一种蛋白质B细胞表位展示系统的构建及验证

目的:建立一种能够有效展示蛋白质B细胞表位的载体系统,以用于表位特异性抗体的制备和表位疫苗的设计。方法:选用抗体的可变区作为蛋白质B细胞表位展示的骨架蛋白,B细胞表位的展示部位为CDR3区。全基因合成骨架蛋白基因,通过重叠PCR将B细胞表位编码序列插入到骨架蛋白基因中,原核表达目的蛋白,利用Sephacryl S-100层析柱进行蛋白纯化,用纯化的蛋白按常规方法免疫小鼠,采用ELISA法检测免疫血清的滴度及特异性,通过Western blot和间接免疫荧光技术进一步验证免疫血清对目的蛋白的识别。结果:成功构建了3种表位展示蛋白,均表现出了很好的抗原性,表位展示蛋白免疫血清具有较好的特异性,能够特异识别所展示的B细胞表位。结论:抗体可变区作为骨架蛋白能够很好地展示B细胞表位,免疫小鼠后获得的免疫血清表现出了较好的特异性。

Construction and Verification of a Novel System for B-cell Epitope Display

B cell epitopes are the sites of the molecules that are recognized by antibodies or B cell receptors.Prediction and verification of B cell epitopes of antigen is useful for specific antibody generation and vaccine design.The immunoglobulin heavy chain variable region was used as the scaffold protein for B cell epitope display,and its CDR3 region was the display site.Three B cell epitopes were displayed in the protein scaffold,including(His)6 tag and two epitopes derived from VEGF.The three recombinant proteins were used to immunize mice,and the immune serum was isolated and analyzed by ELISA,Western blot and immunofluorescence.The results demonstrated that the immune serum exhibited good specificity for the corresponding epitopes and the source protein.This means that the immunoglobulin heavy chain variable region is a good scaffold protein for B cell epitope display.