猪圆环病毒2型Cap蛋白单克隆抗体制备及线性抗原表位鉴定

Preparation of the monoclonal antibody against Cap protein of porcine circovirus type 2 and its antigenic epitope identification

为制备猪圆环病毒2型(PCV2)Cap蛋白单克隆抗体(MAb)及鉴定其抗原表位,本研究采用PCV2去核定位信号Cap蛋白(dCap),通过杂交瘤技术制备1株杂交瘤细胞株(4E2),制备小鼠腹水效价达1∶64 000,亚类鉴定为IgG1/κ链。Western blot分析显示,该株MAb与毕赤酵母表达的dCap和原核表达的Cap蛋白均可以发生特异性免疫反应。采用肽扫描法对MAb的非核定位信号区进行抗原表位鉴定,结果表明该MAb可以识别的线性表位区域为131TKATALTYDPYVNYSS146。本实验结果为进一步研究Cap蛋白的结构和PCV2临床检测方法的建立奠定基础。

To prepare monoclonal antibodies （MAb） against Cap protein of porcine circovirus Ⅱ （PCV2） and identify the antigen epitope, a MAb against PCV2 was prepared by lymphocyte hybridoma technique with the recombinant Cap protein without nuclear location signal （dCap） expressed by P.pastoria. The MAb was belonged to IgG1 subclass and κ light chain. The ELISA titer of MAb was 1：64 000 in ascites. Western blot indicated that MAb was reacted with PCV2-dCap expressed in P.pastoria and PCV2-Cap expressed in E.coli. IFA assay showed the MAb was reacted with PCV2 in PK15 cells. Furthermore, the MAb recognized dCap protein epitope of ^131TKATALTYDPYVNYSS^146 mapped by a serious of prokaryotic expressed peptides. The MAb prepared in this study is useful tool for developing immunological diagnostic techniques of PCV2 and further investigation of the Cap protein.