显示,Cap蛋白和E2蛋白多抗与GETV均有良好的反应性。本研究制备的Cap蛋白与E2蛋白多克隆抗体,为建立ELISA等检测方法以及GETV致病机制的研究提供了参考。

To further understand the protein function of the Getah virus(GETV),the genes of the GETV capsid protein and glycoprotein(E2)were amplified by RT-PCR and were cloned into a pET-32 a(+)vector.Then,both recombinant plasmids were transformed into BL21(DE3)to induce the gene expression.After purification and renaturation of the target proteins expressed in prokaryotic cells,BALB/c mice were immunized three times to prepare specific polyclonal antibodies whose titer was more than 1:105 detected by indirect ELISA.Western blot and IFA results showed that the polyclonal antibodies had high reactivity with GETV,indicating that the prepared polyclonal antibodies were able to recognize the GETV-Cap protein and GETV-E2 protein.The Cap protein and E2 protein polyclonal antibodies were helpful for establishment of a rapid detection method based on immunology,and for verifying virus isolation;which laid a foundation for epidemiological investigation of the Geta virus and research on the pathogenesis of GETV.