SBIR Phase II: Low-cost Detection and Enrichment of Nucleic Acids by Interfacing with Commercially Available Cell Sorters

这个小企业创新研究（SBIR）项目的更广泛的影响/商业潜力将是能够从大量的背景人群中检测和富集罕见的DNA或RNA分子。目前，这种检测是在昂贵的专用仪器上进行的，这些仪器灵敏度低，不能富集。该策略是将荧光识别的靶分子封装在薄油壳中，使其模拟细胞，然后在通常可用的灵敏和高通量细胞分选仪器上处理它们。通过将检测和浓缩工作转移到这样的机器上，研究人员所需要的只是一个简单的工具，将分子封装在薄的油壳中。通过高灵敏度和富集而成为可能的应用是众多的，从稀有病原体或细胞检测到以下游测序为目标的不常见突变序列的靶向捕获。在降低成本和扩大能力之间，这种方法可能会成为一种广泛采用的方法。SBIR第二阶段项目提出开发一种可部署的仪器，通过与现有流式细胞仪连接，可以高通量，低成本地定量样品中的DNA/RNA分子。最初的目标是将最终工具中所需的所有功能快速设计成一个粗略的alpha版本。其功能将包括从迭代优化的硬塑料微流控芯片中产生可靠的液滴，以及通过具有精确反馈控制的先进流体处理系统实现自动化。第二个目标是构建实际的beta版本供客户试用。测试版将用精简的定制工程取代大部分现成的部件，并专注于减小尺寸和成本，同时通过更多迭代芯片优化来改进功能。最终目标是使用beta仪器，使用客户样品和基准测试优化多个应用的方案和试剂。在项目结束时，目标是在客户现场使用仪器生成数据。

The broader impact/commercial potential of this Small Business Innovation Research (SBIR) project will be to enable affordable detection and enrichment of rare DNA or RNA molecules from a large background population. Currently, such detection is performed on expensive, dedicated instruments that have low sensitivity and are not able to enrich. The strategy is to encapsulate the fluorescently-identified target molecules in thin oil shells so that they mimic cells, and then process them on sensitive and high-throughput cell-sorting instruments that are commonly available. By offloading the detection and enrichment to such machines, all that is required by researchers is a simple tool that encapsulates the molecules in thin oil shells. Applications made possible by high sensitivity and enrichment are numerous, from rare pathogen or cell detection to target capture of uncommon mutant sequences with the goal of downstream sequencing. Between the reduced cost and expansion of capabilities, this approach is likely to become a widely adopted method.This SBIR Phase II project proposes to develop a deployable instrument that allows high-throughput, low-cost quantitation of DNA/RNA molecules in a sample by interfacing with existing flow cytometry instruments. The initial objective is to quickly engineer all the functionality desired in that final instrument into a rough alpha version. Its features will include reliable droplet generation from iteratively optimized hard-plastic microfluidic chips, and automation via an advanced fluid handling system with precise feedback control. The second objective is to build the actual beta version for customer trials. The beta version will replace most of the off-the-shelf parts with streamlined, custom engineering and focus on reducing size and cost while still improving function through more iterative chip optimization. The final objective is to use the beta instrument to optimize the protocols and reagents for several applications using customer samples and benchmarking. At the conclusion of the project, the goal is to have instruments at customer sites generating data.