生物素化高分子涂层的制备与评价

制备了一种能固载目标蛋白质, 却没有非特异性蛋白质吸附的高分子涂层. 该涂层是可生物降解的油水两亲性的三嵌段聚合物, 即生物素偶联的聚乙二醇-聚丙交酯-聚赖氨酸共聚物. 将高分子溶解于N,N-二甲基甲酰胺中, 并涂布在预先包被了聚赖氨酸的脱脂玻片基质上, 形成高分子涂层, 在其表面包被一层由明胶和聚N-乙烯基吡咯烷酮组成的封闭剂. 使用酶标免疫分析法, 对高分子涂层表面的生物活性进行评价. 依次将辣根过氧化物酶标记的链亲和素和生物素偶联的小鼠球蛋白抗原和碱性磷酸酯酶标记的马抗小鼠抗体固载在高分子涂层表面上, 通过标记酶与底物作用生色. 分析结果表明, 经过封闭以后, 生物素化的高分子涂层表面能够排斥非特异性的蛋白质; 同时特异性蛋白质之间(如生物素和链亲和素之间、抗原和抗体之间)的相互作用依然保留, 并且固定在表面的蛋白质依然保留其生物活性. 因此生物素化的聚乙二醇-聚丙交酯-聚赖氨酸三嵌段高分子可以作为生物活性材料, 用于蛋白质固载和蛋白质分离及分析.

Preparation and Evaluation of the Biotinylated Polymer Coating

A novel bioactive polymer coating without nonspecific adsorption of proteins was prepared from a biodegradable amphiphilic triblock copolymer,biotinylated poly(ethylene glycol)-b-poly(L-lactide)-b-poly(L-lysine)(PEG-PLA-PLL/biotin)by coating it on a substrate and covering it with a layer of blocking coating consisting of gelatin and poly(N-vinyl pyrrolidone).The bioactive surface was characterized by enzyme linked immunoassay,on the basis of sequential immobilizations of horseradish peroxidase labeled streptavidin,biotinylated mouse globulin,and alkaline phosphatase labeled horse anti mouse IgG(H L).The results showed that the bioactive surface was resistant to nonspecific protein adsorption but allowed specific recognition and combination between biotin and streptavidin,and meanwhile,the combined streptavidin retained its bioactivity.Therefore,subsequent protein immobilization can be realized via the reaction of the streptavidin and biotinylated proteins.