Ultracompact sample-to-answer nucleic acid test on USB stick

在过去十年中，加强疟疾控制的努力使全球疟疾发病率大幅下降，世界卫生组织赞同在世界范围内消除和根除疟疾的宏伟目标。将重点从疟疾控制转向消除疟疾，需要简单有效的方法来快速筛查疟疾。目前的疟疾筛查试验几乎完全依靠显微镜和免疫学试验。由于检测能力有限，这两种方法都可能错过重要信息。核酸检测（NAT）是目前最灵敏的检测方法。不幸的是，迄今为止开发的基于nat的方法大多需要某些基础设施和熟练的技术人员。迫切需要一种改进的疟疾筛查检测，将基于横向流量的检测（可扩展的吞吐量、最少的培训）和NATs（高度敏感和特异性）的好处结合起来。拟议研究的成功将加强国内和全球的卫生保健系统。作为一种平台技术，它将允许向偏远、服务不足和资源有限的社区提供高度敏感的测试。此外，拟议的教育和外展活动将通过独特的以实践为基础的互动活动促进学生的参与。本课题的研究目标是采用多学科交叉的方法，在USB接口设备上探索一种高灵敏度、可扩展的疟疾核酸筛选试验。本研究目标将通过开展以下三个研究任务来实现。任务1，检测优化和验证，目的是提高DNA提取率，提高环介导的等温扩增检测拷贝数的敏感性。任务2，设备集成和仪器仪表，目标是开发一次性微流控芯片和超紧凑USB接口分析仪。任务3，使用从现场收集的实验室存档血液样本对设备进行性能评估，目的是确定其灵敏度、特异性、可重复性和检测限。这项工作的创新之处在于三个方面：(1)高度敏感的疟疾等温扩增分析可以像横向流检测一样简单地进行；(2)USB集线器层树拓扑的级联能力提供了可扩展的核酸检测吞吐量和异步处理能力；(3)多功能平台技术，可以广泛传播到需要便携性、连接性和易用性的情况。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Increased malaria control efforts have resulted in a dramatic reduction in the global malaria incidence over the past decade, and the World Health Organization has endorsed the ambitious goal of achieving worldwide malaria elimination and eradication. A change in focus from malaria control to malaria elimination requires easy and efficient ways to rapidly screen for malaria. Current malaria screening tests rely almost exclusively on microscopy and immunological tests. Both methods could miss significant information due to their limited detection abilities. Nucleic acid testing (NAT) is currently the most sensitive method available. Unfortunately, NAT-based methods developed to date mostly require certain infrastructures and skilled technicians. There is a critical need for an improved malaria screening test that combines the benefits of lateral flow-based testing (scalable throughput, minimal training) and the NATs (highly sensitive and specific). The success of the proposed research would enhance the health care system domestically and globally. As a platform technology, it would allow for delivering the highly sensitive test to remote, underserved and resource-limited communities. In addition, the proposed educational and outreach activity would foster student engagement through unique hands-on-experience-based interactive activities. The research objective of this proposal is to take a multidisciplinary approach to explore a highly sensitive and scalable malaria nucleic acid screening test on a USB interfaced device. This research objective will be achieved by carrying out the following three research tasks. Task 1, assay optimization and validation, with the goal to increase the DNA extraction yield and to increase the loop-mediated isothermal amplification assay copy number sensitivity. Task 2, device integration and instrumentation, with the goal to develop the disposable microfluidic chip and the ultracompact USB interfaced analyzer. Task 3, device performance evaluation with lab-archived blood samples collected from field sites, with the goal to determine its sensitivity, specificity, reproducibility and limits of detection. The innovation of the proposed work are three folds: (1) Highly sensitive malaria isothermal amplification assay could be performed as simple as lateral flow-based testing, (2) Scalable nucleic acid testing throughput and asynchronous processing capability provided by the cascading capability of a tier tree topology of USB hubs, and (3) Versatile platform technology that can be widely disseminated to situations that demand portability, connectivity and ease-of-use.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.