Ultrasensitive, Rapid, Amplification-Free RNA Virus Detection Using Nanodimer-Based Nucleic Acid Target Sequence Recognition

核糖核酸（RNA）病毒是对公共卫生和全球经济的主要威胁。从普通感冒到危及生命的出血热，许多人类疾病都是由RNA病毒引起的。在过去几十年里，人类免疫缺陷病毒、埃博拉病毒、登革热病毒以及最近的冠状病毒（COVID-19）等RNA病毒给全球卫生和全球经济带来了重大负担。在人类标本和环境中快速和高灵敏度地检测RNA病毒的存在对于跟踪病毒污染的速度、防止快速传播、建立预防措施和及时给予治疗至关重要。流行的基于聚合酶链反应（PCR）的病毒检测需要较长的周转时间，并且需要专门的实验室仪器和训练有素的人员。本项目旨在建立一种新型的生物传感微系统，可以提供超灵敏、快速、可靠的RNA病毒检测，而不需要PCR扩增。该项目将产生一种便携式、快速、经济的RNA病毒检测方法，用于即时检测、流行病预防、反生物恐怖主义和环境监测。此外，这项跨学科研究将支持阿克伦大学研究生和本科生的多元化发展。研究结果还将支持阿克伦大学的几个微系统和生物材料研究生课程。项目的技术范围分为多个任务：1)研制一种表面声学预过滤芯片，快速去除连续流动中可能导致传感通道堵塞的微级杂质；2)研究一种基于独特的电扩散电泳效应的新型纳米粒子聚焦方法，该方法可以显著提高病毒粒子浓度，从而在数秒内输出数千次信号；3)研究一种基于创新的纳米二聚体解离实验的病毒检测芯片。4)研究应用于电阻脉冲传感器阵列的独特信号多路复用，这将实现RNA病毒的高通量数字检测，最终5)将三个芯片集成到一个微系统中，并展示其使用灭活模型病毒快速检测超低丰度RNA病毒的实用性。通过此次研究，将推进目前的尖端技术难以实现的超高分辨率和数字化的病毒检测领域。它还将为下一代高通量现场生物传感系统的发展产生许多新的知识和创新。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Ribonucleic acid (RNA) viruses are a major threat to public health and global economy. Numerous human diseases, from the common colds to life-threatening hemorrhagic fevers are caused by RNA viruses. In the past decades, RNA viruses such as human immune deficiency virus, Ebola virus, dengue virus, and most recently coronavirus (COVID-19) impose significant burdens on global health and global economy. Rapid and high-sensitivity detection of the presence of RNA viruses in human specimens and the environment is crucial to track the rate of viral contamination, prevent fast spreading, and establish preventive measures and timely administration of treatments. The popular polymerase chain reaction (PCR)-based virus detection takes lengthy turn-around time and requires specialized laboratory instruments and trained personnel. This project aims to create a novel biosensing microsystem that can provide ultra-sensitive, rapid and reliable detection of RNA viruses with no need of PCR amplification. The project will result in a portable, rapid, economical RNA virus detection method for point-of-care-testing, pandemic prevention, anti-bioterrorism, and environmental monitoring. Furthermore, this cross-disciplinary research will support the development of a diverse cohort of graduate, undergraduate students at the University of Akron. The research results will also support several graduate-level courses on microsystems and biomaterials at the University of Akron.The technical scope of the project are divided into multiple tasks: 1) create a surface acoustic pre-filtering chip that will rapidly remove microscale impurities in continuous flow, which would otherwise cause clogging of the sensing channels, 2) research a novel nanoparticle focusing method based on unique electro-diffusio-phoresis effect, which can significantly increases the virion concentration and thus output signals thousands of times within seconds, 3) research a virus detection chip based on an innovative nanodimer dissociation assay, which in principle, can detect viruses with single virus resolution, 4) research unique signal multiplexing applied on a resistive pulse sensor array, which will enable high throughput, digital detection of RNA viruses, and ultimately 5) integrate the three chips into a microsystem and demonstrate its utility for rapid detection of ultra-low abundance RNA viruses using an inactivated model virus. The research will advance the field of virus detection with ultra-high resolution and digitization capability, which are difficult to achieve using current state-of-the-art methods. It will also generate numerous new knowledges and innovations for the development of the next generation high-throughput, onsite biosensing systems in general.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.