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.

项目概要/摘要 流式细胞术 (FC) 是一种强大的细胞分析工具，依赖于细胞的荧光标记。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， 除其他功能外。 FC 是肿瘤生物学研究、癌症研究和临床领域的重要工具 艾滋病诊断、干细胞治疗、免疫表型分析和癌症免疫治疗。领先者之一 FC 的局限性在于外源添加的荧光团并不是样品中唯一的荧光源。 在某些情况下，自发荧光 (AF) 可能是每个细胞中存在的信号的重要贡献者。 检测通道，通常与发射光谱重叠。 AF 限制测定灵敏度及其原因 数据解释中的问题可能导致错误的科学结论。虽然有几种方法 虽然已采用消除 AF 的方法，但尚无令人满意的解决方案。 Kinetic River 一直在开发利用荧光寿命 (FLT) 作为内在特征的新方法 判别参数并解决流式细胞术中的多指数寿命衰减。在我们完成的 I 期 SBIR，我们成功演示了荧光团中细胞 AF 干扰的自动消除 使用 4 色系统使用时间分辨发射寿命标记人体细胞，建立了该方法的可行性 方法。第二阶段的资助将通过将该系统扩展到 具有实时数据处理功能的全功能 16 色科罗拉多分析仪。此次活动的顺利完成 科罗拉多分析仪将推出第一个自动消除流动中 AF 干扰的解决方案 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 用户获得更准确和定量的信息 结果。消除细胞 AF 干扰将提高灵敏度，这对于检测罕见的 表面标记。科罗拉多平台的这一附加功能将为其提供显着的竞争优势 与当前具有类似多路复用功能的商用 FC 分析仪相比具有优势。这将使 科罗拉多州将取代当前的中级多路复用仪器，这些仪器目前占据了大片地区 快速增长的价值 48 亿美元的 FC 市场。