一种基于微流控免疫磁珠快速检测登革热抗原的方法

A new method for rapid detection of dengue antigen based on microfluidic immune magnetic beads

目的建立一种灵敏、快速、方便的检测登革热抗原新方法,为登革热的临床诊断提供帮助。方法建立了一种基于微流控免疫磁珠快速检测登革热抗原检测方法。使用Solidwork软件设计微流控芯片,采用机械加工法和化学封接方式制备微流控芯片;利用化学偶联反应制备登革热抗体免疫磁珠;在微流控芯片载体上,以HRPTMB-H2O2为显色体系,利用双抗体夹心法原理检测登革热NS1抗原;最后,同时应用该方法与传统ELISA试剂盒对57例临床样品进行检测,分析该方法的正确度,并对两种方法的优缺点进行比较。结果微流控免疫磁珠方法整个检测过程仅需20 min,反应体系只需磁珠量10μg和样品量10μL;验证实验中,该方法可以将阴性和阳性明显区分开,阳性样本有显色,阴性、空白样本均无显色;性能上,该新型ELISA方法与传统ELISA方法结果阴阳性符合率达到100%。结论该新型ELISA检测方法具有简便、快速、节省试剂和所需样本少、灵敏度高、稳定性好、准确性高等优点,可以用于登革热抗原检测。

Objective To establish a sensitive,rapid and convenient method for the detection of dengue antigen and assist clinical diagnosis of dengue.Methods In this paper,we developed a rapid detection method for dengue antigen based on microfluidic immune magnetic beads.Solidwork software was used to design microfluidic chip,which was prepared by mechanical processing and chemical sealing.Immunomagnetic beads of dengue antibody were prepared by chemical coupling reaction.Using HRP-TMB-H2O2 as color system,dengue NS1 antigen was detected on microfluidic chip carrier by double antibody sandwich method.Finally,57 clinical samples were tested by the novel method and traditional ELISA kit,and the accuracy of the method was analyzed,and the advantages and disadvantages of the two methods were compared.Results20 minutes was needed to detect dengue NS1 antigen by using the novel ELISA method,and the reaction system only needed 10μg beads and 10μL samples.In the verification experiment,the method could distinguish the negative from the positive obviously.The positive sample had color rendering,while the negative and blank samples had no color rendering.In terms of detection performance,the coincidence rate between the new ELISA method and the traditional ELISA method reached 100%.Conclusion The novel ELISA detection platform had the advantages of simple,rapid,reagent and sample saving,high sensitivity,good stability and high accuracy,and could be used for the detection of dengue antigen.