免疫印迹法检测牛海绵状脑病和羊瘙痒病

用大肠杆菌表达的牛朊病毒正常成熟蛋白 (BoPrPC)免疫新西兰白兔 ,获得了与朊病毒蛋白 (PrP)反应的抗体T1。根据致病型朊病毒 (PrP^SC)能抵抗蛋白酶消化的特性 ,用蛋白酶K消化脑组织提取物 ,以抗体T1进行免疫印迹反应 ,结果表明从接种羊瘙痒病朊病毒 2 6 3K的金黄地鼠脑组织提取物内检测到抗蛋白酶K消化的致病型PrPSC ,而正常金黄地鼠脑组织中没有抗蛋白酶消化的蛋白。以我国正常牛羊为材料 ,制备其脑组织提取物 ,用上述方法和抗体T1进行检测 ,结果没有发现抗蛋白酶K的任何蛋白存在 ,说明没有牛海绵状脑病和羊瘙痒病存在。用 1A8抗体也获得了同样的结果。这些结果表明可以用自制的抗血清检疫牛海绵状脑病和羊瘙痒病 ,防止其传入我国

Diagnosis of Bovine Spongiform Encephalopathy and Scrapieby Western blot

The rabbits were immuned with bovine prion protein (BoPrPC) which was expressed in E. coli and anti-PrPC antibody (T1) was obtained. According to pathological prion protein (PrPSC) was resistant to protease treatment, extracts of brain tissue were digested with proteinase K and detected by western blot with T1 antibody. The results showed that protease-resistant pathological PrPSC was existed in golden hamster brain tissue which was inoculated with scrapie strain 263 K, but no protein existed in normal golden hamster brain homogenates which was detected with T1 antibody. Several bovines and sheep from Beijing were used for diagnosis of Bovine spongiform encephalopathy (BSE) and scrapie, their brain tissue were freshly collected and homogenated. The homogenates were separated on SDS-PAGE and detected by western blot with T1 antibody. The results indicated no protease-resistant protein(PrPSC) existed, this suggested they were not infected by BSE and scrapie. The same results were obtained with 1A8 antibody from England. These results indicated we could detect BSE and scrapie with T1 antibody.