摘要
目的:以小白鼠为试验模型,检测金黄色葡萄球菌α-溶血素基因工程蛋白单位疫苗的抗体效价水平和免疫保护力,初步评价此亚单位疫苗的免疫效力。方法:Bradfrod法对目的蛋白进行蛋白含量测定,Geoffroy法测定目的蛋白半数溶血效价和活性,建立ELISA方法测定抗体效价,并且通过攻毒得出疫苗免疫保护力。结果:纯化的目的蛋白在53kDa处显示单一条带,蛋白含量为0.1278mg/ml;溶血比活为8012.5HU/mg;所有试验小鼠在首免一周后采集的血清中都检测到了特异性抗体,而抗体效价开始时呈逐渐上升趋势,达最高值后随时间延长逐渐降低。结论:纯化的目的蛋白达到电泳级纯度,且依然保持良好的黏附活性,蛋白疫苗作用机体产生的抗体水平符合抗体消长规律。
Objective：α -Hemolysin of Staphylococcus aureus, which was expressed in E. coli BL21 （DE3） with recombinant pET32a ＋ -α-HL plasmid, was purified with gel filtration chromatography（ GFC ）, and then the engineered subunit vaccine was developed. The immunity effectiveness of this vaccine was evaluated on mouse models. Method：The purified fusion protein was analyzed in SDS-PAGE, and subjected to the evaluation of its median hemolytic dose potency （HD50） was finally analyzed with rabbit erythrocyte. Protein concentration was determined by the method of Bradford. Antibody titers were evaluated on ELISA, and then challenged to gain the immunity protect index. Results ：There is an expected protein band with molecular mass of 53kDa in SDS-PAGE, and the concentration is 0. 1278mg/ml. The hemolysis activity is 8012.5 HU/mg. There are specific antibodies acquired in blood-serum from mice after vaccined and the antibody titers rising until it has arrive the max, then following down. Conclusion ： The purified fusion protein has good fineness and hemolysis activity, the antibody titers initiated by the protein vaccine go with regulation and the immunoprotection is satisfied.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2009年第11期17-22,共6页
China Biotechnology
基金
山东省农科院高技术自主创新基金(2006YCX027
2007YCX017-03)
山东省农科院重大成果培育基金(2006YCG012)资助项目
