Self-Powered Sample Concentrating and CRISPR-based Biosensing for Moile HIV-1 RNA Detection

用于 Moile HIV-1 RNA 检测的自供电样本浓缩和基于 CRISPR 的生物传感

• 批准号: 10228759
• 负责人: Changchun Liu
• 金额: $ 48.13万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10228759
• 关键词: AIDS/HIV problem; Acute; Adopted; Affect; Bioinformatics; Biological Assay; Biosensing Techniques; Blinded; Blood; Blood capillaries; Blood specimen; Cellular Phone; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Country; Cytolysis; Data; Data Reporting; Detection; Developing Countries; Development; Devices; Diagnostic; Early Diagnosis; Early Intervention; Engineering; Ensure; Environment; Epidemic; Equipment; Evaluation; Fingers; Fluorescence; HIV; HIV Infections; HIV diagnosis; HIV-1; Health; Home; Hospitals; Human Resources; Human immunodeficiency virus test; Immunoassay; Individual; Infant; Infectious Diseases Research; International; Laboratories; Membrane; Methods; Microfluidic Microchips; Monitor; Mothers; Nucleic Acid Amplification Tests; Nucleic Acids; Patients; Pennsylvania; Performance; Phase; Plasma; Play; Polymerase; Public Health; RNA; Reaction; Recovery; Research; Resources; Reverse Transcriptase Polymerase Chain Reaction; Reverse Transcription; Risk; Role; Sampling; Sensitivity and Specificity; Signal Transduction; Statistical Data Interpretation; System; Technology; Test Result; Testing; Time; Training; Treatment Effectiveness; Treatment Failure; Universities; Validation; Viral; Viral Load result; Viremia; Whole Blood; Zambia; antiretroviral therapy; base; biomaterial compatibility; clinical application; cost; design; detection platform; detection sensitivity; diagnostic technologies; field study; improved; innovation; instrument; instrumentation; internal control; isothermal amplification; molecular diagnostics; multidisciplinary; next generation; point of care; point-of-care detection; point-of-care diagnostics; portability; recombinase; self testing; seroconversion; tool; transmission process; viral RNA; viral detection; viral rebound

Self-Powered Sample Concentrating and CRISPR-based Biosensing for Mobile HIV-1 RNA Detection Abstract HIV/AIDS has become a major public health concern affecting ~37.9 million people worldwide. Early diagnosis of acute HIV infection during seroconversion window will facilitate early intervention. During antiretroviral treatment (ART) of HIV-infected patients, it requires frequent monitoring of HIV viral load to confirm treatment effectiveness, and to identify viral rebound. HIV viral load testing that quantifies HIV viral RNA (circulating HIV virus) in plasma is the most accurate and reliable approach for the ART monitoring and acute HIV detection. However, current standard HIV viral load testing methods rely on expensive equipment and well-trained personnel, limiting their clinical applications in centralized laboratories and hospital environments. Commercially available immunoassay-based point of care (POC) diagnostic technologies, such as OraQuick® HIV Self-Test (HIVST), are not effective to detect acute HIV infections, as well as ART failure. As a consequence, the lack of a simple, rapid, affordable, POC diagnostic tool for HIV RNA detection leaves many individuals unaware of their condition and impedes timely antiretroviral treatment. To fill this gap, we propose to develop a low-cost (~ $ 5), rapid (< 35 min), and sensitive (<1,000 copies/mL), clustered regularly interspaced short palindromic repeats (CRISPR) biosensing platform for HIV viral load testing using finger- prick volume (~50 µL) of whole blood. In the R61 phase (Aims 1-3), we will: i) develop and optimize highly sensitive and specific CRISPR biosensing technology for next-generation nucleic acid-based molecular diagnostics, and ii) design and fabricate a disposable "blood-to-answer", CRISPR biosensing device that integrates self-powered plasma separation, viral RNA enrichment, and CRISPR biosensing detection. In the Phase 33 (Aims 4-5), we will systematically evaluate the performance of our integrated CRISPR biosensing platform, and rigorously validate its feasibility for clinical application by testing HIV clinical samples in the US and Zambia. If successful, such a simple, rapid, affordable, POC detection platform will enable acute HIV diagnosis and viral load testing at home and be appropriate for resource-limited settings where HIV is most prevalent.

适用于移动设备的自供电样本浓缩和基于 CRISPR 的生物传感 HIV-1 RNA检测 抽象的 HIV/艾滋病已成为影响全球约 3790 万人的主要公共卫生问题。早期诊断 血清转换窗口期间急性艾滋病毒感染的发生将有助于早期干预。抗逆转录病毒治疗期间 HIV感染者的治疗（ART），需要经常监测HIV病毒载量以确认治疗 有效性，并确定病毒反弹。 HIV 病毒载量检测可量化 HIV 病毒 RNA（循环 HIV 血浆中的病毒）是 ART 监测和急性 HIV 检测最准确、最可靠的方法。 然而，当前标准的 HIV 病毒载量检测方法依赖于昂贵的设备和训练有素的人员 人员，限制了其在集中实验室和医院环境中的临床应用。 市售的基于免疫测定的护理点 (POC) 诊断技术，例如 OraQuick® HIV 自检 (HIVST) 不能有效检测急性 HIV 感染以及 ART 失败。作为一个 因此，缺乏简单、快速、经济实惠的用于 HIV RNA 检测的 POC 诊断工具使许多人 个人不了解自己的病情并妨碍及时的抗逆转录病毒治疗。为了填补这一空白，我们建议 开发一种低成本（约 5 美元）、快速（< 35 分钟）、灵敏（<1,000 拷贝/mL）、定期聚类的方法 间隔短回文重复序列 (CRISPR) 生物传感平台，用于使用手指进行 HIV 病毒载量检测 全血的点刺体积 (~50 µL)。在R61阶段（目标1-3），我们将：i）高度开发和优化 用于下一代核酸分子的灵敏且特异的 CRISPR 生物传感技术 诊断，ii) 设计和制造一次性“血液到答案”CRISPR 生物传感设备 集成了自供电血浆分离、病毒 RNA 富集和 CRISPR 生物传感检测。在 第 33 阶段（目标 4-5），我们将系统评估集成 CRISPR 生物传感的性能 平台，并通过在美国测试HIV临床样本严格验证其临床应用的可行性 和赞比亚。如果成功，这样一个简单、快速、经济实惠的 POC 检测平台将能够实现急性 HIV 检测 在家进行诊断和病毒载量检测，适用于艾滋病毒最严重的资源有限的环境 流行。