High volume high throughput affordable parallel acoustic flow cytometry

高容量、高通量、经济实惠的并行声学流式细胞仪

• 批准号: 8575382
• 负责人: STEVEN W GRAVES
• 金额: $ 17.68万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8575382
• 关键词: Acoustics; Address; Animal Model; Area; Bacteria; Binding; Biological Assay; Biological Models; Biomedical Research; Blood; Blood Cells; Blood specimen; Caliber; Calibration; Cells; Cellular Spheroids; Clinical; Clinical Research; Detection; Development; Diagnostic; Effectiveness; Ensure; Flow Cytometry; Generations; Goals; Image; Lasers; Light; Liquid substance; Microspheres; Modeling; Optics; Particle Size; Pathogen detection; Population; Population Analysis; Preparation; Process; Property; Resolution; Sampling; Solutions; Source; Speed; Stream; System; Techniques; Technology; Translating; Ursidae Family; Whole Blood; clinical application; cost; cost effective; data acquisition; design; detector; fluid flow; high throughput analysis; high throughput screening; instrument; meter; novel strategies; particle; pressure; prototype; public health relevance; small molecule libraries; success; tumor; Acoustics; Address; Animal Model; Area; Bacteria; Binding; Biological Assay; Biological Models; Biomedical Research; Blood; Blood Cells; Blood specimen; Caliber; Calibration; Cells; Cellular Spheroids; Clinical; Clinical Research; Detection; Development; Diagnostic; Effectiveness; Ensure; Flow Cytometry; Generations; Goals; Image; Lasers; Light; Liquid substance; Microspheres; Modeling; Optics; Particle Size; Pathogen detection; Population; Population Analysis; Preparation; Process; Property; Resolution; Sampling; Solutions; Source; Speed; Stream; System; Techniques; Technology; Translating; Ursidae Family; Whole Blood; clinical application; cost; cost effective; data acquisition; design; detector; fluid flow; high throughput analysis; high throughput screening; instrument; meter; novel strategies; particle; pressure; prototype; public health relevance; small molecule libraries; success; tumor

DESCRIPTION (provided by applicant): The analytical power of flow cytometry makes it invaluable for numerous biomedical applications that require the enumeration of cell populations and the analysis of multicellular model systems or organisms. However, sample analysis flow rates of typical flow cytometers are limited to less than 250 uL/min, analytical rates are limited o 70,000 cells/s, and particle diameters must be less than 70 um. These limitations are driven by a number of factors that include pressure induced by high linear velocity fluid flows, turbulence in wide channels, and the single point analysis of stochastically arriving particles. Therefore, flow cytometry requires significant additional sample preparation steps to be effective in the analysis of very rare cell populations, uses offline particle concentration to analyze particles in large volume samples, and requires special purpose large flow channel cytometers using low linear velocity hydrodynamic focusing in wide channels to analyze particles that are >70 um in diameter at low analysis rates (200 s-1). Such limitations severely reduce its effectiveness in many critical applications including the detection of rare blood cell populations, the detection of pathogens in liquid samples, and the high throughput analysis model systems (e.g. multicellular model organisms, cellular spheroids, and one-bead-one-compound chemical libraries) that use large particles. To provide the analytical power of flow cytometry to these critical applications, we must dramatically increase the analytical rate, volumetric sample delivery, and the useable particle size of flow cytometers. To this end, we have developed acoustic flow cells that generate up to 300 focused parallel streams of particles using both acoustically resonant micro fabricated channels and multi-node acoustic standing waves. These flow cells focus particles up to 200 um in diameter at volumetric delivery rates as high as 25 mL/min. In this proposal, we will optimize the fluidics and optical properties of our flow cells and couple them with new approaches for highly parallel optical detection to create an affordable parallel acoustic flow cytometer (APAfc) platform. To address the broad set of unmet application needs, the APAfc platform will analyze cells or particles, ranging from 1 to 1000 um in diameter, at flow rates up t 50 mL/min, and at rates up to 1 x 106 particles/s. Importantly, the APAfc will achieve these specifications while retaining the analytical properties of flow cytometry (sensitivity, resolutionof free vs. bound probes, correlated multipara meter analysis) that make it the technology of choice for cell and particle analysis. Furthermore, the APAfc platform will be designed using affordable technologies to ensure that when translated into a commercial product, it will cost about what a low- end flow cytometer does today (~$50 to $100K). We will demonstrate the effectiveness of the APAfc platform using relevant models of clinical and research assays that are directly limited by analytical rates, volumetric throughput, or particle size. Development of the APAfc will have significant impact on both biomedical research and clinical diagnostics. It will provide a prototype instrument that provides highly sensitive and precise multipara meter optical analysis at analytical and volumetric delivery rates sufficient to provide a cost effective solution to routine detection of rare cells in blood or environmental samples, dramatically increase sample processing rates for HTS applications, and dramatically speed the analysis of multicellular particles and model organisms. We anticipate that if we are successful, our approaches to large volume high throughput flow cytometry will bring powerful analytical techniques to bear on a new spectrum of clinical and research problems.

