基于微流控生物传感技术的多种特殊病原体现场快速一体化检测

Rapid detection of infectious disease is of great importance for the scientific control and prevention of epidemic. Nowadays, fulminating infectious disease epidemics break out frequently to significantly threaten on the health of human being. However, current detecting techniques have many limitations such as complicated protocol, high false positive rate, bulky instrumentation, and difficulties in multiplexed detection in field. Based on our previous work, we propose an integrating assay method for the field and multiplexed detection of fulminating infectious diseases. Detection targets are focused on the pathogens of fulminating infectious diseases including Ebola, Lassa fever, Marburg hemorrhagic fever, Rift valley fever, yellow fever and so on. In this project, we utilize technical principles of biochemistry, material chemistry, molecular biology and some other interdisciplines. The superparamagetic nanoparticles are used in the pretreatment of complicated samples, and the electrochemistry biosensor is used to detect the nucleic acid production of direct amplification from RNA. Then, we assemble the above nanomaterials and biosensors on micro-fluidic chip to form a synergistic detection system. Finally, an integrating detection system based the above synergistic assay method and portable equipment will be fabricated for field and multiplexed detection of fulminating infectious disease. It integrates the procedures of nucleic acid extraction, isothermal amplication, sensor detection and signal output into a portable detection platform. With the advantage of convenience, multiplex, high efficiency and sensitivity, the integrating detection system will be suitable for the field and multiplexed detection of fulminating infectious diseases, which will provide technical support for the timely and effective control of such infectious diseases.

传染病快速检测对于疫情的科学防控具有重要意义。本课题立足于烈性传染病疫情频发，严重危害人类健康的现实背景，针对现有检测技术存在操作步骤冗繁、常规DNA等温扩增假阳性率高、检测仪器不易携带，在现场条件下难以多病原检测的问题；在原有工作基础上，以埃博拉、拉沙热、马尔堡出血热、裂谷热和黄热病等烈性传染病病原体为检测对象，综合运用生物化学、材料化学、分子生物学等交叉学科技术原理，通过合成超顺磁性纳米新材料用于样本快速前处理，研发电化学生物传感检测模块用于病原体RNA直接等温扩增结果快速判读，研究检测过程各单元操作在微流控芯片的系统集成和协同效应；研制可在现场条件快速筛查多种烈性病原体的检测新技术和便携设备，具备“核酸提取-等温扩增-传感检测-信号输出”的一体化检测功能，具有快速、灵敏、多通道、操作简单等优点；实现多种重要烈性传染病病原体的现场快速筛查，为此类传染病的及时有效防控提供技术支持。

传染病现场快速检测对于疫情的科学及时防控具有重要意义。除了新型冠状病毒疫情，埃博拉、猴痘等非洲地区的传染病频发并不断外溢，严重危害全球人类健康，亟需发展现场适用的检测技术设备。本项目综合运用纳米材料、生物传感、微流控芯片等交叉学科技术，分别构建了基于假病毒载体的病原体核酸检测参考品及模拟样本，建立了基于单分散纳米磁珠的现场快速前处理方法；根据待检测病原基因组特点，设计了等温扩增引物和探针并优化了检测反应条件，建立了传染病病原体核酸等温扩增方法和生物传感检测技术；将各个单元操作兼容整合到10通道微流控芯片上，构建微流控一体化检测设备以及配套检测试剂，并研制便携式微流控多重病原检测箱，实现了埃博拉、马尔堡、猴痘等非洲地区传染病以及新冠、流感等呼吸道传染病病毒的现场快速检测，模拟或临床样本实验结果表现出很高的特异性和准确性，检测限达到10-1000拷贝/mL，优于荧光定量PCR法。建立了基于丝网印刷电极的多通道电化学生物传感检测技术，并构建了流动相倏逝波光纤生物传感检测系统，实现了新冠病毒和甲型流感病毒的现场检测；基于集成化的设计和光纤传感实现了低成本的自动化检测，灵敏度和特异性均优于商业化的ELISA和胶体金试纸，对SARS-CoV-2奥密克戎临床样本的特异性和灵敏度分别达到100%和94%，整个检测过程仅需10分钟。该传感检测设备尺寸小巧、操作简单、传感界面可再生，有望用于传染病现场快速筛查，有助于资源有限地区的疫情应急防控。总之，本项目研发的基于微流控纳米生物传感技术的便携式传染病检测技术和设备，具有灵敏、简便、适合现场的优势，并在疫情处置现场获得应用，显著提高了疫情防控的时效性和准确性。