Eliminating Interference from Autofluorescence in Flow Cytometry

消除流式细胞术中自发荧光的干扰

• 批准号: 10484468
• 负责人: Giacomo Vacca
• 金额: $ 144.18万
• 依托单位: KINETIC RIVER CORPORATION
• 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 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10484468
• 关键词: Acquired Immunodeficiency Syndrome; Acute Myelocytic Leukemia; Algorithmic Analysis; Algorithmic Software; Algorithms; Antigens; Benchmarking; Biological; Biological Assay; Biomedical Research; Cancer Biology; Cell Count; Cell Membrane Permeability; Cells; Clinical; Collection; Color; Colorado; Communicable Diseases; Data; Data Analyses; Detection; Diagnosis; Diagnostic; Discipline; Dyes; Event; Excision; Fiber; Flow Cytometry; Fluorescence; Freeze Drying; Funding; Grant; Human; Immunologist; Immunology; Immunophenotyping; Kinetics; Label; Laboratories; Lasers; Lead; Lymphocyte; Malignant Neoplasms; Measures; Methods; Microbiology; Microspheres; Molecular Biology; Monitor; Myeloid Cells; NADH; Oncology; Output; Oxidation-Reduction; Patients; Phase; Ploidies; Research; Research Personnel; Rivers; Sampling; Shapes; Signal Transduction; Small Business Innovation Research Grant; Source; Speed; Stains; Surface; System; Technology; Testing; Time; Tumor Biology; United States National Institutes of Health; Viola; analog; cancer immunotherapy; clinically relevant; commercialization; computerized data processing; data acquisition; data handling; data quality; design; drug development; experimental study; fluorophore; improved; instrument; novel; optical spectra; programs; prototype; receptor; response; signal processing; stem cell therapy; time resolved data; time use; tool

PROJECT SUMMARY/ABSTRACT Flow cytometry (FC) is a powerful cell analysis tool relying on fluorescently labeling of cells. It can provide information on cell count, shape, size, DNA content, redox state, membrane permeability, and surface receptors, among other features. FC is an essential tool in research and clinical fields for tumor biology research, cancer and AIDS diagnostics, stem cell therapy, immunophenotyping, and cancer immunotherapy. One of the leading limitations of FC is that exogenously added fluorophores are not the only source of fluorescence in a sample. Autofluorescence (AF) can be, in some circumstances, a significant contributor to the signal present in each of the detection channels, often overlapping with the emission spectra. AF limits assay sensitivity and causes problems in data interpretation, which can lead to erroneous scientific conclusions. While several approaches have been employed to eliminate AF, there are no satisfactory solutions available. Kinetic River has been developing novel methods utilizing fluorescence lifetime (FLT) as an intrinsic discriminating parameter and resolving multi-exponential lifetime decays in flow cytometry. In our completed Phase I SBIR, we successfully demonstrated the automated removal of cellular AF interference in fluorophore- labeled human cells using time-resolved emission lifetimes using a 4-color system, establishing feasibility of the method. This Phase II grant will advance these findings towards commercialization by expanding the system into a fully operational 16-color Colorado analyzer with real-time data processing. The successful completion of the Colorado analyzer will lead to the first solution that automatically eliminates interference from AF in flow cytometer experiments. It will become a game-changing platform enabling cell biologists, immunologists, oncology researchers, diagnostic companies, and other FC users to obtain more accurate and quantitative results. Removal of cellular AF interference will yield enhanced sensitivity, which is important for detecting rare surface markers. This added capability of the Colorado platform will provide it with a significant competitive advantage over current commercial FC analyzers of similar multiplexing capabilities. This will enable the Colorado to replace current mid-level multiplexing instruments, which currently occupy a large swath of the rapidly growing $4.8 billion FC market.

项目摘要/摘要 流式细胞仪（FC）是一种强大的细胞分析工具，依靠细胞的荧光标记。它可以提供 有关细胞计数，形状，大小，DNA含量，氧化还原状态，膜渗透性和表面受体的信息， 除其他功能。 FC是肿瘤生物学研究的研究和临床领域的重要工具 以及AIDS诊断，干细胞疗法，免疫表型和癌症免疫疗法。领导之一 FC的局限性是外源添加的荧光团不是样品中荧光的唯一来源。 在某些情况下，自动荧光（AF）可以是每个信号中存在的重要贡献者 检测通道，通常与发射光谱重叠。 AF限制测定敏感性和原因 数据解释的问题，这可能导致错误的科学结论。而几个方法 已被用来消除AF，没有令人满意的解决方案。 动力学河一直在开发利用荧光寿命（FLT）作为固有的新方法 在流式细胞仪中区分参数和解决多指数寿命衰减。在我们的完成中 第一阶段SBIR，我们成功地证明了荧光团中细胞AF干扰的自动去除 使用时间分辨的排放寿命使用4色系统标记为人类细胞，建立了可行性 方法。该第二阶段赠款将通过将系统扩展到 具有实时数据处理的完全运行的16色科罗拉多分析仪。成功完成 科罗拉多州分析仪将导致第一个解决方案，该解决方案自动消除流量中的AF干扰 细胞仪实验。它将成为一个改变游戏规则的平台，使细胞生物学家，免疫学家， 肿瘤学研究人员，诊断公司和其他FC用户，以获取更准确和定量的 结果。切除细胞AF干扰将产生增强的灵敏度，这对于罕见 表面标记。科罗拉多州平台的这种附加功能将为它提供重要的竞争力 优于当前类似多重功能的当前商业FC分析仪。这将使 科罗拉多州以替换当前的中级多路复用器械，目前占据了一大堆 迅速增长了48亿美元的FC市场。