液相蛋白质芯片检测血清C反应蛋白方法的建立

Liquid chip:a new method for detection of serum high-sensitivity C-reactive protein

目的建立液相蛋白质芯片检测血清C反应蛋白(CRP)的方法,探讨该方法诊断冠心病的临床应用价值。方法用聚苯乙烯荧光微珠,将抗CRP单克隆抗体包被在微珠上,将另一针对CRP不同抗原表位的单克隆抗体生物素化;包被好的微珠加入到96孔板中,将待测血清分别加入各孔,用CRP标准品建立标准曲线,各孔中加入生物素化抗CRP单抗和PE荧光素标记的链霉亲合素。在Bio-Plex液相蛋白质芯片分析仪上检测296份血清样品。同一血清标本同时用免疫比浊法测定。比较两种方法对CRP检测的敏感性和特异性。结果液相蛋白质芯片检测CRP方法对诊断冠心病的敏感性(78.3%)和特异性(87.1%)较免疫比浊法的敏感性(63.0%)和特异性(86.5%)高,χ2=28.94,P<0.001。结论建立了一种检测血清CRP的新方法,对诊断冠心病有临床应用价值。

Objective To establish a new method using liquid chip for detection of serum high-sensitivity C-reactive protein （CRP） and estimate its values in clinical application for diagnosis of coronary heart disease （CHD）. Methods A monoclcnal antibody to CRP was coupled to the fluorescent beads of carboxylated polystyren and another monoclonal antibody to different epitope of CRP was biotinylated. The antibody-coupled beads were added into each well of 96-well microtiter plate and the diluted standard or sample was in turn added to the corresponding wells. Following keeping the plate in dark Streptavidin-PE were added into each well. The plate was then proceeded on Bio-Plex protein array system to read the results. A total of 296 samples were detected by this method and immunoturbidimetry simultaneously. The sensitivity and specificity of the 2 methods were compared. Results The sensitivity of the established liquid chip method was 78.3% and the specificity was 87.1% ,while in the immunoturbidimetry were 63.0% and 86.5% respectively. Conclusion Using the liquid chip system to detect serum CRP is a credible method. The establishment of this method will offer feasibility for using liquid chip system to detect multiple protein markers in diagnosis of CHD and other diseases.