描述（由申请人提供）：流式细胞仪的分析能力使其对于需要枚举细胞种群的众多生物医学应用以及多细胞模型系统或生物体的分析而言是无价的。但是，典型流式细胞仪的样本分析流速限制为小于250 ul/min，分析速率限制为O 70,000单元/s，颗粒直径必须小于70 um。这些局限性是由许多因素驱动的，这些因素包括由高线性速度流动引起的压力，宽通道中的湍流以及随机到达颗粒的单点分析。因此，流式细胞仪需要大量的额外样品制备步骤，以在分析非常罕见的细胞种群的分析中，使用离线颗粒浓度来分析颗粒中的颗粒 大容量样品，需要特殊目的的大型流通通道细胞仪，使用低线性流体动力学聚焦在宽通道中，以低分析速率（200 S-1）分析直径> 70 um的颗粒。这种局限性严重降低了其在许多关键应用中的有效性，包括检测稀有血细胞群体，检测 液体样品中的病原体以及使用大颗粒的高吞吐量分析模型系统（例如多细胞模型生物，细胞球体和单珠一个化学库）。为了为这些关键应用提供流式细胞仪的分析能力，我们必须大大提高分析速率，体积样品输送和流式细胞仪的可用粒径。为此，我们开发了声流细胞，使用声学谐振的微型制造通道和多节点的声波产生了多达300个聚焦的颗粒流粒子流。这些流动细胞以高达25 ml/min的体积递送速率聚焦直径高达200 um。在此提案中，我们将优化流动池的流体和光学特性，并将它们与高度平行的光学检测方法相结合，以创建一个负担得起的平行声流式细胞仪（APAFC）平台。为了满足广泛的未满足应用需求，APAFC平台将分析直径为1到1000 UM的细胞或颗粒，以t 50 ml/min的流速和高达1 x 106个颗粒/s的速率。重要的是，APAFC将达到这些规格，同时保留流式细胞仪的分析特性（灵敏度，自由分辨率与结合探针，相关的多拉仪分析），这使其成为细胞和粒子分析的首选技术。此外，APAFC平台将使用负担得起的技术设计，以确保将其翻译成商业产品时，低端流式细胞仪今天的作用将花费成本（约50至$ 100k）。我们将使用相关的临床和研究测定模型来证明APAFC平台的有效性，这些模型直接受到分析速率，体积吞吐量或粒径的限制。 APAFC的开发将对生物医学研究和临床诊断都有重大影响。它将提供一种原型仪器，可在分析和体积交付速率上提供高度敏感和精确的多抗光学分析，足以提供成本效益 解决血液或环境样品中稀有细胞的常规检测，大大提高了HTS应用的样品处理速率，并大大加快了对多细胞颗粒和模型生物的分析。我们预计，如果我们成功，我们的大量吞吐量流式细胞仪的方法将带来强大的分析技术，以在新的临床和研究问题上承受。