鲫鱼血清和皮肤粘液IgM的分离纯化及部分性质的鉴定

采用盐析法结合葡聚糖凝胶柱 ,分离纯化鲫鱼血清IgM ;然后制备兔抗鲫鱼血清IgM多克隆抗体 ,将其偶联到Sepharose 4B上制成亲和柱 ,用于分离纯化皮肤粘液IgM。结果表明 :33%～ 4 5 %硫酸铵溶液沉淀处理可以去除鲫鱼血清中除IgM外的很多杂蛋白 ,再经葡聚糖凝胶柱纯化 ,IgM纯度可达 80 %以上 ,其重链和轻链的分子量分别为 79和 2 5kDa ;以兔抗鲫鱼血清IgM多克隆抗体亲和柱分离皮肤粘液IgM ,分离效果良好 ,IgM重链的分子量为 88kDa ;Westernblot显示兔抗鲫鱼血清IgM多克隆抗体识别的是血清和皮肤粘液IgM的重链部分。用ELISA测定鲫鱼血清中IgM含量在一年中的变化 ,结果表明IgM在春夏季的含量高于秋冬季

Purification and Partial Characterization of Immunoglobulin M from Carassius auratus Serum and Skin Mucus

Serum IgM in Carassius auratus was purified by precipitation of ammonium sulfate saturated solution followed by Sephadex G 200 chromatography.Then we made rabbit polyclonal antibody against serum IgM and linked it to Sepharose 4B to made affinity column,with which IgM was separated from skin mucus.The results showed that many other proteins were removed from serum by precipitation of ammonium sulfate,IgM could reach a purity of above 80% by Sephadex G 200 chromatography purified further,and the molecular weights of its heavy chain and light chain were 79?kDa and 25?kDa respectively.The rabbit polyclonal antibody affinity column had good capacity of purifying skin mucus IgM,whose heavy chain was 88?kDa.The rabbit antibody only recognizes the heavy chain of serum and skin mucus IgM by Western blot analysis.The result of ELISA indicated that IgM concentration in serum was higher in spring and summer than in autumn and winter.