A Microfluidic/Nanofluidic System for Rapid Single Cell Proteomics

用于快速单细胞蛋白质组学的微流控/纳流控系统

• 批准号: 10547040
• 负责人: Aaron T Timperman
• 金额: $ 40万
• 依托单位: FLUIDISPEC LLC
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10547040
• 关键词: Adsorption; Biological; Biological Process; Biological Sciences; Cell Separation; Cells; Charge; Complex; Cytolysis; Data; Detection; Development; Digestion; Dimensions; Disease; Electrophoresis; Fee-for-Service Plans; Health; Health Sciences; Hela Cells; Heterogeneity; Image; Individual; Injections; Ions; Liquid Chromatography; Mass Spectrum Analysis; Measurement; Methods; Metric System; Microfluidics; Modification; Optics; Peptides; Performance; Pharmaceutical Preparations; Pharmacotherapy; Population; Post-Translational Protein Processing; Power Sources; Preparation; Protein Analysis; Proteins; Proteome; Proteomics; Pump; Research; Robot; Sample Size; Sampling; Speed; Stimulus; System; Technology; Time; Tissues; base; cell type; cellular imaging; commercialization; cost; design; detector; environmental stressor; improved; improved functioning; innovation; insight; instrumentation; liquid chromatography mass spectrometry; mass spectrometer; nanofluidic; nanoscale; novel; optical imaging; performance tests; research and development; response; stressor; tool; tumor; voltage

Proteomics typically involves the analysis of the protein components of large populations of cells. Nearly all the of a cell’s machinery are proteins, so many diseases and responses to stimuli and stressors are manifested at the protein level. Proteomes are challenging to characterize fully for many reasons. Some of the key attributes of a proteome that make its characterization so challenging are: the large number of proteins expressed in a single cell, the wide dynamic range of protein relative abundance, and the numerous modifications that can switch proteins between active an inactive states. To further increase complexity, significant heterogeneity exists at the cellular level that is averaged out by bulk analyses. To better understand biological and health implications of cellular heterogeneity, single cell proteomics has emerged. However, improvements in the enabling analytical technologies are needed as single-cell proteomics adds the additional challenge of extremely small sample size. This application focuses on the development of a novel electrokinetic microfluidic/nanofluidic system for improved single-cell proteome analyses that is a significant deviation from the current ultra-performance liquid chromatography (UPLC)-mass spectrometry (MS) based approaches. The proposed electrokinetic microfluidic/nanofluidic system is expected to offer improved performance from: reduced sample loss, increased sample concentration eluted into the mass spectrometer, reduced peptide contamination, high efficiency separations, and rapid analyses. The Aims of this application focus on: 1) fabrication and optimization of the key functional modules, and 2) integration of these functional modules into a complete system and measurement of the performance of the complete system. The primary metrics for system performance will be the speed of the analysis and the number of protein groups identified. If successful, this application will provide a novel system for single-cell proteomics that will improve the analysis speed without sacrificing depth of proteome coverage.

蛋白质组学通常涉及对大量细胞的蛋白质成分的分析。 细胞的所有机制都是蛋白质，因此许多疾病以及对刺激和压力的反应都与蛋白质有关。 由于多种原因，蛋白质组的表征具有挑战性。 蛋白质组的关键属性使其表征如此具有挑战性，其中包括： 单细胞中表达的蛋白质的数量、蛋白质相对丰度的宽动态范围以及 许多修饰可以使蛋白质在活性和非活性状态之间切换。 由于复杂性，细胞水平上存在显着的异质性，通过批量分析来平均。 更好地了解细胞异质性的生物学和健康影响，单细胞蛋白质组学已经 然而，作为单细胞，需要改进支持分析技术。 蛋白质组学增加了极小样本量的额外挑战。 开发新型动电微流体/纳米流体系统以改进单细胞蛋白质组 与当前超高效液相色谱法有显着偏差的分析 所提出的基于电动微流控/纳米流控（UPLC）-质谱（MS）的方法。 系统预计将通过以下方式提供改进的性能：减少样品损失、增加样品 浓度洗脱进入质谱仪，减少肽污染，效率高 该应用的目标集中在：1）制造和优化。 关键功能模块，2）将这些功能模块集成到一个完整的系统中 衡量整个系统性能的主要指标。 如果成功，将是分析的速度和识别的蛋白质组的数量。 该应用将为单细胞蛋白质组学提供一种新颖的系统，从而提高分析速度 不牺牲蛋白质组覆盖深度。