Direct bioelectronic detection of SARS-CoV-2 from saliva using single-molecule field-effect transistor array

使用单分子场效应晶体管阵列直接生物电子检测唾液中的 SARS-CoV-2

• 批准号: 10266395
• 负责人: Kenneth L Shepard
• 金额: $ 81.73万
• 依托单位: QUICKSILVER BIOSCIENCES, INC.
• 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/10266395
• 关键词: 2019-nCoV; Address; Binding Proteins; Biological Assay; Biological Sciences; COVID test; COVID-19 detection; COVID-19 diagnostic; COVID-19 pandemic; COVID-19 testing; Carbon Nanotubes; Clinical; Communicable Diseases; Complex; Defect; Detection; Development; Devices; Diagnosis; Diagnostic tests; Differential Diagnosis; Electrolytes; Gold; Hour; ImProv; Immobilization; Immunoassay; Individual; Infection; Infectious Agent; Influenza; Kinetics; Laboratories; Llama; Membrane Proteins; Nanotubes; New York; Nucleic Acid Amplification Tests; Nucleocapsid Proteins; Optics; Phase; Polymerase Chain Reaction; Population Surveillance; Preparation; Process; Proteins; Public Health; Quantitative Reverse Transcriptase PCR; Reader; Reagent; Recombinants; Research Personnel; Reverse Transcription; SARS-CoV-2 transmission; Saliva; Sampling; Semiconductors; Sensitivity and Specificity; Site; Small Business Innovation Research Grant; Specificity; Specimen; Structural Protein; Symptoms; Technology; Testing; Time; Training; Transistors; Universities; Vaccines; Viral; Viral Load result; Viral Proteins; Virus; antibody detection; antigen detection; antigen test; base; biobank; cost; design; diagnostic assay; env Gene Products; experimental study; improved; innovation; metal oxide; molecular array; molecular diagnostics; multiplex detection; nanobodies; particle; pathogen; point of care; point of care testing; programs; protein structure; saliva sample; sensor; single molecule; temporal measurement; tool; transmission process; viral RNA; viral detection

Direct bioelectronic detection of SARS-CoV-2 from saliva using single-molecule field-effect transistor array Nucleic acid tests have become the gold-standard for diagnostic testing for COVID-19, usually performed in specialized laboratories. Most are based on reverse-transcription quantitative polymerase chain reaction (qRT-PCR). The time required for specimen transport and processing results in a turnaround time that is typi- cally several days. The few rapid (<1 hour) point-of-care (POC) tests are more expensive, still require sample preparation and specialized reagents, and do not have the throughput needed for population surveillance. Di- rect testing for the virus, which also reduces requirements for multiple reagents, is a necessary step to improv- ing diagnostic testing. While four such antigen tests have been approved for detection of SARS-CoV-2 based on immunoassays to the N protein, sensitivity is limited and no quantitation of viral load is possible. We will address this gap by using DiagnostikosTM, an in-development rapid POC platform for direct, real- time, multiplexed, quantitative bioelectronic detection of biomolecules that employs an all-electronic detection device that functions at the single-molecule level. These single-molecule field-effect transistors (smFETs) are arrayed on a complementary metal-oxide-semiconductor (CMOS) integrated circuit chip. Chips will interface with an envisioned USB-stick-form-factor reader device. Robust single-domain antibodies, known as nanobod- ies and immobilized on these devices, are used for sensitive detection of viral particles and viral debris. The use of multiple nanobodies for a single protein and nanobodies for different proteins in a single assay allows for significant improvements in specificity. Nanobodies will be specific for one or more of the four major struc- tural proteins in SARS-CoV-2; the nucleocapsid (N) protein engulfing the viral RNA, the spike (S) protein, the membrane (M) protein and the envelope (E) protein. No sample preparation or specialized reagents are re- quired for detection, and the device will be designed to operate with saliva, which has very recently been shown to be a reliable medium for detecting SARS-CoV-2. Individual sensor chips can be manufactured at a cost of $35. With the addition of other nanobodies, these large dense arrays can also allow detection of many pathogens in a single test. In this Direct-To-Phase-2 SBIR program we will pursue several key innovations that are required to make such a platform possible, including isolation of nanobodies for key structure proteins of SARS-CoV-2 (Specific Aim 1), development of the smFET platform for antigen detection (Specific Aim 2), development of large CMOS arrays of these smFET devices (Specific Aim 3), and verification of detection in increasingly complex samples up to and including clinical samples (Specific Aim 4). This project is a partnership between university researchers who developed the smFET technology and a venture-based start-up venture, Quicksilver Biosci- ences, spun out to commercialize smFET technology and develop smFET/CMOS arrays for molecular diag- nostic applications.

唾液中SARS-CoV-2的直接生物电学检测 使用单分子场效应晶体管阵列 核酸检测已成为新冠肺炎诊断测试的黄金标准，通常进行 在专门的实验室里。大多数都是基于逆转录定量聚合酶链式反应 (qRT-PCR)。样品运输和处理所需的时间导致周转时间通常是- 凯莉有几天了。少数几种快速(&lt；1小时)护理点(POC)测试更昂贵，仍需要样本 制备和专门的试剂，没有人口监测所需的产量。迪- 对病毒的RECT检测也减少了对多种试剂的要求，这是改善这种情况的必要步骤。 ING诊断测试。虽然已经批准了四种这样的抗原测试来检测基于SARS-CoV-2的病毒 在对N蛋白的免疫分析中，敏感性是有限的，无法对病毒载量进行定量。 我们将通过使用DiagnostikosTM来解决这一差距，这是一个正在开发的快速PoC平台，可直接、实时地 使用全电子检测的生物分子的时间、多路复用、定量生物电子检测 在单分子水平上起作用的装置。这些单分子场效应晶体管(SmFET)是 阵列在互补的金属氧化物半导体(CMOS)集成电路芯片上。芯片将连接到 具有设想的USB棒形状因数读取器设备。强健的单域抗体，被称为纳米棒- 固定在这些设备上的离子交换膜用于敏感地检测病毒颗粒和病毒碎片。这个 单个蛋白质的多个纳米小体和不同蛋白质的纳米小体在一次检测中允许 在特异度方面的显著改进。纳米体将针对四种主要结构中的一种或多种- SARS-CoV-2中的结构蛋白；吞噬病毒核糖核酸的核衣壳(N)蛋白，尖峰(S)蛋白， 膜(M)蛋白和包膜(E)蛋白。不需要样品制备或专门的试剂- 需要进行检测，该设备将被设计为与唾液一起工作，这是最近被 证明是检测SARS-CoV-2的可靠介质。单个传感器芯片可以在一个 费用是35美元。随着其他纳米体的加入，这些大而密集的阵列也可以检测到许多 一次测试中的病原体。 在这个直接到第二阶段的SBIR计划中，我们将追求几项关键的创新，这些创新需要 这样的平台是可能的，包括分离SARS-CoV-2(特异性)关键结构蛋白的纳米抗体 目标1)，开发用于抗原检测的smFET平台(特定目标2)，开发大型 这些smFET器件的cmos阵列(具体目标3)，并在日益复杂的检测中进行验证 样本达到并包括临床样本(具体目标4)。该项目是两所大学之间的合作项目 开发smFET技术和基于风险的初创企业QuickSilver Biosi的研究人员-- 将smFET技术商业化，并开发用于分子诊断的smFET/cmos阵列。 不可靠的应用程序。