摘 要IgM 是鱼体特异性免疫系统重要组成部分，在鱼体应答外源刺激、病原感染中发挥重要作用。本文通过对养殖大黄鱼尾静脉采血，离心获得血清后，用饱和硫酸铵盐析法粗提鱼体血清 IgM，再利用 Protein A 特异性结合免疫球蛋白分子恒定区特性，进一步采用 Protein A 亲核层析柱对粗提液进行了纯化。纯化后的蛋白经冷冻干燥、浓缩、重悬后，采用 Bradford 试剂盒法对其进行了浓度测定，实验测得的纯化蛋白浓度约为 0.5mg/mL。对纯化后的蛋白进行聚丙烯酰胺凝胶电泳（SDS-PAGE）分析，结果显示，纯化蛋白不含其他条带，表明其纯度高，不含杂质纯化蛋白由两条分子量大小分别为约 70KDa 和 30KDa 的条带组成，这与鱼类IgM 的重轻链分子大小吻合，表明实验纯化得到的即为大黄鱼 IgM。综上，采用饱和硫酸铵粗提结合 Protein A 亲和层析纯化，能获得较好的大黄鱼血清 IgM 纯化效果。通过对纯化的大黄鱼血清 IgM 进行特性分析，将有助于加深对大黄鱼免疫系统的理解。关键词：大黄鱼；盐析；亲和层析；IgMAbstractIgM is an important component of fish-specific immune system, which plays an important role in fish response to exogenous stimuli and pathogen infection. This passage through collecting blood from tail vein of large yellow croaker culture, after centrifugation to obtain serum, use salting out saturated ammonium sulfate to extracted the serum IgM, and then using Protein A specific binding immunoglobulin molecular homeostatic region, the crude extract was further purified by Protein A nucleophilic chromatography column. The purified protein after freeze-dring concentration and suspension, the concentration of the purified protein used the Bradford kit to determined, the result of concentration is about 0.5mg/ml. The purity and molecular weight of IgM was checked by SDS-PAGE, the obtained IgM showed obviously two main bands with the molecular weight of 70 kDa and 30 kDa, this is consistent with the size of the heavy and light chains of fish IgM, indicating that the IgM obt...
