Interfacing Droplets with Mass Spectrometry for Single-Cell Analysis

本申请涉及RFA-GM-1-1009，特殊、非常规研究促进知识加速（尤里卡）（R 01）。任何生物系统的基本单位都是细胞，单细胞水平的故障可能导致毁灭性的疾病。因此，要了解任何生物系统的生理和功能，重要的是要实现单细胞分辨率的测量。该提案旨在采用先进的质谱技术，当与新的液滴操作技术适当结合时，将使我们能够满足使用质谱分析具有高灵敏度和信息含量的单细胞内容物的巨大挑战。具体来说，本项目的目标是：设计和构建一个质谱仪，具有适当的接口，用于电离和分析液滴中捕获的单细胞内容物。该接口将是一个液滴纳米实验室平台，具有空气中液滴的光学捕获和定位，将用于实现单液滴内容物的有效电离和质谱分析的离子传输。大气压化学电离（APCI）将用于分析单细胞的小分子（例如代谢物和脂质）含量。MALDI将用于分析单细胞的肽和蛋白质含量。细胞是任何生物系统的基本单位;因此，要了解生物体的功能和故障，就必须在单细胞水平上进行测量。然而，单细胞的生化分析是非常具有挑战性的，这是由于可获得的样品的微量以及样品的高度复杂性。在这个拟议的项目中，我们的目标是通过推动质谱法的灵敏度极限来解决这些挑战，以分析液滴中存在的微小但高度浓缩的单细胞衍生样品。 公共卫生相关性：细胞是任何生物系统的基本单位;因此，要了解生物体的功能和故障，就必须在单细胞水平上进行测量。然而，单细胞的生化分析是非常具有挑战性的，这是由于可获得的样品的微量以及样品的高度复杂性。在这个拟议的项目中，我们的目标是通过推动质谱法的灵敏度极限来解决这些挑战，以分析液滴中存在的微小但高度浓缩的单细胞衍生样品。

This application addresses RFA-GM-1-1009, Exceptional, Unconventional Research Enabling Knowledge Acceleration (EUREKA)(R01). The basic unit of any biological system is the cell, and malfunctions at the single-cell level can result in devastating diseases. To understand the physiology and functions of any biological system, therefore, it is important to achieve measurements with single-cell resolutions. This proposal aims to employ advanced mass spectrometric technologies which, when appropriately combined with new droplet manipulation techniques, will allow us to meet the tremendous challenge of analyzing the contents of single cells with high sensitivty and information content using mass spectrometry. Specifically, the aim of this project is: Design and construct a mass spectrometer with the appropriate interface for ionizing and analyzing the contents of single cells trapped in liquid droplets. The interface will be a droplet nanolab platform with the optical trapping and positioning of droplets in air, which will be used towards achieving the efficient ionization of single-droplet contents and ion transport for mass spectrometry analysis. Atmospheric pressure chemical ionization (APCI) will be used for the analysis of small-molecule (e.g. metabolites and lipids) contents of single cells. MALDI will be used for the analysis of peptide and protein contents of single cells. The cell is the basic unit of any biological system; to understand the functions and malfunctions of an organism will thus necessitate measurements at the single-cell level. Yet the biochemical analysis of single cells is highly challenging, owing to both the minute amounts of sample available as well as the high complexity of the sample. In this proposed project, we aim to tackle these challenges by pushing the limit of sensitivity of mass spectrometry for analyzing the minute but highly concentrated single-cell derived samples present in droplets. PUBLIC HEALTH RELEVANCE: The cell is the basic unit of any biological system; to understand the functions and malfunctions of an organism will thus necessitate measurements at the single-cell level. Yet the biochemical analysis of single cells is highly challenging, owing to both the minute amounts of sample available as well as the high complexity of the sample. In this proposed project, we aim to tackle these challenges by pushing the limit of sensitivity of mass spectrometry for analyzing the minute but highly concentrated single-cell derived samples present in droplets.