鸡血清载脂蛋白AⅠ的分离纯化及其抗血清的制备

Separation and purification of apolipoprotein AⅠ from chicken serum and preparation of its specific antiserum

用磷钨酸沉淀制备鸡血清HDL,乙醇丙酮混合液(体积比为1∶1)脱脂后采用DEAESepharoseCL6B离子交换柱层析法分离鸡血清apoAⅠ,分离获得的apoAⅠ在尿素SDSPAGE电泳中呈现单一条带,其分子质量约为27788u。将提取的apoAⅠ对新西兰雄兔进行多次免疫,制备得兔抗鸡apoAⅠ抗血清,其效价为1∶16。在免疫电泳和双向免疫扩散试验中抗血清均只呈现1条清晰沉淀带。试验结果表明,用磷钨酸沉淀法制备兔抗鸡apoAⅠ抗血清,过程简便,时间短且分离效果好。

The HDL of laying hen was isolated and purified from pooled serum by precipitating with phosphotungstic acid. The apolipoprotein AⅠ was obtained from HDL by delipitating with acetone-ethanol(Ψ=1)and purified by ion exchange chromatography with DEAE sepharose CL-6B. The identification by 12.5% urea-SDS-PAGE showed that the purified apolipoprotein AⅠ have only one band and its mole- cular weight is 27788u. The specific antiserum was prepared by immunizing New Zealand male rabbit with the obtained protein for several times. The titer of the prepared specific antiserum was 1∶16, and the specific antiserum showed a single sedimentary band on immune electrophoresis and double immunodifusion. The results showed that the preparation of the rabbit-anti chicken antiserum by the phosphotungstic acid precipitation method was simple, saved times and could achieve good results.