奶牛α_s-酪蛋白多克隆抗体的制备、纯化及鉴定

Preparation,Purification and Identification of Bovine α_s-Casein Polyclonal Antibody

本试验旨在制备高纯度和特异性的奶牛αs-酪蛋白多克隆抗体,为鉴定奶牛乳腺合成的αs-酪蛋白提供试验材料。选用4只健康新西兰大白兔,每2周免疫1次奶牛αs-酪蛋白,待血清达到抗体效价后,对兔进行颈动脉放血并分离血清,利用饱和硫酸铵法和蛋白A树脂纯化抗体,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和免疫印迹(Western-blot)法分别用于鉴定纯化后抗体的纯度和特异性。结果表明,经过饱和硫酸铵法和蛋白A树脂2步纯化后得到了纯度较高的抗体,同时可以特异性结合奶牛αs-酪蛋白。本试验制得的抗体可以用于鉴定奶牛乳腺合成的αs-酪蛋白,为进一步研究奶牛αs-酪蛋白合成的调控提供了有效的科研材料。

This experiment was conducted to produce polyclonal antibody against bovine αs-casein with high purity and specificity to provide experimental materials for identification of αs-casein synthesized by mammary gland of dairy cows.Before bloodletting from jugular arteries and preparing serum,four healthy New Zealand white rabbits were immunized with bovine αs-casein every two weeks until antibody titer was desirable.Methods of saturated ammonium sulfate and protein A resin were used to purify the antibody,and purity and specificity of the antibody were identified by methods of SDS-PAGE and Western-blot,respectively.The results showed that antibody with high purity was obtained by the two steps of purification,and it was also specific for binding to bovine αs-casein.The antibody produced from this experiment can be used to identify αs-casein synthesized by bovine mammary gland,and is a kind of effective material in research on regulation of bovine αs-casein synthesis.