金黄色葡萄球菌5型荚膜多糖偶联抗原的制备及免疫原性分析

Preparation and immunogenicity analysis of Staphylococcus aureus type 5 capsular polysaccharide conjugated to BSA antigen

为建立金黄色葡萄球菌5型荚膜多糖(CP5)纯化与偶联技术方法,获得具有免疫原性的CP5偶联抗原,采用高压破碎菌体的方法释放荚膜多糖,酶处理去除核酸和蛋白质,通过离子交换层析进一步纯化,得到纯化的CP5,并对其纯度和反应原性进行检测;采用己二酸二酰肼(ADH)间桥法制备CP5-牛血清白蛋白(CP5-BSA)偶联抗原,加佐剂免疫小鼠,进行抗体水平测定。结果表明,纯化的CP5纯度为675.1g/kg,经用免疫琼脂双扩散试验检测,纯化多糖仅与同型菌株抗血清发生沉淀反应,偶联抗原CP5-BSA可以刺激小鼠产生CP5免疫应答。此研究结果为奶牛乳房炎金黄色葡萄球菌亚单位疫苗的研制提供了试验依据。

To establish methods for purification and to conjugation of Staphylococcus aureus type 5 capsular polysaccharide（CP5） and to prepare CP5-conjugated complete antigen with immunogenicity,the prototype strain ATCC-49521 of S.aureus was autoclaved to release capsular polysaccharide.The nucleonic acid and protein in the released capsular polysaccharide was removed by different enzyme digestion.Crude capsular polysaccharide was further purified by ion exchange chromatography.The purity and reactinogenicity of CP5 was assayed.Bovine serum albumin（CP5-BSA） conjugated antigen was prepared by adipic acid dihydrazide（ADH） spacing method,and adjuvant（ISA 70 VG） was added to immunize mice with the anti-CP5 antibody detected.The results showed that the purity of CP5 was 675.1 g/kg,the purified CP5 reacted only with anti-homeotype strain serum by double immunodiffusion test and the CP5-BSA conjugate could elicit anti-CP5 immune response.The experiment provided experimental evidence for development of S.aureus subunit vaccine to bovine mastitis.