Rapid Diagnostic Test for Respiratory Syncytial Virus by Digital Nanobubbles

数字纳米气泡对呼吸道合胞病毒的快速诊断测试

• 批准号: 10155417
• 负责人: Zhenpeng Qin
• 金额: $ 50.06万
• 依托单位: UNIVERSITY OF TEXAS DALLAS
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10155417
• 关键词: Address; Affect; Antibiotics; Antibodies; Antiviral Therapy; Area; Bar Codes; Binding; Biological Assay; Biosensing Techniques; Biosensor; Bypass; Cell Culture Techniques; Cessation of life; Child; Childhood; Chimeric Proteins; Clinical; Clinical Sensitivity; Color; Communicable Diseases; Consumption; Coupled; Coupling; Data; Detection; Development; Device or Instrument Development; Devices; Diagnosis; Diagnostic; Diagnostic Procedure; Diagnostic Sensitivity; Diagnostic tests; Disease; Evaluation; Formulation; Future; Generations; Gold; Health care facility; Healthcare; Immunoassay; Infant; Interdisciplinary Study; Laboratories; Lasers; Lateral; Lead; Measures; Membrane Proteins; Methods; Microspheres; Nanosphere; Nucleic Acids; Optics; Outcome; Performance; Physiologic pulse; Pneumonia; Polymerase Chain Reaction; Preparation; Process; Proteins; Quantum Dots; Rapid diagnostics; Reading; Reagent; Respiratory syncytial virus; Rod; Sampling; Secondary to; Sensitivity and Specificity; Signal Transduction; Specificity; Specimen; Speed; Surface; System; Testing; Time; Vaccine Therapy; Virion; Virus; Work; absorption; analog; antibody conjugate; base; clinical translation; computerized data processing; cost; design; detection limit; diagnostic assay; digital; genetic strain; improved; innovation; isothermal amplification; laboratory experience; lateral flow assay; nanoGold; nanobubble; nanoparticle; nanorod; nanoscale; novel; operation; pediatric patients; performance tests; plasmonics; point-of-care diagnostics; prototype; research clinical testing; respiratory pathogen; response; sensor; solid state; success; superinfection; viral detection; virus culture

ABSTRACT Respiratory syncytial virus (RSV) is one of the most common causes to pediatric death globally. Rapid RSV diagnostics is important for judicial use of antibiotics, to reduce disease spread in healthcare facilities, and to enable prompt treatment since several RSV antiviral therapies are on the horizon. Current diagnostic methods rely on time-consuming laboratory-based tests including virus culture and polymerase chain reaction (PCR), and rapid diagnostic tests (e.g., lateral flow immunoassay, LFA) are not sufficiently sensitive as standalone diagnosis. Therefore, there is an unmet need for rapid and ultrasensitive diagnostic tests for RSV. The plasmonic coupling assay is a rapid colorimetric diagnostic test that makes use of the optical response of gold nanoparticles (AuNPs) during the process of target recognition to analyze its concentration. Despite its easy operation, the sensitivity of the plasmonic coupling assay is limited. In this proposed work, we aim to substantially improve the limit of detection (LOD) of the plasmonic coupling assay by innovative digital nanobubble detection. Specifically, we propose to directly detect intact RSV particles with antibody-conjugated AuNPs that recognize the RSV surface fusion (F) protein. AuNPs bind to multiple RSV F proteins and lead to plasmonic coupling. Ultrashort laser pulse selectively activates coupled AuNPs due to their enhanced absorption compared with a single AuNP. This greater optical absorption leads to nanoscale cavitation bubbles, i.e. nanobubbles, which can be measured easily from the bubble-induced and transient scattering. Single nanobubble generation leads to a sensitive digital detection with “on” and “off” signals. Our preliminary results suggest ~3 orders of magnitude improvement of LOD in detecting RSV. In this proposed work, we will firstly innovate the AuNP probe by optimizing the AuNP formulation (size, concentration and conjugation), investigating the non-spherical nanoparticle for more efficient virus binding, and exploring the asymmetric antibody-coated Janus nanoparticles for controlled binding. Next, we will design and build a prototype device for automated sample loading, reading, and data processing for the digital nanobubble assay. Lastly, we will test this assay with clinical RSV isolates (from diverse genetic strains) and clinical specimens. By comparing with current PCR and rapid diagnostic tests, we will establish the clinical sensitivity and specificity for the digital nanobubble test. This test is fast (< 30 minutes), low-cost (AuNP reagent cost is similar to lateral flow immunoassay), and highly sensitive and specific for RSV. Furthermore, the direct detection of virus particles eliminates the need of extensive sample preparation such as nucleic acid extraction. Success of our project will meet urgent demands of rapid and sensitive RSV diagnostics and address a major healthcare need for pediatric patients that are affected by RSV.

抽象的 呼吸综合病毒（RSV）是全球小儿死亡的最常见原因之一。快速RSV 诊断对于司法使用抗生素很重要，以减少医疗机构中的疾病传播以及对 启用迅速治疗，因为几种RSV抗病毒疗法即将到来。当前的诊断方法 依靠耗时的基于实验室的测试，包括病毒培养和聚合酶链反应（PCR）和 快速诊断测试（例如，侧流免疫测定，LFA）不足以像独立诊断那样敏感。 因此，对RSV的快速和超敏感诊断测试的需求未满足。等离子耦合 测定是一种快速的比色诊断测试，可利用金纳米颗粒的光学响应（AUNPS） 在目标识别过程中以分析其浓度。尽管操作很容易，但 等离子耦合测定有限。在这项拟议的工作中，我们旨在大大提高 通过创新的数字纳米泡检测检测等离子耦合测定法（LOD）。具体来说，我们 提议直接用抗体偶联的AUNP直接检测完整的RSV颗粒，以识别RSV表面 融合（F）蛋白。 AUNP与多种RSV F蛋白结合，并导致血浆耦合。超短激光脉冲 与单个AUNP相比，由于其增强的抽象，选择性激活了AUNP的耦合。这 更大的光学滥用会导致纳米级气泡气泡，即纳米泡，可以轻松测量 来自气泡引起的瞬态散射。单纳米泡产生导致敏感的数字 用“ ON”和“ OFF”信号检测。我们的初步结果表明〜3个数量级的改善。 检测RSV的LOD。在这项拟议的工作中，我们将首先通过优化AUNP来创新AUNP探测器 形成（尺寸，浓度和制剂），研究非球纳米颗粒以提高效率 病毒结合，并探索用于控制结合的不对称抗体涂层纳米颗粒。下一个， 我们将设计和构建用于自动示例加载，阅读和数据处理的原型设备 数字纳米泡测定。最后，我们将使用临床RSV分离株（来自潜水员遗传菌株）测试该测定法 和临床标本。通过与当前的PCR和快速诊断测试进行比较，我们将建立临床 数字纳米泡检验的灵敏度和特异性。该测试很快（<30分钟），低成本（AUNP试剂 成本类似于横向流量免疫测定），并且对RSV高度敏感且具有特异性。此外，直接 病毒颗粒的检测消除了需要大量样品制备的必要性，例如提取核酸。 我们项目的成功将满足快速和敏感的RSV诊断的紧急需求，并解决主要的问题 对受RSV影响的儿科患者的医疗保健需求。