Quantitative microfluidic NAT-on-USB: towards routine HIV viral load testing

定量微流控 NAT-on-USB：走向常规 HIV 病毒载量检测

• 批准号: 1912410
• 负责人: Weihua Guan
• 金额: $ 35.25万
• 依托单位: Pennsylvania State Univ University Park
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1912410&HistoricalAwards=false
• 关键词: Quantitative; microfluidic; NAT; USB; towards

HIV, the virus that causes AIDS, continues to be a significant public health issue. Fortunately, HIV can be effectively managed if people of HIV can be identified and their virus is controlled to undetectable levels by antiretroviral therapy. Increasing access to routine viral load testing can help reduce the treatment failure rate by early detection of viral rebound caused by either drug resistance or poor therapy adherence. To this end, HIV self-testing, a process in which individual who wants to know HIV status collects a specimen, performs a test and interprets the result in private, has become an empowering and innovative approach. Existing HIV self-testing methods rely almost exclusively on lateral flow based test to detect host antibody response to HIV infection. They could miss a significant portion of asymptomatic individuals during the 6-12 weeks of early infection window; they also lack the ability for detection of viral rebound. Nucleic acid testing is currently the only method for viral load quantification. Nevertheless, its use by laypersons is limited due to the sample handling and assay complexity. The proposed research has the potential to significantly enhance the treatment outcomes for individuals of HIV under antiretroviral therapy. In addition, with educational and outreach activities, the proposed research is well positioned to enhance the integrative learning experience and to engage multidisciplinary students at many levels.The primary research objective of this proposal is to explore an ultra-compact quantitative nucleic acid testing (NAT) on a disposable microfluidic chip with a USB analyzer to detect HIV-1 viral rebound that is simple enough for laypersons to test themselves to monitor treatment adherence. The goal is to develop and validate a whole blood-based test that can semi-quantitatively assess the presence of HIV-1 RNA at cell concentrations as low as 1000 copies/ml. The test can be performed similarly to a home blood glucose test with a single step of finger-prick blood loading. To this end, four research tasks regarding the microfluidic sample preparation, amplification assay, USB analyzer integration, and analytical validation will be pursued. Task 1. Automated microfluidic sample preparation. A streamlined microfluidic chip for automated plasma separation and RNA extraction from whole blood will be developed. Task 2. An HIV-1 Real-time reverse transcription loop-mediated isothermal amplification assay will be optimized and the lowest possible limit of detection will be explored. Task 3. Analyzer integration and small-scale prototyping. The analyzer hardware and software will be developed and integrated for easy and robust operation. Task 4. Laboratory validation with control samples. The prototyped device will be validated in lab settings using HIV-1 plasma samples spiked into whole blood.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

艾滋病毒（HIV）是导致艾滋病的病毒，仍然是一个重大的公共卫生问题。幸运的是，如果能够识别艾滋病毒感染者，并通过抗逆转录病毒治疗将其病毒控制在不可检测的水平，就可以有效地控制艾滋病毒。增加常规病毒载量检测的机会可以通过及早发现由耐药性或治疗依从性差引起的病毒反弹来帮助降低治疗失败率。为此，艾滋病毒自我检测，即想要了解艾滋病毒状况的个人私下收集样本、进行检测并解释结果的过程，已成为一种赋权和创新的方法。现有的 HIV 自检方法几乎完全依赖于基于侧向层析的测试来检测宿主抗体对 HIV 感染的反应。在早期感染窗口期的 6-12 周内，他们可能会漏掉很大一部分无症状个体；他们还缺乏检测病毒反弹的能力。核酸检测是目前病毒载量定量的唯一方法。然而，由于样品处理和测定的复杂性，外行人士的使用受到限制。拟议的研究有可能显着提高接受抗逆转录病毒治疗的艾滋病毒患者的治疗效果。此外，通过教育和推广活动，拟议的研究能够很好地增强综合学习体验并吸引多个层次的多学科学生。该提案的主要研究目标是探索一种超紧凑的定量核酸检测（NAT） ）在带有 USB 分析仪的一次性微流控芯片上检测 HIV-1 病毒反弹，这对于非专业人士来说非常简单，可以自我测试以监测治疗依从性。目标是开发和验证一种基于全血的测试，可以半定量评估细胞浓度低至 1000 拷贝/毫升时 HIV-1 RNA 的存在。该测试的执行方式与家庭血糖测试类似，只需一步手指刺血加载即可。为此，将开展关于微流体样品制备、扩增测定、USB分析仪集成和分析验证的四项研究任务。任务 1. 自动化微流体样品制备。将开发一种用于自动血浆分离和全血 RNA 提取的简化微流控芯片。任务 2. 将优化 HIV-1 实时逆转录环介导的等温扩增测定，并探索尽可能低的检测限。任务 3. 分析器集成和小规模原型设计。分析仪的硬件和软件将被开发和集成，以实现简单、稳健的操作。任务 4. 使用对照样品进行实验室验证。该原型设备将在实验室环境中使用掺入全血的 HIV-1 血浆样本进行验证。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力优点和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Sensitive and specific CRISPR-Cas12a assisted nanopore with RPA for Monkeypox detection

灵敏且特异的 CRISPR-Cas12a 辅助纳米孔与 RPA 用于猴痘检测

• DOI: 10.1016/j.bios.2023.115866
• 发表时间: 2024-02
• 期刊: Biosensors and Bioelectronics
• 影响因子: 12.6
• 作者: Ahamed, Md. Ahasan;Khalid, Muhammad Asad;Dong, Ming;Politza, Anthony J.;Zhang, Zhikun;Kshirsagar, Aneesh;Liu, Tianyi;Guan, Weihua
• 通讯作者: Guan, Weihua

Handheld Purification-Free Nucleic Acid Testing Device for Point-of-Need Detection of Malaria from Whole Blood

手持式免纯化核酸检测设备，用于从全血中即时检测疟疾

• DOI: 10.1021/acssensors.2c02169
• 发表时间: 2023-02
• 期刊: ACS Sensors
• 影响因子: 8.9
• 作者: Kshirsagar, Aneesh;Choi, Gihoon;Santosh, Vishaka;Harvey, Tara;Bernhards, Robert Cory;Guan, Weihua
• 通讯作者: Guan, Weihua

Programmable magnetic robot (ProMagBot) for automated nucleic acid extraction at the point of need

可编程磁性机器人 (ProMagBot)，用于在需要时自动提取核酸

• DOI: 10.1039/d3lc00545c
• 发表时间: 2023-08
• 期刊: Lab on a Chip
• 影响因子: 6.1
• 作者: Politza, Anthony J.;Liu, Tianyi;Guan, Weihua
• 通讯作者: Guan, Weihua

Quantitative Analysis of Factors Affecting the Event Rate in Glass Nanopore Sensors

玻璃纳米孔传感器事件发生率影响因素的定量分析

• DOI: 10.1021/acssensors.9b01540
• 发表时间: 2019-10
• 期刊: ACS Sensors
• 影响因子: 8.9
• 作者: Nouri, Reza;Tang, Zifan;Guan, Weihua
• 通讯作者: Guan, Weihua

Programmable Magnetic Robot (ProMagBot) For Fully Automated Nucleic Acid Sample Preparation at The Point of Need

可编程磁性机器人 (ProMagBot) 用于在需要时全自动制备核酸样品

• 发表时间: 2022-05
• 期刊: Hilton Head Workshop
• 作者: A. J. Politza; T. Liu
• 通讯作者: T. Liu