New Technologies for detecting extracellular fluxes

检测细胞外通量的新技术

• 批准号: 10213904
• 负责人: WILLIAM R KOBERTZ
• 金额: $ 37.17万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 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 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10213904
• 关键词: Action Potentials; Animals; Astrocytes; Bacteria; Benchmarking; Brain; Cardiac; Cell membrane; Cells; Coculture Techniques; Communication; Data; Disease; Engineering; Fluorescence; Fluorescence Resonance Energy Transfer; Glutamates; Glycocalyx; Goals; Grant; Health; Homeostasis; Human; Individual; Ion Channel; Ions; Kinetics; Label; Mammalian Cell; Membrane Potentials; Membrane Proteins; Membrane Transport Proteins; Metabolic; Microglia; Muscle; Muscle Contraction; Muscle function; Neurons; Pathway interactions; Periodicity; Physiological; Positioning Attribute; Proteins; Quality Control; Reagent; Research Personnel; Rest; Series; Signal Transduction; Surface; Techniques; Technology; Thick; Time; Tissues; Viral; Visualization; Wheat Germ Agglutinins; cell type; chemical genetics; extracellular; genetic approach; heart function; human disease; nanometer; neuronal excitability; new technology; non-genetic; polyhistidine; prevent; prototype; sensor; small molecule; spatiotemporal; sugar; technology development; technology research and development; uptake

Abstract: The goal of this Focused Technology and Development grant is to repurpose intracellular fluorescent ion and metabolite sensors to detect extracellular fluxes. The three orthogonal approaches directly target the cell’s glycocalyx: the nanometer thick, sugar‐coating where ion and metabolite concentrations vary greatly during ion channel and membrane transporter activity. Aim 1 describes an approach to attach any fluorescent protein sensor to cells metabolically-labeled with an unnatural azidosugar. Aim 2 is a non-genetic approach that labels all mammalian cells—including human cells—with both small molecule and protein fluorescent sensors. Aim 3 is a chemical genetic approach to achieve cell-type-specific labeling with small molecule and protein fluorescent sensors. Completion of the aims will convert these working prototypes that utilize three major classes of fluorescent sensors (small molecule, protein, and FRET) into technologies that enable the visualization of ions and metabolites at the surface of a primary cells, tissues, and potentially in living animals.

抽象的： 这项重点技术和开发拨款的目标是重新利用细胞内 荧光离子和代谢物传感器检测细胞外通量。三个正交 方法直接针对细胞的糖萼：纳米厚的糖衣，其中离子 离子通道和膜转运过程中代谢物浓度变化很大 活动。目标 1 描述了一种将任何荧光蛋白传感器附着到细胞上的方法 用非天然叠氮糖进行代谢标记。目标 2 是一种非遗传方法 用小分子和蛋白质标记所有哺乳动物细胞（包括人类细胞） 荧光传感器。目标 3 是一种实现细胞类型特异性标记的化学遗传方法 具有小分子和蛋白质荧光传感器。目标的完成将把这些 利用三大类荧光传感器（小分子、 蛋白质和 FRET）转化为能够实现离子和代谢物可视化的技术 原代细胞、组织的表面，也可能存在于活体动物的表面。