利用霍乱毒素 B亚单位展示猪 O型口蹄疫病毒 VP1蛋白主要抗原表位

摘要

The object of this study is to find out whether cholera toxin subunit B ( CTB) is suitable for displaying GH loop epitope of type O FMDV ( foot⁃and⁃mouth disease virus) . The chimeric gene CTB⁃GHloop was expressed in Escherichia coli. SDS⁃PAGE was used to detect the expression and the solubility of the recombinant protein. Ganglioside M1 coated plate was involved as antigen to identify antibody by ELISA. The immunogenicity of chimeric CTB⁃GHloop protein was evaluated on pigs by detection of FMDV specific antibody and lymphocyte proliferation activity against GHloop epitope. Recombinant protein was identified in the form of pentamers by Wester⁃blot. Antibody test results revealed that piglet immunized with 200μg recombinant CTB⁃GHloop protein developed not only higher titer of antibodies in a short period of time, but also more vigorous T cell immune response compared with the commercial vaccines. It is concluded that CTB protein could be used as a carrier to display other antigenic epitope with no influence on the formation of pentamers, and the recombinant protein CTB⁃GHloop may be a promising candidate for FMDV subunit vaccine research.%为探讨以霍乱毒素B亚单位为载体制备口蹄疫亚单位疫苗的可行性，利用E． coli表达CTB⁃GHloop嵌合基因，用 SDS⁃PAGE分析目的基因的表达及表达产物的可溶性，利用神经节苷脂（ GM1）为抗原鉴定重组蛋白五聚体的形成。将目的蛋白浓度调整为200μg／ml，以白油佐剂乳化制备疫苗，免疫健康仔猪，免疫后利用ELISA测定特异性抗体水平，评价免疫后体液免疫反应，利用淋巴细胞增殖实验评价细胞免疫水平。结果表明嵌合基因在E． coli中获得高效表达，重组蛋白可溶，并能够形成五聚体；Western⁃blot结果显示重组蛋白能够与口蹄疫病毒（ FMDV）阳性血清发生反应；以每头仔猪200μg免疫，试验猪产生较高的抗体水平与细胞免疫反应。