基于荧光纳米颗粒及试纸条对痕量细菌进行快速测定的研究

Bacteria detection is highly related to a lot of things including the testing of food safety and disease diagnosis.It has very important practical significance and great study value to develop sensitive, convinent, and speedy assay for trace bacteria detection especially with the ability of real-time and onsite analysis. Based on the previous study and focus on bacteria detection, the applicant would like to develop an immunosensing method with high sensitivity and speed for trace bacteria detection. This assay will be build up by using core-shell fluorescent silica nanoparticles, lateral flow test strip, immunological techniques and biological labeling methods as well as specific recognition and binding between antigen and atibody. In this proposal, the superior signal brightness and high photostability of fluorescent silica nanoparticles are combined with the promising advatanges of lateral flow test strip including short assay time, high specificity, small amount of sample requirement, and easily handling format. This platform takes full advantages of fluoresenct silica nanoparticles and lateral flow test strip together for quickly screening of trace bacteria,which would lay a foundation in theory and practice to build up the rapid analyzing abiligy and also hold wide application prospects as well as far-reaching study significance.

食品安全检验、疾病诊断等领域都涉及到细菌的检测。发展高灵敏、便捷、快速的细菌检测方法，尤其是能够现场、实时地对痕量致病菌进行快速测定具有非常重要的现实意义和研究价值。本项目拟在申请人已有的工作基础之上，仅仅围绕细菌检测这一研究领域，利用基于核壳型荧光纳米颗粒建立的生物纳米信号放大技术，结合层析试纸条技术、生物标记技术和免疫学技术，以抗原-抗体的特异性识别与结合作用为基础，发展高灵敏的免疫生物传感试纸条检测方法，从而实现对痕量细菌的快速测定。该方法有效利用荧光纳米颗粒荧光强度高、抗光漂白性能强等多种特点，充分发挥层析试纸条技术操作简便、快速、特异性好、以及所需样品量小等诸多优点，将二者有机结合并应用于对痕量细菌的快速筛查，为形成实际快速检测能力奠定了理论和实践基础，具有广阔的应用前景和深远的研究意义。

试纸条技术具有快速、操作简便、成本低等诸多优点，近年来已经引起了越来越广泛的关注；量子点作为一种性能优良的新型荧光纳米材料，在分析检测中发挥着愈来愈重要的作用。本课题充分发挥二者的优势，发展了一种基于量子点荧光纳米颗粒的免疫层析试纸条平台，以破伤风抗体和狂犬抗体为检测对象，取得了较为满意的实验结果。首先，我们制备了一系列性能优良的量子点荧光纳米颗粒，包括CdTe量子点，CdSe/ZnS量子点以及环境友好型Cu:Zn−In−S/ZnS量子点。在此基础之上，采用EDC/NHS连接法，优化各项实验参数，在量子点表面成功修饰上了羊抗人IgG（FC）抗体，建立了较为成熟的荧光纳米颗粒抗体修饰方法，并进一步与试纸条技术相结合，对破伤风抗体进行了快速灵敏测定。为了避免Cd2+离子带来的二次污染，我们重点研究了基于环境友好型Cu:Zn−In−S/ZnS量子点的免疫层析试纸条技术，并对破伤风抗体进行了测定。该方法特异型好，灵敏度高（0.001 IU/mL），相比于胶体金试纸条提升了10倍以上，检测便捷、快速，整个测定可在15分钟内完成，且表现出较好的稳定性，4℃下可保存3个月。接下来，我们进一步将该平台予以拓展，同时对破伤风抗体和狂犬抗体进行了快速测定，也取得了较好的实验结果。该平台的建立，将促进试纸条技术以及荧光纳米颗粒标记技术的融合与发展，具有广阔的应用前景，可进一步应用于医疗检验、食品安全等领域。