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

过去十年来，疟疾控制力度的加大导致全球疟疾发病率大幅下降，世界卫生组织也认可了在全球范围内消除和根除疟疾的雄心勃勃的目标。从疟疾控制到消除疟疾的重点转变需要简单有效的方法来快速筛查疟疾。目前的疟疾筛查测试几乎完全依赖于显微镜检查和免疫学测试。由于检测能力有限，这两种方法都可能会错过重要信息。核酸检测（NAT）是目前最灵敏的方法。不幸的是，迄今为止开发的基于 NAT 的方法大多需要一定的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测试，将基于侧流的测试（可扩展的通量、最少的培训）和 NAT（高度敏感和特异性）的优点结合起来。拟议研究的成功将加强国内和全球的医疗保健系统。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基于实践经验的互动活动来促进学生的参与。本提案的研究目标是采用多学科方法探索在 USB 接口设备上进行高度灵敏且可扩展的疟疾核酸筛查测试。本研究目标将通过开展以下三项研究任务来实现。任务 1，测定优化和验证，目标是提高 DNA 提取率并提高环介导等温扩增测定拷贝数灵敏度。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的血液样本评估设备性能，目的是确定其灵敏度、特异性、再现性和检测限。拟议工作的创新之处在于三个方面：（1）高灵敏度疟疾等温扩增测定可以像基于侧流的测试一样简单地进行，（2）USB集线器的层树拓扑的级联能力提供可扩展的核酸测试吞吐量和异步处理能力，以及（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NSF的法定使命，并被认为值得支持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。

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.