RAPID: Microelectrode Array Sensors for SARS-CoV-2 and Other RNA Viruses

RAPID：用于 SARS-CoV-2 和其他 RNA 病毒的微电极阵列传感器

• 批准号: 2034498
• 负责人: Gangli Wang
• 金额: $ 19.76万
• 依托单位: Georgia State University Research Foundation, Inc.
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2034498&HistoricalAwards=false
• 关键词: RAPID; Microelectrode; Array; Sensors; SARS

There is an urgent need for accurate measurement tools to test for the presence of the novel coronavirus SARS-CoV-2 in samples, so that this information may be used for diagnosing the novel coronavirus infectious disease 2019 (COVID-19). Obtaining test results on the timeline of minutes is essential to limiting spread of the virus and helping individuals and communities make appropriate decisions. Alarmingly high rates of missed virus presence (false negatives) have been found using some of the current fast testing routes. Those routes typically rely on sample amplification that is time-consuming and resource-intensive, and due to their complicated multi-step sample treatments, those routes are more likely to generate false negatives. The immediate goal of this project, led by Dr. Gangli Wang and colleagues at Georgia State University, is to design and develop electrochemical sensors whose "switched-on" response to SARS-CoV-2 presence results from target interactions with specific chemical species in the nucleic acids of the virus. The response is rapidly intensified to allow for trace-level detection of virus, potentially within minutes. The design principle provides several benefits, such as drastically simplified sample pretreatment, fast turn-around time, and simplified one-step detection with greatly decreased false negative outcomes. These essential qualities pave the way for future point-of-use applications that go beyond SARS-CoV-2. The project provides a rare opportunity for fundamental measurement science to address an ongoing societal and global crisis through cutting-edge research. In addition, the project provides a rich and diverse educational experience for students and postdoctoral fellows at Georgia State University, an institution with large numbers of minority students and students from low-income backgrounds. Scientific concepts and results are disseminated through classroom teaching, local events, attendance at scientific meetings, and publications. Additional educational and outreach activities provide opportunities to many in the Atlanta Metro Area and beyond through an on-going NSF undergraduate research program and the Atlanta Science Festival, as well as several other recently established mentor programs at Georgia State University. The enabling intellectual merit is the in-situ signal amplification mechanism in signal-on electrochemical sensors via redox cycling. The novel design drastically improves the lower limit of detection (LoD) to be competitive with and possibly surpass that of existing methods and eliminates the need for error-prone, multi-step sample treatments, such as enrichment, labeling, or amplification. The concept, employed in single-entity electrochemistry, is fundamentally different from the sample amplification strategy adopted in most nucleic acid analysis tools being used. In principle, the LoD is anticipated to approach utmost levels, i.e., single copies of ribonucleic acid (RNA) where the response time is correlated through statistical analysis. The sensor specificity to the SARS-CoV-2 specific RNA sequence(s) is based on their recognition by electrode-immobilized probes. With key components pre-assembled as integral parts of the sensor, near-zero background may be established, which is the prerequisite to improve the LoD and resolve binding of single nucleic acids. Binding with the target sequence turns on the designed electron-transfer pathway: the redox molecule on the recognition probe is then abled to repeatedly mediate electron transfers between the sensor electrode and the co-reactants in solution. The signal-on mechanism further mitigates nonspecific adsorption concerns. Individual sensors are assembled into arrays to simultaneously detect multiple target RNAs. The multiplex signal readout design represents another intellectual merit for enhanced sensor performance. The mechanistic insights from cyclic voltammetry and low LoD achieved with pulse voltammetry are envisioned as critical for the formulation of optimal and generalizable measurement methods and protocols. Synthetic samples, common biomatrices, viral RNA extractions, and virus-infected cell lysates are selected to establish calibration profiles and detection strategies. Comparison of the results with those from laboratory analysis techniques, such as reverse-transcription polymerase chain reaction, provides further validation of the sensor efficacy. In addition to offering qualitative Yes/No answers for fast testing, the sensor approach may provide quantitative information in absolute concentrations over a wide dynamic range, an outcome of great value to the COVID-19 pandemic, as well as to other on-going and future needs in the biological sensing community.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.

迫切需要准确的测量工具来检测样本中是否存在新型冠状病毒SARS-CoV-2，以便这些信息可以用于诊断新型冠状病毒传染病2019年(新冠肺炎)。在几分钟的时间线上获得检测结果对于限制病毒的传播和帮助个人和社区做出适当的决定至关重要。使用目前的一些快速检测路线，已经发现了令人震惊的高漏检率(假阴性)。这些方法通常依赖于耗时和资源密集型的样本扩增，而且由于其复杂的多步骤样本处理，这些方法更有可能产生假阴性。这项由佐治亚州立大学王刚博士及其同事领导的项目的直接目标是设计和开发电化学传感器，这种传感器对SARS-CoV-2存在的“开启”反应是通过与病毒核酸中特定化学物种的目标相互作用而产生的。反应迅速加强，可能在几分钟内就能进行痕量级别的病毒检测。该设计原理提供了几个优点，如大大简化了样品前处理，快速周转时间，简化了一步检测，大大减少了假阴性结果。这些基本素质为超越SARS-CoV-2的未来使用点应用程序铺平了道路。该项目为基础测量科学提供了一个难得的机会，通过尖端研究来解决正在进行的社会和全球危机。此外，该项目为佐治亚州立大学的学生和博士后研究员提供了丰富和多样化的教育经验，佐治亚州立大学是一所拥有大量少数民族学生和低收入背景学生的机构。科学概念和成果通过课堂教学、当地活动、出席科学会议和出版出版物来传播。其他教育和外展活动通过正在进行的NSF本科生研究计划和亚特兰大科学节，以及佐治亚州立大学最近建立的其他几个导师计划，为亚特兰大大都市区和其他地区的许多人提供了机会。其智能优势在于通过氧化还原循环实现信号导通电化学传感器中的原位信号放大机制。这种新颖的设计极大地提高了检测下限(LOD)，与现有方法竞争，甚至可能超过现有方法，并消除了对易出错的多步骤样品处理的需要，如浓缩、标记或放大。这一概念在单一实体电化学中使用，与目前使用的大多数核酸分析工具中采用的样本放大策略有根本不同。原则上，LOD预计将达到最高水平，即单拷贝核糖核酸(RNA)，其中反应时间通过统计分析进行关联。针对SARS-CoV-2特异性核酸序列(S)的传感器特异性是基于它们被电极固定的探针识别的。将关键部件预先组装成传感器的组成部分，可以建立近零的本底，这是提高LOD和解决单核酸结合的先决条件。与靶序列的结合开启了所设计的电子转移途径：识别探针上的氧化还原分子随后能够重复地调节传感器电极和溶液中的共反应物之间的电子转移。信号开启机制进一步减轻了对非特异性吸附的担忧。单个传感器被组装成阵列，以同时检测多个目标RNA。多路信号读出设计代表了增强传感器性能的另一个智能优点。从循环伏安法和脉冲伏安法获得的低LOD的机械洞察力被认为是制定最佳和可推广的测量方法和协议的关键。选择合成样品、普通生物标记物、病毒RNA提取物和病毒感染细胞裂解物来建立校准曲线和检测策略。将结果与逆转录聚合酶链式反应等实验室分析技术的结果进行比较，进一步验证了传感器的有效性。除了为快速测试提供是/否的定性答案外，传感器方法还可以在广泛的动态范围内提供绝对集中的定量信息，这一结果对新冠肺炎大流行以及生物传感社区其他持续和未来的需求具有重要价值。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智力优势和更广泛的影响审查标准进行评估，被认为值得支持。