检测肌糖蛋白C夹心ELISA方法的建立及初步应用

Establishment and primary application of sandwich ELISA method to detect tenascin-C

目的:以已经制备的抗肌糖蛋白C(TN-C)的单克隆抗体(mAb)为基础,建立能定量检测肌糖蛋白C浓度的夹心ELISA方法,并初步于临床血清标本检测。方法:将3株mAb(克隆号分别为:1A8、3H7和4D6)制备腹水后纯化,分别与辣根过氧化物酶交联后,两两配对,以重组肌糖蛋白C蛋白为检测抗原,分析抗体之间最佳组合;利用建立的夹心ELISA方法检测收集的临床骨肉瘤患者和正常人血清标本。结果:包被1A8 mAb,HRP-4D6配对敏感性最高;骨肉瘤患者血清中肌糖蛋白C浓度明显高于正常人。结论:成功建立检测肌糖蛋白C的夹心ELISA方法,并利用其检测到肌糖蛋白C在骨肉瘤患者中的浓度异于正常人。

AIM： To establish a sandwich method to detect tenascin-c on the basis of preparation of monoclonal antibodies（mAbs） against tenascin-C（TN-C）.METHODS： The ascites of three stains of mAbs（No.1A8,3H7 and 4D6） were prepared and purified.The mAbs were conjugated with HRP and paired,respectively.The recombinant TN-C was taken as standard to analyze the optimal combination between mAbs.The sera TN-C concentrations of patients with osteosarcoma and the normal persons were evaluated with the sandwich ELISA method.RESULTS： Among these mAbs,the most sensitivity was obtained when combined the coated 1A8 with HRP-4D6.The sera TN-C concentrations of patients with osteosarcoma were significantly higher than the normal controls.CONCLUSION： The sandwich ELISA method to detect TN-C was established successfully.The sera TN-C concentrations of patients with osteosarcoma and the normal persons were found distinct with the sandwich method.