Microfluidic Sample Preparation Device for Environmental DNA Separation and Washing **********

检测环境DNA（eDNA），从有机体分泌到环境中的浓度非常低，已成为一个强大的方法来识别水生系统中的危险或有风险的物种。精密生物监测（PBI）为现场检测稀有eDNA提供了突破性的技术。它包括用于过滤淡水样品的样品制备单元和用于感测eDNA的检测单元。该公司的问题与复杂的淡水性质有关，这使得样品过滤变得复杂，而样品中的悬浮物作为抑制剂，对系统的检测限和灵敏度产生负面影响。上述问题非常重要，因此必须研究一种样品制备解决方案，该解决方案必须1）对广泛的水样有效，2）可与公司的平台集成，3）便携式和电池供电，以及4）低成本。针对环境样品制备的特点，提出了一种微流控颗粒分离、DNA分离和DNA洗涤装置。多半径连续弯曲微通道将用于使用弹性力和惯性力从样品中顺序提取抑制性颗粒，所述弹性力和惯性力可以基于感兴趣的颗粒的尺寸施加到所述颗粒。两名研究助理（RA）和一名博士生将进行两阶段的研究。在第一阶段，我们将表征水样的颗粒和生物含量。在第二阶段，我们将使用第一阶段的信息和我们对流体-颗粒流动的了解来开发和测试微流体装置。从大到小的颗粒将依次从样品中分离，捕获在微珠上的DNA内容物将与其他分子内容物分离，并洗涤到干净的缓冲液中，用于PBI系统的检测。我们提出的设备是创新的，因为它将能够纯化已经以非常低的浓度分散在抑制性颗粒中的DNA分子，同时能够处理具有非牛顿特性的复杂流体，而不是常用的水基溶液。**

Detection of environmental DNA (eDNA), excreted from organisms into the environment at very low concentrations, has emerged as a powerful method for identification of dangerous or at-risk species in aquatic systems. Precision Biomonitoring (PBI) offers a ground-breaking technology for detection of rare eDNA on-field. It includes a sample preparation unit for filtration of freshwater samples and a detection unit for sensing eDNA. Company's problem is associated with the complex freshwater properties making sample filtration complicated, while the suspended matter in the samples act as inhibitors negatively affecting the detection limit and sensitivity of the system. The above problem is significant for which a sample preparation solution must be researched that is 1) effective on a broad range of water samples, 2) integrate-able with company's platform, 3) portable and battery-operated, and 4) low cost.**We propose developing a microfluidic particle separation, DNA isolation and DNA washing device to meet the attributes of environmental sample preparation. Multi-radius serial curved microchannels will be used to sequentially extract inhibitory particles from the sample using elastic and inertial forces that can be applied to particles of interest based on their sizes. Two Research Assistants (RAs) and one PhD student will conduct a two-phase research. In Phase 1, we will characterize the particle and biological content of water samples. In Phase 2, we will use the information in Phase 1 and our knowledge of fluid-particle flow to develop and test the microfluidic device. Particles from large to small will be sequentially separated from the sample and the DNA content, captured on microbeads, will be separated from the other molecular content and washed into clean buffers for detection with the PBI system. Our proposed device is innovative because it will be capable of purifying DNA molecules that are already dispersed at very low concentrations among the inhibitory particles, while being able to process complex fluids with non-Newtonian properties instead of the commonly-used water-based solutions. **