【摘要】 采用饱和硫酸氨分步沉淀和Sephadex G-200凝胶层析的方法,首次分别纯化制备了健康非免疫状态下中华鲟(Acipenser sinensis)、史氏鲟(Acipenser schrenckii)和达氏鳇(Huso dauricus)的血清免疫球蛋白(Ig),并采用聚丙烯酰胺凝胶电泳(PAGE)、SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)、蛋白免疫印迹(Western blotting)及免疫琼扩实验等方法对其Ig及Ig亚单位的分子量和部分特性进行了分析。PAGE及SDS-PAGE的结果显示:史氏鲟,中华鲟和达氏鳇IgM的相对分子量分别为867kD,896kD和924kD;3种鲟鱼Ig的重链分子量均为88kD,都具有29kD的轻链,其中达氏鳇还另有一分子量约为26kD的轻链蛋白。分子量的测定及计算结果显示鲟鱼的Ig为四聚体。Western blotting的检测结果表明,3种鲟鱼Ig的重链与其Ig具有同样的抗原性,在硝酸纤维素膜上可被兔抗鲟Ig多克隆抗体所识别,而轻链的Western blotting检测结果则呈阴性。免疫沉淀反应的结果显示,3种鲟鱼的血清及其Ig与相互之间的兔抗Ig血清有免疫沉淀反应,但与兔抗鲤Ig血清无免疫沉淀反应,这表明3种鲟科鱼类的Ig在结构和序列上是较为相似的,而与鲤鱼等高等硬骨鱼类的Ig存在较大的差别。更多还原

【Abstract】 Serum immunoglobulin(Ig) of Amur sturgeon Acipenser schrenckii,Chinese sturgeon Acipenser sinesis and Huso sturgeon Huso dauricus which were healthy and unimmuned,was first time respectively purified by using saturation ammonium sulfate precipitation followed by Sephadex G-200 gel column chromatography.Molecular weight and partial characterization of Ig and heavy chain and light chain of Ig were measured by polyacrylamide gel electrophoresis(PAGE),SDS polyacrylamide gel electrophoresis(SDS-PAGE),Western blotting and Ouchterlony method.The results show that Ig molecular weight of Amur sturgeon,Chinese sturgeon and Huso sturgeon is 867 kD,896 kD and 924 kD respectively.Heavy chain molecular weight of Ig in the three sturgeons’ serum is all 88 kD and light chain of Amur sturgeon and Chinese sturgeon is 29 kD,but there is an extra light chain of 26 kD in Huso sturgeon serum.The results of measurements and calculation show that Ig of sturgeon is tetramer.In Western blotting,Ig and Ig heavy chain of the sturgeons have the same antigenicity and can be detected by antibody of rabbit anti the sturgeon Ig respectively in colloxylin membrane while light chain cannot be detected.Serum and Ig in serum of the three sturgeons can react with serum of rabbit anti each of the three sturgeons’ Ig,but cannot react with serum of rabbit anti Ig of carp Cyprynus carpio.The Ouchterlony results show that Ig of sturgeons are immune homologous in structure and sequence,but have little immune homologous characters compared with Ig of high-classification Osteichthyes such as carp.更多还原