原核表达牦牛朊蛋白的纯化及其活性测定  

Purification and activity determination of recombinant prion protein of yak expressed in prokaryotic cell

作　　者：伍志伟[1] 吴润[1] 刘湘涛[2] 陈豪泰[2] 

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070 [2]中国农业科学院兰州兽医研究所农业部畜禽病毒学重点开放实验室,甘肃兰州730046

出　　处：《甘肃农业大学学报》2005年第2期133-137,共5页Journal of Gansu Agricultural University

基　　金：农业部畜禽病毒学重点开放实验室基金(2002-01);兰州兽医研究所所长基金资助(2004-2005)

摘　　要：以原核表达的GST-BoPrP(23～242)融合蛋白为抗原,免疫BALB/C小鼠,制备抗牦牛朊蛋白的特异性抗血清。经Western blotting和间接ELISA鉴定,该抗血清可与牦牛重组成熟PrP(23～242)和牛脑组织提取物发生反应,蛋白酶K消化各抗原可消除免疫反应,但不与GST蛋白和E.coli BL21(DE3)的菌体蛋白发生反应,表明该抗血清为抗牦牛重组成熟PrP(23～242)的抗血清,其效价高达1∶12800,并能识别黄牛脑组织中的天然朊蛋白。原核表达的GST-BoPrP(23～242)融合蛋白能有效地刺激免疫动物产生PrP特异性抗体,所制备的抗血清可适用于天然朊蛋白的检测。Using prokaryotic expressed GST-BoPrP(23～242) fusion protein as antigen, BALB/C mice were immunized to prepare the specific antiserum against the yak prion protein. By Western blotting and indirect ELISA detecting, the antiserum could react with the yak recombinant mature PrP(23～242) and the extraction of bovine brain tissue, respectively. After digestion of the antigens by proteinase K, the immune reaction disappeared. However the antiserum could not be reacted with the GST protein and the full lysate of E. coli BL21(DE3). The results indicated that the antiserum was the one against the yak recombinant mature PrP(23～242) with its titer higher than 1:12800, and it could detect the native prion proteins from the brain tissue homogenate of cattle. The GST-PrP(23～242) fusion protein could efficiently stimulate the immunized animals to produce the PrP-specified antibody, and the antiserum could be used for detecting the native prion proteins.

关 键 词：牦牛 重组成熟朊蛋白 纯化 活性测定 抗血清 原核表达 疯牛病 

分 类 号：S858.23[农业科学—临床兽医学] S859.797[农业科学—兽医学]