基于反向荧光增强和抗体定向固定技术的AMI两种标志物POCT检测研究

由于目前POCT荧光免疫层析检测技术存在信号强度低，背景噪音高及抗体免疫活性低的缺陷，难以实现对急性心肌梗死（AMI）痕量血清标志物肌钙蛋白I（cTnI）和肌酸激酶同工酶（CK-MB）的同步检测，本项目拟将反向荧光增强技术与抗体定向固定技术有机结合，研发一种快速、灵敏的POCT免疫层析试纸条，同步检测cTnI和CK-MB。通过增强纳米金大作用范围内的荧光淬灭性能，放大检测信号并降低背景噪音；通过真菌I型疏水蛋白(HGFI)定向固定抗体，高效保持抗体的免疫活性，从而显著提高检测灵敏度。并利用两种荧光染料的双标记成功实现cTnI和CK-MB同步检测。在确定关键参数的基础上，进行临床血清样本的检测，与临床电化学发光免疫检测方法进行比较，验证该方法的有效性。本课题的开展将为实现急性心肌梗死的快速准确诊断提供一种新方法。

At present, it’s difficult for the POCT fluorescence immunochromatographic assay(FICA) to realize the rapid and high sensitive detection of the acute myocardial infarction biomarkers (cTnI and CK-MB) in the serum because of the low signal strength, high background noise and the instable antibody immunocompetence. In this subject, we will combine the "reverse" fluorescence enhancement technology and antibodies oriented immobilization technology to overcome these shortcomings, as follows: the fluorescence quenching property of gold nanoparticles in a wide spacing range is enhanced so as to realize the signal amplification and background noise elimination; the HGFI is applied to orientedly immobilize the antibody in order to keep the high immunocompetence; the cTnI and CK-MB molecules are simultaneously detected by the two kinds fluorescent dye labelling. After the key parameters are determined, the FICA is further applied in the clinical serum samples detection and the results are compared with those of the electrochemiluminescence immunoassay. The project will provide a new method for rapid and accurate diagnosis of acute myocardial infarction.