NEXT GENERATION OPTICALLY ACTIVATED LARGE PARTICLE SORTING

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 几种研究途径迫切需要大颗粒的高速分选， 包括整个生物体的选择、多细胞颗粒和分子的选择 在大颗粒上合成的组合库。迄今为止，高速分拣主要 依赖于基于电荷的液滴分选，结合颗粒的多参数光学分析， 流式细胞仪，这是一个不可或缺的技术，为许多生物医学应用，包括 稀有细胞分离、染色体分选和细胞展示分子选择等。 然而，基于液滴的流分选机在大颗粒被分选时具有显著的局限性。 考虑了首先，增加颗粒尺寸需要更大的孔，以防止堵塞和影响。 液滴断裂点。随着分选孔直径的增加，分选孔会遭受越来越大的湍流， 这需要使用较低的线速度并限制分选速率（<1000/s）。这个， 有效地限制了颗粒尺寸（<100 μ m）并导致了替代的大颗粒分选方法， 包括机械流转向和微通道流体切换。我们的工作重点是 大颗粒分选的解决方案，这将导致分选机的发展， 直径可达1 mm，速度可与目前传统的基于液滴的分选机（>104/s）相媲美。 为了实现这一目标，我们将针对当前分选技术的每一个技术限制， 大颗粒系统的独特解决方案。首先，我们将开发高速同步粒子 输送系统，以克服颗粒输送的统计不确定性并最大限度地分选 通过提供已知的粒子位置来分类事件。其次，我们将利用我们最近的 低成本流式细胞术在激光和数据采集系统方面的发展， 低线速度并行流式细胞术分析仪，以最大化颗粒通量。三是 创建不受颗粒尺寸限制的液滴按需分选器。虽然这些技术 发展将最适用于大颗粒分选，他们也将有附带的好处 传统的排序，因为它们将大大降低系统成本，并创造有价值的并行 分析技术，用于高速分选。最后，我们将构造高速大粒子 用于内部测试的分选机和关键的外部合作，以高速分选大颗粒， 适体和肽的选择以及罕见肿瘤微球体的收集。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Several avenues of research have a critical need for high speed sorting of large particles, including the selection of whole organisms, multicellular particles and the selection of molecules from combinatorial libraries synthesized on large particles. High speed sorting has to date primarily relied on charge based droplet sorting coupled with multiparameter optical analysis of particles in a flow cytometer, which is an indispensable technique for numerous biomedical applications including rare cell isolation, chromosome sorting and cellular display molecular selection among many others. However, droplet based flow sorters have significant limitations when large particles are considered. First, increasing particle size requires largerorifices to prevent clogging and effects on droplet break off points. Sorting orifices suffer increasing turbulence as their diameter increases, which requires the use of lower linear velocities and restricts sorting rates (<1000/s). This, effectively limits particle size (<100 ¿m) and has led to alternative large particle sorting approaches, including mechanical stream diversion and micro-channel fluidic switching. Our effort focuseson solutions to large particle sorting that will result in the development of sorters that will sort particles up to 1 mm in diameter at rates comparable to current conventional droplet based sorters (>104/s). To accomplish this, we will target each technical limitation of current sorting technology with unique solutions for large particle systems. First, we will develop high speed synchronous particle delivery systems to overcome the statistical uncertainty of particle delivery and maximize sorting rates by providing known particle positions for sorting events. Second, we will leverage our recent low cost flow cytometry developments in lasers and data acquisition systems to create inexpensive low linear velocity parallel flow cytometry analyzers to maximize particle throughput. Third, we will create droplet on demand sorters that are not limited by particle size. While these technical developments will be most applicable to large particle sorting, they will also have ancillary benefits to conventional sorting, as they will dramatically reduce system cost and create valuable parallel analysis technology for high speed sorting. Finally, we will construct high-speedlarge particle sorters for internal testing and key external collaborations to sort large particles at high rates for selection of aptamers and peptides as well as rare tumor microspheroid collection.