猪伪狂犬病病毒gB蛋白单克隆抗体的制备及鉴定

Preparation and identification of a monoclonal antibody against pseudorabies virus g B protein

为了制备猪伪狂犬病病毒(PRV)gB蛋白单克隆抗体,本研究利用原核表达系统表达gB蛋白的主要抗原区(540~734 aa),获得重组蛋白免疫BALB/c小鼠。利用杂交瘤技术,经过细胞筛选和亚克隆,获得1株能稳定分泌抗gB蛋白的单克隆抗体,将其命名为1C10。经间接免疫荧光试验(IFA)和免疫印迹试验(Western-blot)验证,1C10具有良好的特异性,能识别PRV感染细胞时表达的gB蛋白。亚类鉴定结果表明,1C10重链为IgG2a型,轻链为kappa型。ELISA结果表明,纯化后的腹水效价高于1∶128000。综上,本研究成功表达了PRV gB蛋白,并制备了抗PRV gB蛋白单克隆抗体,为进一步探究PRV gB蛋白的结构和功能以及后期建立PRV诊断方法及致病机制的研究奠定了基础。

In order to prepare monoclonal antibodies(MAbs)against the gB protein of porcine pseudorabies virus(PRV),the main antigen region(540—734aa)of gB protein was expressed by a prokaryotic expression system,and the recombinant protein was used to immunize BALB/c mice.Using hybridoma technology,a MAb against the gB protein was obtained through the cell screening and subcloning,and named as1C10.Indirect immunofluorescence assay(IFA)and Western-blot assay verified that 1C10 has a good specificity and could recognize the gB protein expressed in cells infected with PRV.The identification of isotyping revealed that the 1C10 was of IgG2a type for heavy chain and kappa type for light chain.The ELISA data showed that the purified ascites was higher than 1∶128000.In summary,PRV gB protein was successfully expressed and a MAb against PRV gB protein were prepared in this study,laying the foundation for further study on the structure and function of PRV gB protein,as well as the establishment of diagnostic methods and pathogenic mechanisms in the later stage.