基于ELISA的神经胶质酸性蛋白自身抗体的血清含量测定及其在脑胶质母细胞瘤诊断中的应用

 目的 发展基于ELISA的血清神经胶质酸性蛋白(glial fibnillary acidic protein,GFAP)自身抗体含量测定方法,用于脑胶质瘤患者的诊断。方法 克隆GFAP基因至pGEX 6p 1载体,使用大肠埃希菌中纯化得到的GFAP重组蛋白包被96孔板,经封闭后依次孵育稀释的血清样本和偶联HRP的羊抗人二抗,最后加入TMB底物显色,硫酸终止显色反应,测定各孔吸光度。结果 pGEX 6p 1 GFAP质粒在大肠埃希菌Rosetta菌株中表达产生了大量GFAP重组蛋白,所得蛋白经洗脱纯化后用作ELISA反应底物。ELISA实验中,脑胶质瘤患者血清D值明显高于正常人,差异有显著统计学意义。结论 使用GFAP作为抗原的ELISA实验可作为应用GFAP自身抗体进行脑胶质瘤患者血清诊断的一种候选方法。

ELISA based serum glial fibnillary acidic protein autoantibody detection and its application in glioblastoma diagnosis

Objective To develop a autoantibody detection method for serum glial fibnillary acidic protein(GFAP) based on ELISA used for glioma diagnosis.Methods We cloned GFAP into pGEX-6p-1 vector,purified its recombinant protein from transformed Rosetta E.coli strain,and then applied it for antigen coating step in ELISA.After being blocked with 5% BSA,and sequentially incubated with diluted sera sample and HRP conjugated goat anti-human secondary antibody,TMB substrate was added to visualize ELISA result.The colorat...