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，2019 年冠状病毒 (COVID-19) 已影响 620 万美国人，并导致 19 万人死亡 2020 (WHO website); a roughly 3% mortality.检测不足和身份不明的无症状感染者之间 个人而言，实际感染人数可能是报道的 6 至 24 倍。 SARS-CoV-2 (CoV-2) 是导致疾病的病毒，会导致一系列症状；在某些情况下，会出现严重的呼吸道疾病 阻碍呼吸的疾病，可能导致住院和死亡。 CoV-2 可以在人与人之间传播 人通过鼻子和喉咙粘膜吸入病毒，传播后 接触受污染的表面或吸入雾化病毒。不幸的是，COVID-19 很可能 流行到 2021 年及以后。 我们必须提高检测 CoV-2 的能力。首先，需要进行测试来诊断患有以下疾病的个体： 有症状或无症状，以减少社区传播。其次，监控聚集区域 空气传播的病毒可以为关闭空间或实施消毒和缓解措施的决定提供依据 an area.我们建议在两种检测设备中使用电化学生物传感器，1）诊断设备 用于即时检测 CoV-2 的呼气分析仪和 2) 用于实时、连续监测的机载检测器 a large space. 我们开发了一种新型超灵敏、基于抗体的电化学生物传感器来检测 CoV-2 重复结合域 (RBD) 刺突蛋白。该技术基于微免疫电极（MIE） 生物传感器由 Cirrito 实验室首创，用于研究神经退行性疾病中的蛋白质动力学 (2,3)。 该生物传感器采用伏安法测量酪氨酸氨基酸的氧化情况；氧化是释放 生物传感器测量的电子作为电流的变化。抗体共价连接至 电极表面提供选择性。我们的原型 CoV-2 生物传感器对 2 飞克/毫升敏感， 与目前对低皮克/毫升范围敏感的几种 CoV-2 抗原测试相比。 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，并测试 几个参数可提高灵敏度和寿命。目标 2 将构建一个测试呼吸分析仪，该分析仪将利用 雾化器产生充满病毒的空气，其中含有气溶胶液滴，类似于含有特定成分的呼吸 CoV-2 病毒颗粒的浓度。 Aim 3 将在现实环境中测试机载生物传感器。 Co-I 查克拉巴蒂的实验室具有模仿现实世界环境条件的独特能力，尤其是 在大气气溶胶的背景下，对于测试和优化生物传感器的性能是必要的 field deployment.大气条件包括相对湿度 (RH) 和温度，以及常见的 室内发现的空气污染物。 寻找检测 CoV-2 的新方法，并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 流行病。