Nanobody-Based Electrochemical Biosensor for Real-Time Detection of Aerosolized SARS-CoV2

基于纳米抗体的电化学生物传感器，用于实时检测气溶胶 SARS-CoV2

• 批准号: 10264330
• 负责人: John R Cirrito
• 金额: $ 43.33万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-21 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10264330
• 关键词: 2019-nCoV; Aerosols; Affinity; Air; Alzheimer' s Disease; American; Amino Acids; Amyloid beta-Protein; Antibodies; Area; Atmosphere; Binding; Biosensor; Blood; Breathalyzer Tests; Breathing; COVID-19; Cell Nucleus; Cessation of life; Collection; Communities; Contact Tracing; Coronavirus; Data; Detection; Development; Devices; Diagnosis; Diagnostic; Dimensions; Disease; Disinfection; Dust; Electrodes; Electrons; Environment; Environmental Risk Factor; Future; Goals; Hospitalization; Hour; Human; Humidity; Individual; Inhalation; Laboratories; Lead; Liquid substance; Llama; Longevity; Measurement; Measures; Membrane; Membrane Proteins; Methodology; Monitor; Mucous Membrane; Nebulizer; Nerve Degeneration; Nose; Observational Study; Particulate Matter; Performance; Persons; Pharyngeal structure; Physical condensation; Property; Protein Dynamics; Proteins; Rapid diagnostics; Recombinants; Reporting; Respiration; Saliva; Sensitivity and Specificity; Signal Transduction; Soot; Specificity; Surface; Symptoms; Technology; Temperature; Testing; Tetrahydrocannabinol; Time; Touch sensation; Tube; Tyrosine; United States; Viral; Virus; aerosolized; air sampling; antigen test; atmospheric aerosols; atmospheric conditions; base; carbon fiber; design; detection limit; detector; mortality; nanobodies; novel; oxidation; pandemic disease; particle; pathogen; pollutant; prototype; residence; respiratory; sensor; transmission process; web site

ABSTRACT/PROJECT SUMMARY Coronavirus 2019 (COVID-19) has afflicted 6.2 million Americans and killed 190,000 as of early September 2020 (WHO website); a roughly 3% mortality. Between a shortage in testing and unidentified asymptomatic individuals, the actual number of those infected could be 6 to 24-fold higher than that reported. SARS-CoV-2 (CoV-2), the virus underlying the disease, results in a range of symptoms; in select cases a severe respiratory illness that impedes breathing that could lead to hospitalization and death. CoV-2 is transmitted person-to- person via inhalation of the virus through mucosal membranes of the nose and throat from transfer after touching a contaminated surface or by inhaling aerosolized virus. Unfortunately, COVID-19 is likely to be prevalent well into 2021 and beyond. We must increase our ability to test for CoV-2. First, testing is needed to diagnose individuals that are symptomatic or asymptomatic to reduce community spread. And second, monitoring gathering areas for airborne virus that could inform the decision to shutdown a space or implement disinfection and mitigation of an area. We propose to use an electrochemical biosensor in two detection devices, 1) a diagnostic breathalyzer for instant detection of CoV-2 and 2) an airborne detector for real-time, continuous surveillance of a large space. We have developed a novel ultra-sensitive, antibody-based electrochemical biosensor to detect CoV-2 repeat binding domain (RBD) spike protein. The technology is based on a micro-immunoelectrode (MIE) biosensor pioneered by the Cirrito laboratory to study protein dynamics in the setting of neurodegeneration (2,3). The biosensor uses voltammetry to measure the oxidation of tyrosine amino acids; oxidation is the release of electrons that the biosensor measures as a change in current. Antibodies are covalently attached to the electrode surface to provide selectivity. Our prototype CoV-2 biosensor is sensitive to 2 femtogram/ml, compared to several current CoV-2 antigen tests that are sensitive to the low picogram/ml range. The proposal will first (Aim 1) optimize our CoV-2 biosensor to detect CoV-2 viral particles, as well as test several parameters to increase sensitivity and longevity. Aim 2 will build a test breathalyzer that will utilize a nebulizer to generate virus laden air containing aerosol droplets similar to a breath that contain defined concentrations of CoV-2 viral particles. Aim 3 will test the airborne biosensor in a realistic environment. Co-I Chakrabarty’s laboratory has unique capabilities of mimicking real-world environmental conditions, especially in the context of atmospheric aerosols, necessary for testing and optimizing the biosensor’s performance for field deployment. Atmospheric conditions include relative humidity (RH) and temperature, as well as common airborne pollutants found indoors. Finding novel means to detect the CoV-2, as well as create a platform to detect other and future pathogens, would enable us to limit the viral spread throughout the community in the current and future pandemics.

摘要/项目摘要 截至9月初，冠状病毒2019(新冠肺炎)已困扰620万美国人，导致19万人死亡 2020年(世卫组织网站)；死亡率约为3%。在检测短缺和不明症状之间 就个人而言，实际感染人数可能是报告人数的6至24倍。SARS-CoV-2 这种疾病背后的病毒(CoV-2)会导致一系列症状；在某些情况下，严重的呼吸道疾病 阻碍呼吸的疾病，可能导致住院和死亡。CoV-2在人与人之间传播 人通过鼻和喉部粘膜吸入病毒后转移 接触受污染的表面或吸入雾化病毒。不幸的是，新冠肺炎很可能是 一直流行到2021年及以后。 我们必须提高检测CoV-2病毒的能力。首先，需要进行测试来诊断符合以下条件的个人 有症状或无症状，以减少社区传播。第二，监测聚集区 空气传播的病毒，可告知决定关闭空间或实施消毒和缓解 一个区域。我们建议在两个检测设备中使用电化学生物传感器：1)诊断 用于即时检测CoV-2的呼气测定仪和2)用于实时、连续监测的气载探测器 一个很大的空间。 我们开发了一种新型的基于抗体的超灵敏电化学生物传感器来检测CoV-2 重复结合结构域(RBD)刺突蛋白。这项技术基于微型免疫电极(MIE)。 由Cirrito实验室首创的生物传感器，用于研究神经退行性变环境中的蛋白质动力学(2，3)。 生物传感器使用伏安法来测量酪氨酸氨基酸的氧化；氧化是 生物传感器测量为电流变化的电子。抗体以共价方式附着在 电极表面提供选择性。我们的原型CoV-2生物传感器对2毫微克/毫升敏感， 与目前几种对低皮克/毫升范围敏感的CoV-2抗原检测相比， 该提案将首先(目标1)优化我们的CoV-2生物传感器，以检测CoV-2病毒颗粒，以及测试 增加敏感度和寿命的几个参数。Aim 2将建造一个测试酒精测定器，它将使用 喷雾器产生含有病毒的空气，含有气雾滴，类似于呼吸，含有限定的 CoV-2病毒颗粒的浓度。目标3将在现实环境中测试空气中的生物传感器。CO-I 查克拉巴蒂的实验室具有独特的模拟真实世界环境条件的能力，特别是 在大气气溶胶的背景下，测试和优化生物传感器的性能 野战部署。大气条件包括相对湿度(RH)和温度以及常见的 在室内发现的空气污染物。 寻找新的方法来检测CoV-2，以及创建一个检测其他和未来的平台 病原体，将使我们能够在当前和未来限制病毒在整个社区的传播 大流行。