桃拉综合征病毒VP1高变区蛋白抗体制备及初步特性分析

Polyclonal antibody specific for hypervariable region of VP1 capsid protein of Taura syndrome virus( TSV) prepartion and characteristic analysis

目的:制备高效价的桃拉综合征病毒主要结构蛋白VP1高变区蛋白抗体,分析其免疫特性,为研究免疫诊断试剂做参考。方法:将p ET-VP1重组载体转化入大肠杆菌BL21,在IPTG诱导下进行表达。重组VP1蛋白纯化后免疫新西兰家兔,用琼脂扩散试验及ELISA检测抗体效价,用与VP1纯化蛋白特异性结合的单克隆噬菌体做竞争抑制试验。结果:p ETVP1重组蛋白在大肠杆菌BL21中呈可溶性高效表达。抗血清经琼扩试验鉴定效价达1∶26,ELISA效价达1∶217。与VP1蛋白特异结合的单克隆噬菌体能阻断部分抗血清与VP1蛋白的结合活性,但不影响最终检测阳性的判断。结论:制备的VP1高变区蛋白抗体效价高,VP1蛋白少数区域的位点变异并不影响多克隆抗体的结合活性,可用作免疫检测诊断试剂。

Objective： Preparation and immune characteristic analysis of polyclonal antibody against hypervariable region protein of Taura syndrome virus major capsid protein VP1 as a reference for studies on immunological diagnosis reagent. Methods ： The recombinant vector pET-VP1 was transformed into E. coli BL21 for protein expression. Immunizing a New Zealand rabbit with purified VPI protein, the titer of anti-VP1 serum was determined by Agar diffusion test and ELISA. Monoclonal phage specific binding to the purified VP1 protein was used for competitive inhibition test. Results： The VP1 protein was soluble and high expression in E. coli BL2I. The biological activity titer of anti-VP1 serum reached 1：26, 1：217 determined by Agar diffusion test and ELISA respectively. A litter binding activity of antiserum and VPI protein could be blocked by monoclonal phage, but would not affect the final positive result. Conclusion： High titer antibody Preparation of the VP1 hypervariable region protein. The binding activity of the polyclonal antibody with VP1 protein was not affected by the mutations of VP1 protein in minority areas, so the antiserum could be used as immu- nological detection diagnosis agent.