摘要
目的 制备 14 3 3特异性抗体并试用于朊病毒病患者脑脊液标本检测。方法 以原核细胞表达的谷光甘肽 S 转移酶 (GST) 14 3 3蛋白为抗原 ,免疫家兔。用酶联免疫吸附测定法(ELISA)及Westernblot方法检测制备的抗血清效价和特异性。结果 ELISA检测显示所制备的抗体与纯化的GST 14 3 3蛋白反应效价为 1∶40 96 0 0 0 ;Westernblot结果证实所制备的抗体可有效识别原核细胞表达的及脑组织提取物中存在的 14 3 3蛋白。对 2例具有较为典型脑电图改变、临床可疑诊断为克罗伊茨费尔特 雅各布病 (Creutsfeldt Jacobdisease,CJD)病人的脑脊液检测发现其 14 3 3蛋白为阳性 ;另外 4例伴有痴呆、昏迷症状病人脑脊液 14 3 3蛋白为阴性 ,其中 3例进行的脑组织活检显示致病性朊蛋白 (PrPSc)阴性 ,无典型CJD病理改变 ;其余 2 4例无痴呆症状的脑炎及脑病病人脑脊液 14 3 3蛋白均为阴性。以上结果与用标准 14 3 3抗体所获结果完全吻合。结论 利用原核细胞表达的GST 14 3 3蛋白可有效地诱导免疫动物产生特异性抗体 ,所制备的抗体可用于CJD病人脑脊液 14 3 3蛋白的检测。
Objective Preparation of specific antibody of neuron protein 14 3 3 and application for the diagnosis of prion diseases. Methods The rabbits were immunized with prokaryotic expressed GST 14 3 3 fusion protein and the collected antiserum was evaluated by ELISA and Western blot assays. Results ELISA revealed that the reactive titer of the prepared antibody against purified 14 3 3 protein was up to 1:4 096 000; Western blot assays confirmed that both prokaryotic expressed,and the natively presenting 14 3 3 proteins in the brain extracts were recognized by the prepared antibody. Two cases of clinically diagnosed Creutzfeldt Jakob disease (CJD)with typical EEG changes showed the positive 14 3 3 protein in cerebral spinal fluid (CSF). Four patients with dementia and narcoma but without typical EEG change were 14 3 3 negative in CSF,three of them being even further tested by brain biopsies without typical histological change of CJD and presentation of protease resistant PrP Sc protein. 35 samples CFS collected from patients without dementia were 14 3 3 negative in CSF. All the results were the same as that of standard 14 3 3 polyclonal antibody.Conclusion Using purified prokaryotic expressed GST 14 3 3 fusion protein as antigen,the specific antibody was elicited in the immunized animals. The prepared antibody can be used in identification of 14 3 3 protein in CSF for the diagnosis of CJD.
出处
《中华神经科杂志》
CAS
CSCD
2000年第5期302-304,共3页
Chinese Journal of Neurology
基金
国家 8 6 3高科技发展计划项目! (86 3 10 2 11 0 3 0 5 )
