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 细胞和多细胞结构的分选和浓缩对生物医学研究的许多领域都很有价值 在细胞疗法中越来越重要。浓缩过程使用物理属性来执行 细胞的高速分离。然而，由于大多数细胞的物理相似性，这些方法不 提供高纯度的产品。因此，需要基于多参数的排序才能获得纯单元格 人口。荧光激活细胞分选(FACS)可以基于&gt；20细胞标记和 对于稀有细胞群的分类非常有用。不幸的是，FACS是一个缓慢、低量的过程。因此， 分离稀有细胞群或需要多个标记才能区分的大量细胞仍然是一个 具有挑战性、耗时且昂贵的过程，需要多个步骤，每个步骤都容易丢失信元。 因此，我们设计了一种自动分拣流程，将敏感的多色检测结合在一起 FACS具有高速和高体积比的物理分离技术。我们的过程，称为 位置辅助负粒子排斥(PANR)，使用声波驻波来定位细胞 用于流式细胞仪分析和下游流体分选细胞。RPANR后的分选样品不稀释 通过鞘或其他流体，因此可以通过任何细胞浓缩技术快速采集样本。因为. 声学流式细胞术提供的高容积率以及实现重复循环的简单性 PANR，rPANR提供了一种快速温和地对细胞进行分选的途径。此外，rPANR系统将与 直径从1到1000微米的颗粒，这将使其适用于稀有细胞和大型细胞结构。这 项目将演示rPANR流程，以创建一条通向商业rPANR的清晰开发途径 从109个细胞/毫升中分选最少10个细胞的仪器，时间不超过1小时。此外，~gt；10毫升/分钟的流速 将实现显示多个标记的细胞的高速丰富。最后，它将能够对颗粒进行分类 到直径1毫米，速率超过10,000/S。这些特征使rPANR过程的演示成为 创建一种新工具的第一步，该工具将对生物医学研究、制药发现和 随着时间的推移，这是一种非常有用的细胞治疗工具。