Eu标记RT-PCR检测丙型肝炎病毒RNA

目的 采用一种新的铕螯合物BHHCT，制备高灵敏度荧光标记物，建立检测丙型肝炎病毒（HCV）RNA的方法。方法 采用柱层析纯化的方法抽提血清中HCV RNA。以5′端带有生物素的引物进行RT－PCR Tth酶一步法扩增，通过固定于微孔板上的捕获探针与带有生物素的PCR扩增产物杂交后，利用标记有铕的链霉亲合素与生物素的特异性结合，将铕连接到待测核酸上，在特定波长的激光光激发下发出荧光，进行检测。结果 铕标记检测法与酶标法的变异系数、敏感性、特异性分别为10．43％、100％，100％与16．25％，94％，100％。结论 Eu标记RT－PCR Tth酶一步法检测HCV RNA，具有敏感、特异、快速、重复性好、不受环境因素影响和无溴乙锭污染等优点，可望取代酶标法及电泳法。

Detection of hepatitis C virus by using a new europium fluorescent chelate as a label

Objective To establish a method which utilize the new europium fluorescent chelate BHHCT as label for highl sensitive detection of hepatitis C virus.Methods HCV RNA extracted by chromatography was one-round amplified by Tth enzyme PT-PCR in which,the primers were pre-labeled with biotin.The amplified products were hybridized with capture probe pre-fixed on the microplate.By conjugation of biotin with europium-labeDNAled streptavidin,Europium was linked to the target DNA,and fluorescent of europium was measured.Results The CV values,sensitivity and specifity of this method were 10.43%,100%,100% respectively,compared with enzyme-labeled assay 16.25%,94%,100%,Conclusion Europium labelled RT-PCR assay is a sensitive,specific,fast and precise method for detection of hepatitis C virus.