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: 10627753
• 负责人: Zhenpeng Qin
• 金额: $ 50.08万
• 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/10627753
• 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; Darkness; Data; Detection; Development; Device or Instrument Development; Devices; Diagnosis; Diagnostic; Diagnostic Equipment; Diagnostic Procedure; Diagnostic Sensitivity; Diagnostic tests; Disease; Evaluation; Formulation; Future; Generations; 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; Training; Vaccines; Virion; Virus; Work; absorption; analog; antibody conjugate; clinical translation; computerized data processing; cost; design; detection limit; diagnostic assay; digital; genetic strain; improved; innovation; isothermal amplification; 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 呼吸道合胞病毒 (RSV) 是全球儿童快速死亡的最常见原因之一。 诊断对于抗生素的司法使用、减少医疗机构中的疾病传播以及 由于目前的诊断方法即将出现，因此可以立即进行治疗。 依赖耗时的实验室测试，包括病毒培养和聚合酶链反应 (PCR)，以及 快速诊断测试（例如侧流免疫分析，LFA）作为独立诊断不够灵敏。 因此，对 RSV 等离子体耦合的快速和超灵敏诊断测试的需求尚未得到满足。 测定是一种快速比色诊断测试，利用金纳米颗粒 (AuNP) 的光学响应 在目标识别过程中分析其浓度尽管操作简单，但灵敏度较高。 在这项拟议的工作中，等离子体耦合测定是有限的，我们的目标是大幅提高限制。 通过创新的数字纳米气泡检测进行等离激元耦合测定（LOD）。 直接建议使用识别 RSV 表面的抗体偶联 AuNP 来检测完整的 RSV 颗粒 AuNP 与多个 RSV F 蛋白结合并导致超短激光脉冲。 与单个 AuNP 相比，其吸收能力更强，因此可以选择性地激活耦合的 AuNP。 更大的光学吸收导致纳米级空化气泡，即纳米气泡，可以轻松测量 来自气泡诱导和瞬态散射的单个纳米气泡的产生导致了灵敏的数字。 我们的初步结果表明，“开”和“关”信号的检测性能提高了约 3 个数量级。 LOD 检测 RSV 在这项拟议的工作中，我们将首先通过优化 AuNP 来创新 AuNP 探针。 配方（尺寸、浓度和结合），研究非球形纳米颗粒以提高效率 病毒结合，并探索不对称抗体包被的 Janus 纳米颗粒进行控制结合。 我们将设计和构建一个原型设备，用于自动样品加载、读取和数据处理 最后，我们将使用临床 RSV 分离株（来自不同的遗传株）来测试该测定。 通过与现有的PCR和快速诊断测试进行比较，我们将建立临床样本。 数字纳米气泡测试的灵敏度和特异性 该测试快速（< 30 分钟）、低成本（AuNP 试剂）。 成本与侧流免疫分析相似），并且对 RSV 具有高度敏感性和特异性。此外，直接检测。 病毒颗粒的检测消除了核酸提取等大量样品制备的需要。 我们项目的成功将满足快速、灵敏的 RSV 诊断的迫切需求，并解决一个重大问题 受 RSV 影响的儿科患者的医疗保健需求。