T7 RNA聚合酶催化的荧光扩增技术检测人CEA

Detection of human carcinoma embryonic antigen using fluorescent amplification catalyzed by T7 polymerase technique

目的:建立检测人癌胚抗原(CEA)的T7 RNA聚合酶催化的荧光扩增技术(fluorescent amplification catalyzed by T7 polymerase technique,FACTT).方法:以亲和素作为连接分子,连接生物素化的检测抗体和生物素化的DNA,加入T7RNA聚合酶进行转录扩增反应,对生成的RNA产物进行荧光检测,并同时进行夹心酶联接免疫吸附剂测定(ELISA)方法检测人CEA.结果:成功的建立了检测人CEA的T7 RNA聚合酶催化的荧光扩增技术,其检测CEA的灵敏度达2×10^(-3)mg/L,比夹心ELISA方法灵敏度(0.25 mg/L)高125倍.结论:T7 RNA聚合酶催化的荧光扩增技术较夹心ELISA方法具有更高的敏感性,有可能作为一种新的检测方法用于临床的早期诊断.

AIM： To establish a new method for the detection of human carcinoma-embryonic antigen （CEA） using fluorescent amplification catalyzed by T7 polymerase technique （FACTT）, METHODS： Avidin was used to bridge biotinylated detection antibody and biotinylated DNA, T7 RNA polymerase was added to perform RNA amplification, RNA products were quantified by adding the RNA intercalating dye RiboGreen, CEA was determined by the enzyme-linked immunosorbent assay （ELISA）, RESULTS： The detection limit of FACTT for human CEA was 2 × 10^-3 mg/L, which was 125 times more sensitive than ELISA （0,25 mg/L）, which was performed in parallel, CONCLUSION： The FACTT has a higher sensitivity than sandwiched ELISA for the detection of human CEA, and may be a powerful and very sensitive tool for early stage clinical diagnosis.