表面等离子体共振传感器技术在检测甲型流感病毒中的应用研究

   滕娟1，徐云庆2，顾大勇2，史蕾2，刘春晓2，赵纯中2，杨燕秋2，孙秋香1，欧青叶2，季明辉2

    1.中山大学公共卫生学院，广东 广州 510080；2.深圳国际旅行卫生保健中心

    摘要：目的建立一种基于表面等离子体共振传感器技术检测人类甲型流感病毒的方法，并进行初步诊断应用。方法分别设计基于甲型流感M基因的引物和探针，制备RT-PCR扩增产物，优化并确定基于传感技术的实验条件，建立高灵敏度和高特异度的表面等离子体共振传感器方法。结果缓冲液的盐离子强度和pH值是影响杂交效果的关键因素，对其优化后标准曲线参数良好，甲型流感病毒标准品的Ru值与其相应梯度稀释标准品浓度呈良好的线性关系(R2 > 0.99)，同时，该传感器在200 mmol/L氯化钠的磷酸盐缓冲液中可以有效区分DNA的完全匹配与单碱基错配，以乙型流感探针作为质控，该探针并不与甲型流感扩增产物发生特异性杂交。结论成功建立了高灵敏度和高特异度的甲型流感病毒表面等离子体共振传感器检测方法，为临床诊断或感染病例提供了技术平台。

    关键词：表面等离子共振传感技术；甲型流感病毒；检测；应用

    中图分类号：R511.7　文献标识码：B

    Surface plasmon resonance biosensor fordetecting

    influenza A virus in human

    TENG Juan*, XU Yun-qing, GU Da-yong, SHI Lei,LIU Chun-xiao, ZHAO Chun-zhong, YANG Yan-qiu,

    SUN Qiu-xiang, ’OU Qing-ye, JI Ming-hui

    *Sun Yat-sen University, School of PublicHealth, Guangzhou, Guangdong 510080, China

    Abstract:   Objective  To establish a detection method for influenza-A virus based onsurface plasmon resonance (SPR) biosensor.  Methods    Design respectively primers and probe ofinfluenza A virus based on M gene and prepare RT-PCR product. Asensitive and specific SPR method was established andoptimized.    Results     Saltionic strength and pH of the buffer were key elements thatinfluence hybridization. The standard curve showed that thereflectivity shifts and PCR product concentration had a good linearrelationship (R2 > 0.99). In addition, the prepared biosensormethod can effectively discriminate perfect complementary and otherthree types of mismatch: base substitute, base insertion and basedeletion in 200 mmol/L NaCl-PBS buffer. Conclusion    The established SPR biosensor methodis sensitive and specific, which could be used as a prospectiveassay for detecting influenza-A virus. This system offers a goodplatform for diagnosing other causative organism.

    Key words:   Surface plasmonresonance biosensor； Influenza A virus; Detection; Application

    《中国国境卫生检疫杂志》