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目的:建立一种基于免疫-RCA的甲型流感方便快速的筛查方法。方法:根据甲型流感M基因的特异序列设计一条环化引物,在聚合酶和连接酶的共同作用下形成DNA环状单链,双引物进行RCA指数扩增,扩增产物通过固相于96孔板标记探针杂合,酶免疫结合显色判断结果。结果:应用本方法检测100份流感样病例样本,其结果与荧光PCR检测结果符合率达到97%,特异度为100%,灵敏性为94%。结论:利用RCA恒温扩增,产物为大量DNA重复单元的特性,将其与酶联免疫结合起来,进行甲型流感的快速筛查,方法简便、快速,具有推广意义。
Objective: To establish a convenient and rapid screening method for influenza A (RCA) -based influenza. Methods: According to the specific sequence of influenza A (M) gene of influenza A, a circularized primer was designed to form a single circular DNA loop under the action of polymerase and ligase. Double primers were used to amplify the RCA index. The amplified product was separated by solid phase at 96 Genetically labeled probe hybridization, enzyme immunoassay combined with color judgment. Results: 100 samples of influenza-like illness were detected by this method. The results were in good agreement with those of fluorescence PCR with the accuracy of 97%, the specificity of 100% and the sensitivity of 94%. CONCLUSION: The product is a large number of DNA repeat units by constant temperature amplification of RCA. Combined with enzyme-linked immunosorbent assay (ELISA), the rapid screening of influenza A method is simple and rapid and has the promotion significance.