Context rich mass spectrometry of molecular localization and cellular interactions

分子定位和细胞相互作用的上下文丰富质谱分析

• 批准号: 9751908
• 负责人: Alexander Revzin
• 金额: $ 47.88万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-06 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9751908
• 关键词: Address; Affect; Antibodies; Antigen-Presenting Cells; Biological; Biological Phenomena; Biological Sciences; Biosensor; Caliber; Cell Communication; Cell Surface Proteins; Cell secretion; Cell surface; Cells; Cellular Structures; Cytometry; Cytotoxic T-Lymphocytes; Development; Electron Microscopy; Ensure; Event; Flow Cytometry; Future; Glass; Goals; Heart; Hydration status; Hydrogels; Immune; Infection prevention; Invaded; Ions; Knowledge; Label; Leprosy; Location; Mass Spectrum Analysis; Metals; Methodology; Methods; Molecular; Multiparametric Analysis; Phenotype; Positioning Attribute; Preparation; Protein Analysis; Proteins; Rare Earth Metals; Sampling; Science; Shapes; Specific qualifier value; Surface; Suspensions; Synapses; T-Lymphocyte; Technology; Time; Vacuum; base; biological systems; cytokine; cytotoxic; ethylene glycol; experimental study; immunological synapse; instrument; instrumentation; interest; macrophage; nano; nanoscale; new technology; novel; novel strategies; pathogen; pressure; protein expression; relating to nervous system; tool

ABSTRACT How does the molecular composition of cells and cell-cell contacts change over time or after experimental manipulations? This fundamental question is asked across all fields of biomedical science and is at the heart of most studies focused on determining cellular and molecular mechanisms of biological phenomenon. Yet, our ability to answer this question is severely constrained by the limitations of current methods to detect the protein composition of particular cellular structures. Imagine the rapid progress biological science could make if we could detect more than a handful of proteins at specified locations on cells in each experiment. The objective of our project is to develop a new approach to achieve this goal—called “context-rich mass cytometry” –a technology that enables multi-parametric analysis of proteins in the context of cellular interactions. Mass spectrometry has recently emerged as a powerful tool for cell analysis. One version of this technology called mass cytometry or CyToF is used for multi-parametric analysis of protein expression in single cells. It is similar to flow cytometry in that cells are labeled with antibodies and analyzed on cell-by-cell basis in high- throughput manner. However, unlike flow cytometry which employs fluorescently-labeled antibodies and is limited to ~12 parameters, mass cytometry employs rare earth metals as antibody labels and may be used to analyze up to 60 intracellular or cell surface markers. While an exciting technology, mass cytometry requires that cells be extracted from their native microenvironment and homogenized into single cell suspension prior to analysis. This makes connecting the protein signature to the microenvironment context very challenging. The objective of our project is to develop “context-rich mass cytometry” –a technology that enables multi- parametric analysis of proteins in the context of cellular interactions. Once developed this mass spectrometry approach will be used to determine the composition and organization of proteins within immune synapses. This new knowledge enabled by our technology may, in the future, be parlayed into strategies and therapies for preventing infections.

抽象的 细胞的分子组成和细胞与细胞的接触如何随时间或实验后发生变化 这个基本问题是整个生物医学科学的核心问题。 大多数研究的重点是确定生物现象的细胞和分子机制。 我们回答这个问题的能力受到当前检测方法的局限性的严重限制 想象一下，如果特定细胞结构的蛋白质组成，生物科学将会取得怎样的快速进步。 在每个实验中，我们可以在细胞上的特定位置检测到多种蛋白质。 我们项目的目标是开发一种新方法来实现这一目标，称为“上下文丰富的大众” 细胞计数术”——一种能够在细胞环境中对蛋白质进行多参数分析的技术 互动。 质谱法最近已成为该技术的一种强大工具。 称为质谱流式细胞术或 CyToF 用于单细胞中蛋白质表达的多参数分析。 与流式细胞术类似，细胞用抗体标记，并在高通量分析中逐个细胞地进行分析。 然而，采用荧光标记抗体的各种流式细胞术是不可行的。 限于约 12 个参数，质谱流式细胞仪采用稀土金属作为抗体标记，可用于 分析多达 60 个细胞内或细胞表面标记物虽然是一项令人兴奋的技术，但质谱流式分析需要。 细胞从其天然微环境中提取并匀浆成单细胞悬浮液，然后再进行 这使得将蛋白质特征与微环境联系起来非常具有挑战性。 我们项目的目标是开发“上下文丰富的质谱细胞术”——一种能够实现多 一旦开发出这种质谱法，就可以在细胞相互作用的背景下对蛋白质进行参数分析。 该方法将用于确定免疫突触内蛋白质的组成和组织。 我们的技术所带来的新知识在未来可能会被运用到策略和治疗中 用于预防感染。