DiagnosDisk - a highly sensitive point-of-care test for detecting SARS-CoV-2 antigen in saliva

DiagnosDisk - 一种用于检测唾液中 SARS-CoV-2 抗原的高度灵敏的现场检测

• 批准号: 10285991
• 负责人: James Lai
• 金额: $ 23.33万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-25 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10285991
• 关键词: 2019-nCoV; Acute; Address; Adopted; Antibodies; Antigens; Biological Assay; Blood Plasma Volume; COVID-19; COVID-19 detection; COVID-19 pandemic; COVID-19 patient; COVID-19 screening; Centers for Disease Control and Prevention (U.S.); Cessation of life; Contact Tracing; Coupling; Data; Deposition; Detection; Development; Devices; Diagnosis; Early Diagnosis; Enzyme-Linked Immunosorbent Assay; Equipment; Evaluation; Exhibits; Goals; Gold; Gold Colloid; Human Resources; Hydrophobicity; Image; Immunoassay; Individual; Infection; Lateral; Location; Malaria; Measures; Membrane; Noise; Nucleocapsid; Patients; Plasma; Polymers; Precipitation; Prostate-Specific Antigen; Proteins; Reagent; Reverse Transcriptase Polymerase Chain Reaction; SARS-CoV-2 antigen; SARS-CoV-2 infection; SARS-CoV-2 negative; SARS-CoV-2 positive; Saliva; Salivary; Sampling; Scanning; Sensitivity and Specificity; Signal Transduction; Specimen; Temperature; Testing; Time; Training; United States; Viral Antigens; Viral Load result; Virus; Visual; acute infection; antibody conjugate; antigen binding; antigen detection; design; detection assay; detection limit; hydrophilicity; improved; innovation; medical attention; novel; pandemic disease; point of care testing; rapid test; saliva sample; salivary assay; sample collection; scale up; screening; success; viral RNA

Project Summary/Abstract SARS-CoV-2, a global pandemic, has caused more than 6 million infections and 180,000 deaths in the United States alone during the past 7 months. Both the WHO and CDC emphasize the need for point-of-care (POC) tests to expand diagnosis of acute infections to control the pandemic (i.e. isolating infected individuals). The current gold standard test for diagnosing acute SARS-CoV-2 infection is reverse transcriptase polymerase chain reaction (RT-PCR) with nasopharyngeal samples. The assay is highly sensitive, but requires well-trained personnel for sample collection and locations with specialized equipment. Therefore, the turnaround time varies from one to few days, delaying isolation, contact tracing, and medical attention. These challenges can be addressed with lateral flow immunoassays (LFIA) to detect viral antigens in self-collected saliva samples. Compared to nasopharyngeal specimens, studies have demonstrated that assays with saliva samples are more consistent and the saliva itself may contain higher viral load. However, existing LFIA for detecting salivary antigens exhibit poor sensitivity (≤ 40%) caused by the use of small sample volumes and sample dilution. To achieve timely and accurate screening of suspected COVID-19 cases, this project aims to develop a rapid assay for detecting SARS-CoV-2 antigens in saliva with high sensitivity to meet WHO's target product profile. We hypothesize that using ≥500 µL saliva as the assay sample can significantly improve the sensitivity because a larger specimen volume provides more analytes for detection. The new test will employ temperature-responsive polymer-antibody conjugates in conjunction with DiagnosDisk, a novel flow-through assay device. The approach is innovative in in using enriched saliva specimens for more consistent assay results, enhancing antigen detection by scaling up the saliva specimen volume, and adopting the well- developed sandwich immunoassay for DiagnosDisk with temperature-responsive polymer reagents that simplify the design while maximizing the sensitivity. We will employ polymer-antibody conjugates that efficiently isolate analytes in saliva (Aim 1) and fabricate the DiagnosDisk device to accommodate larger volume saliva specimens to improve the detection limit (Aim 2). The sensitivity of the new rapid assay, which combines polymer-antibody conjugates and DiagnosDisk, will be evaluated using saliva specimens from COVID-19 positive and negative patients (Aim 3).

项目概要/摘要 SARS-CoV-2 是一场全球大流行，已在美国造成超过 600 万人感染和 18 万人死亡 过去 7 个月内仅各州。 WHO 和 CDC 都强调即时护理 (POC) 的必要性 扩大急性感染诊断范围的测试，以控制大流行（即隔离感染者）。这 目前诊断急性 SARS-CoV-2 感染的金标准测试是逆转录酶聚合酶 鼻咽样本的链式反应（RT-PCR）。该检测灵敏度高，但需要经过良好培训 样本采集人员和配备专业设备的地点。因此，周转时间 时间从一到几天不等，导致隔离、接触者追踪和医疗护理延迟。这些挑战可以 可以通过侧流免疫分析 (LFIA) 来检测自采集唾液样本中的病毒抗原。 与鼻咽样本相比，研究表明唾液样本的检测更有效 更一致，唾液本身可能含有更高的病毒载量。然而，现有的 LFIA 用于检测 由于使用小样本量和样本，唾液抗原的灵敏度较差（≤ 40%） 稀释。为了实现及时、准确地筛查 COVID-19 疑似病例，该项目旨在开发 一种快速检测唾液中 SARS-CoV-2 抗原的方法，灵敏度高，符合 WHO 的目标产品 轮廓。我们假设使用≥500 µL唾液作为检测样本可以显着提高灵敏度 因为更大的样本体积可以提供更多的分析物用于检测。新的测试将采用 温度响应型聚合物-抗体缀合物与 DiagnosDisk（一种新型流通式）结合使用 测定装置。该方法的创新之处在于使用富集的唾液样本进行更一致的检测 结果，通过扩大唾液样本体积来增强抗原检测，并采用良好的 使用温度响应聚合物试剂开发了 DiagnosDisk 夹心免疫测定法， 简化设计，同时最大化灵敏度。我们将采用聚合物-抗体缀合物，有效 分离唾液中的分析物（目标 1）并制造 DiagnosDisk 设备以容纳更大容量的唾液 样本以提高检测限（目标 2）。新快速检测的灵敏度结合了 将使用来自 COVID-19 的唾液样本评估聚合物抗体缀合物和 DiagnosDisk 阳性和阴性患者（目标 3）。