Five-laser Aurora spectral flow cytometer

五激光 Aurora 光谱流式细胞仪

• 批准号: 10426837
• 负责人: Remi J Creusot
• 金额: $ 36.08万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10426837
• 关键词: Biomedical Research; Biopsy; Blood; Cells; Color; Cytometry; Detection; Flow Cytometry; Fluorochrome; Funding; Human; Incentives; Lasers; Life; Maintenance; Masks; Measures; Medical center; Organoids; Patients; Principal Investigator; Proteins; Reagent; Reporter; Research Personnel; Resolution; Sampling; Services; Side; Signal Transduction; Surface; Technology; Testing; Tissues; United States National Institutes of Health; Universities; Viola; Work; cell type; cost; design; detector; flexibility; humanized mouse; instrument; lymph nodes; operation; optical spectra; particle; single cell analysis; tool

PROJECT SUMMARY Flow cytometry is an essential tool for the single cell analysis of a wide variety of cell types to assess the expression of surface and intracellular markers and perform functional measures. Conventional flow cytometry has been unable to leverage the large variety of fluorochromes available for marker analysis, because the technology is focused on a narrow window of their emission spectrum, in which many of these fluorochromes overlap, limiting the number of markers that can be measured. In contrast, spectral flow cytometry acquires the full emission spectrum of each fluorochromes, which enables high resolution characterization of fluorochromes (or fluorescent proteins) that can be used in conjunction. Moreover, the autofluorescence emitted by unstained cells can also be acquired and subtracted for better detection of weaker fluorescent signals. The 5-laser Aurora from Cytek is one of the most powerful spectral flow cytometry instruments currently available and the need for this technology is supported (1) by the fact that our two most high-end conventional flow cytometers (LSRII) are reaching end of serviceable life in one year, and (2) by increasing demand for greater capabilities, including more parameters analyzed per sample, as well as better sensitivity for dim markers and for small particles. With the 5-laser Aurora, investigators can expand the number of markers assessed to well beyond 30 markers: Cytek has validated a 40-color panel for human blood and Dr. Farber at the Columbia University Irving Medical Center (CUIMC) is applying panels of 25-35 parameters for several of her projects with several substitutions made possible by the flexibility of panel design. This capability is attractive to many Flow Cytometry Core users who have limited amount of precious material to work with, including cells isolated from small tissue biopsies, humanized mice (particularly blood and lymph node samples), patient airway washes and tissue organoids. This technology is also ideal as an upgrade option for most of our LSRII users, as opposed to mass cytometry, which requires acquiring or generating all new reagents. When testing the Aurora, users have been able to overcome major roadblocks in their projects, such as the ability to identify tissues cells whose autofluorescence was masking the expression of a fluorescent reporter. Users also validated the detection of particles or cell products in the 100-200 nm size range, made possible by the Aurora's violet side scatter detector. In this proposal, 34 principal investigators at CUIMC have indicated how their projects (84% of which are NIH- funded) will be significantly enhanced by upgrading from conventional flow cytometry instrument to the 5-laser Aurora. Thus, the addition of this instrument will have a beneficial and long-lasting impact on biomedical research encompassing a broad variety of fields. Generous institutional support is provided in the form of incentives and coverage of maintenance costs, and the operation of the 5-laser Aurora will be sustainable in the long-term because its maintenance is comparable to conventional instruments in use.

项目摘要 流式细胞仪是对各种细胞类型进行单细胞分析的重要工具，以评估 表面和细胞内标记的表达并执行功能度量。常规流式细胞仪 一直无法利用可用于标记分析的各种荧光素 技术集中在其发射光谱的狭窄窗口上，其中许多荧光色素中的许多 重叠，限制可以测量的标记数。相比之下，光流式细胞仪获得了 每个荧光色素的全发射光谱 （或荧光蛋白）可以结合使用。此外，未染色发出的自动荧光 还可以获取和减去细胞，以更好地检测较弱的荧光信号。 Cytek的5-laser Aurora是目前可用的最强大的光流式细胞仪器之一 我们的两个最高端的常规流程支持了这项技术的需求（1） 细胞仪（LSRII）在一年内达到了可维修寿命的终结，（2）通过增加对更大的需求 功能，包括每个样本分析的更多参数，以及对昏暗标记和 用于小颗粒。使用5射线Aurora，调查人员可以将评估的标记数扩大到井 超过30个标记：Cytek已验证了一个40色的人类血液和Farber博士的哥伦比亚博士。 大学欧文医学中心（CUIMC）正在将25-35个参数的面板用于她的几个项目 面板设计的灵活性使几个替代成为可能。这种功能对许多流动都有吸引力 细胞仪核心用户使用的珍贵材料有限，包括与隔离的细胞 小组织活检，人源化小鼠（尤其是血液和淋巴结样本），患者气道洗涤和 组织器官。对于我们大多数LSRII用户而言，这项技术也是升级选项的理想选择 质量细胞仪，需要获取或生成所有新试剂。测试极光时，用户有 能够克服其项目中的主要障碍，例如鉴定组织细胞的能力 自荧光正在掩盖荧光报告基因的表达。用户还验证了检测 Aurora的紫罗兰侧散射检测器使100-200 nm尺寸范围内的颗粒或细胞产物成为可能。 在该提案中，CUIMC的34名主要研究人员指出了他们的项目（其中84％是NIH- 资助）将通过从常规流式细胞仪仪升级到5射线来显着增强 极光。因此，该仪器的添加将对生物医学研究产生有益且长期的影响 包括各种各样的领域。以激励措施的形式提供慷慨的机构支持 维护成本的覆盖范围以及5型激光Aurora的运行将在长期内可持续 因为其维护与使用的常规仪器相媲美。