Demonstration of repeated Positionally Assisted Negative particle Rejection for High-Speed Sorting

用于高速分选的重复位置辅助负粒子剔除演示

• 批准号: 10081332
• 负责人: STEVEN W GRAVES
• 金额: $ 24.74万
• 依托单位: BENNUBIO INC
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10081332
• 关键词: 3D Print; Acoustics; Algorithms; Antibodies; Area; Biological Models; Biomedical Research; Caliber; Cell Separation; Cell Therapy; Cells; Cellular Spheroids; Centrifugation; Clinic; Collection; Computer software; Consumption; Detection; Development; Flow Cytometry; Fluorescence-Activated Cell Sorting; Information Systems; Liquid substance; Magnetism; Mechanics; Microspheres; Modeling; Modification; Optics; Pathway interactions; Population; Positioning Attribute; Process; Reaction Time; Sampling; Sorting - Cell Movement; Speed; Stream; System; Target Populations; Technology; Therapeutic; Time; base; data acquisition; design; instrument; interest; millisecond; novel strategies; particle; physical property; physical separation; tool

Project Summary Sorting and enrichment of cells and multicellular constructs is valuable to many areas of biomedical research and is of growing importance for cellular therapies. Enrichment processes use physical properties to perform high-speed separation of cells. However, due to the physical similarity of most cells, these approaches do not provide a high purity product. Therefore, sorting based on multiple parameters is needed to obtain pure cell populations. Fluorescence activated cell sorting (FACS) can separate cells based on >20 cellular markers and is very useful for sorting of rare cell populations. Unfortunately, FACS is a slow, low volume process. Thus, isolation of rare cell populations or large numbers of cells that require multiple markers to distinguish remains a challenging, time consuming, and expensive process that requires multiple steps that are each prone to cell loss. Therefore, we have designed an automated sorting process that combines the sensitive multicolor detection of FACS with the high speed and high volumetric rate of physical separation technologies. Our process, termed Positionally Assisted Negative particle Rejection (PANR), uses an acoustic standing wave to position the cells for flow cytometry analysis and downstream fluidics to sortthe cells. The sorted sample after rPANR is not diluted by sheath or other fluids so sample can be rapidly collected via any cell concentration technology. Because of the high volumetric rate that acoustic flow cytometry offers and the simplicity of implementing repeated cycles of PANR, rPANR offers a pathway to rapidly and gently sort cells. In addition, the rPANR system will be usable with particles from 1 to 1000 µm in diameter, which will make it useful for rare cells and large cellular constructs. This project will demonstrate the rPANR process to create a clear development pathway to a commercial rPANR instrument that sorts as few as 10 cells from a 109 cells/ml in less < 1hr. Furthermore, flow rates of >10 mL/min will enable high-speed enrichment of cells displaying multiple markers. Finally, it will be able to sort particles up to 1 mm in diameter at rates of over 10,000/s. These features make demonstration of the rPANR process the first step in the creation of a new tool that will be of great value to biomedical research, pharma discovery, and, in time, a very useful tool for cellular therapeutics.

项目摘要 细胞和多细胞构建体的分类和富集对于生物医学研究的许多领域都是有价值的 并且对细胞疗法的重要性越来越重要。富集过程使用物理特性执行 细胞的高速分离。但是，由于大多数细胞的身体相似性，这些方法没有 提供高纯度产品。因此，需要基于多个参数进行排序以获得纯单元 人群。荧光激活的细胞分选（FACS）可以根据> 20个细胞标记分离细胞，并 对于分类稀有细胞群体非常有用。不幸的是，FACS是一个缓慢，低量的过程。那， 隔离稀有细胞群或需要多个标记来区分的大量细胞仍然存在 具有挑战性，耗时和昂贵的过程，需要多个步骤，每个步骤都容易损失细胞。 因此，我们设计了一个自动分类过程，该过程结合了敏感的多色检测 具有高速和高体积速率的物理分离技术的FACS。我们的过程称为 位置辅助负粒子排斥（PANR），使用声学驻波将细胞定位 用于流式细胞仪分析和下游流体，以对细胞进行分类。 RPANR后的样品未稀释 通过鞘或其他液体，可以通过任何细胞浓度技术迅速收集样品。由于 声流式细胞术提供的高容量速率以及实施重复周期的简单性 PANR，RPANR提供了快速和轻轻分类细胞的途径。此外，RPANR系统将与 直径为1至1000 µm的颗粒，这将使其可用于稀有细胞和大细胞构建体。这 项目将展示RPANR过程，以创建通往商业RPANR的清晰开发途径 从少于1hr中的109个细胞/mL分类的仪器少于10个细胞。此外，流速> 10 mL/min 将实现显示多个标记的单元格的高速增强。最后，它将能够对粒子进行排序 直径为1毫米，速度超过10,000/s。这些功能使RPANR过程演示 创建一种新工具的第一步，该工具将对生物医学研究，制药发现以及，以及 随着时间的流逝，一种非常有用的细胞治疗工具。