血小板膜抗原的提取及其免疫原性研究

目的:建立血小板外膜蛋白复合物(outer membrane protein complex,OMPC)的提取方法,为研制具有临床诊断价值的多克隆抗体提供依据.方法:分别以DTT裂解法、SDS破碎法、NP-40裂解法和TritonX-100裂解法提取血小板膜抗原,利用SDS-PAGE电泳、动物实验、蛋白免疫印迹分析和ELISA评价裂解效果和免疫原性.结果:在4种方法中,TritonX-100裂解法效果最好,其主要含有分子量从50～160 kD的几乎所有血小板膜糖蛋白和少量小分子的蛋白.结论:TritonX-100裂解血小板获得的OMPC溶解性好、免疫原性强,具有广泛的抗原交叉反应性,以此作为动物免疫或作为偶联多糖抗原的载体具有很好的应用前景.

Extraction and Immunogenicity of Platelet Membrane Antigen

Objective:To establish an extraction technique for the outer membrane protein complex(OMPC) from the platelets and to lay a basis for producing polyclonal antibodies against OMPC for medical diagnosis.Methods:Four methods for extracting OMPC from the platelets were compared in this study,that is,DTT schizolysis,SDA quassation,NP-40 schizolysis and TritonX-100 schizolysis.Then,SDS-PAGE electrophoresis,rabbits test,western blot and ELISA were used to evaluate extraction effect and immunogenicity.Results:Of the four methods for extracting OMPC,TritonX-100 produced the best outcome.The OMPC product contained almost all platelet membrane glycoprotein components with molecular weight ranging from 50 to 160 kD and some small molecule proteins.Conclusions:The TritonX-100 extracted OMPC of platelets possessed good solubility,high immunogenicity and broadly cross reactivity.This seemed to be highly potential when it was used to immunize animals or regarded as a carrier of polysaccharide antigens.