From Single Cells to Tissues: a Novel Mass Spectrometry Approach for Bioanalysis

从单细胞到组织：一种新颖的生物分析质谱方法

• 批准号: 9469104
• 负责人: Zhibo Yang
• 金额: $ 2.01万
• 依托单位: UNIVERSITY OF OKLAHOMA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9469104
• 关键词: Animals; Antineoplastic Agents; Atlas of Cancer Mortality in the United States; Biological; Biology; Biomedical Research; Biomimetics; Breast Cancer Cell; Cell Line; Cell physiology; Cells; Cellular Spheroids; Cellular biology; Ceramides; Chemicals; Cholesterol; Clinical Research; Cultured Cells; Detection; Devices; Doxorubicin; Eukaryotic Cell; Fiber; Goals; Image; Imaging Techniques; In Situ; In Vitro; Individual; Investigation; Knowledge; Lipids; Location; Mammary Neoplasms; Maps; Mass Spectrum Analysis; Measurement; Measures; Medical Research; Methods; Microscopy; Modeling; Molecular; Molecular Analysis; Motion; Mus; Organism; Paclitaxel; Peptides; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Pharmacotherapy; Population; Population Heterogeneity; Preparation; Research; Resolution; Sampling; Science; Silicon Dioxide; Slice; Software Engineering; Spatial Distribution; Stable Isotope Labeling; Structure; System; Techniques; Technology; Time; Tissue Sample; Tissue imaging; Tissues; Tumor Biology; Tumor Tissue; Tumor-Derived; analytical method; animal tissue; base; biological research; cancer cell; experimental study; human disease; imaging modality; imaging study; improved; innovative technologies; instrumentation; ionization; miniaturize; neoplastic cell; novel; scaffold; single cell analysis; tool; tumor

There is a fundamental lack of understanding in biology on the single cell level, especially in human disease biology. This gap in knowledge encompasses both how individual cells function and also how single cells combine and collaborate to construct multi-cellular creations, such as how many cancer cells associate to form a tumor. Traditionally, bioanalysis has largely been conducted through large numbers (i.e. thousands or millions) of cells being destroyed and mixed together to form a completely artificial construct: the cell lysate. Cell lysates are prepared from a large number of heterogeneous cells, either present in tissue samples or grown in culture. Bioanalysis of lysates are an amalgamated average of many heterogeneous cells subjected to artificial sample preparation, and therefore, standard bioanalytical methods have been largely unable to define single cell biology. Recently, single cell analysis (SCA) has become possible through advancements in many experimental methods. Due to its unparalleled detection sensitivity, mass spectrometry (MS) is one of the most promising analytical methods to be deployed for single cell analysis. Single-cell mass spectrometry (SCMS) and mass spectrometry imaging (MSI) are emerging techniques that could potentially revolutionize bioanalytical science in basic scientific and biomedical research. We have developed a miniaturized sampling and ionization device, the Single-probe, capable of performing single cell mass spectrometry (SCMS) in real time under ambient conditions. The Single-probe, with a sampling tip smaller than eukaryotic cells (<10 μm), can be inserted into individual living cells to sample the intracellular compounds for immediate MS analysis. The Single-probe SCMS technique has produced highly sensitive detection of intracellular compounds, including cell metabolites, lipids and anticancer compounds, inside living single cells. The Single-probe has also been used successfully in MSI experiments to reconstruct the spatial distribution of species (e.g. lipids and cell metabolites) from sections of animal tissue. Our long-term research goal is to fully establish the Singleprobe based techniques in SCMS and MSI for broad utilization in bioanalysis. The objective of this application is to develop the Single-probe MS technology as a first-in-class tool for quantitative SCMS (qSCMS) in cells and for MSI-based investigation of tumors. This research objective will be accomplished through completing the following three Specific Aims: 1) create quantitative single cell mass spectrometry (qSCMS) for compounds inside of single cells from cultured cell lines; 2) use the Single-probe MS to analyze single cells in in vitro tumors; and, 3) perform Single-probe MSI measurements of the in vitro and animal-derived tumors. The Single-probe MS is an innovative technology in that it provides new experimental capabilities for single cell analysis and tissue imaging. The proposed research is significant because the Single-probe MS experimental applications (i.e. qSCMS and MSI), once developed, will allow new understandings in single cell and tumor biology that are not currently possible.

人们对单细胞水平的生物学缺乏根本性的了解，尤其是在人类 疾病生物学。这种知识差距既包括单个细胞如何发挥作用，也包括单个细胞如何发挥作用。 细胞结合并协作构建多细胞创造物，例如有多少癌细胞与 形成肿瘤。传统上，生物分析主要是通过大量（即数千或 数百万）细胞被破坏并混合在一起形成完全人工的结构：细胞裂解物。 细胞裂解物由大量异质细胞制备而成，这些细胞存在于组织样品中或 在文化中成长。裂解物的生物分析是许多异质细胞的合并平均值 人工样品制备，因此，标准生物分析方法基本上无法 定义单细胞生物学。最近，随着技术的进步，单细胞分析（SCA）已成为可能。 许多实验方法。由于其无与伦比的检测灵敏度，质谱 (MS) 是一种 用于单细胞分析的最有前途的分析方法。单细胞质谱分析 （SCMS）和质谱成像（MSI）是新兴技术，可能会彻底改变 基础科学和生物医学研究中的生物分析科学。我们开发了一种小型化采样 电离装置，单探针，能够实时进行单细胞质谱（SCMS） 环境条件下的时间。单探针，采样头比真核细胞小（<10 μm）， 可以插入单个活细胞中，对细胞内化合物进行采样，以便立即进行 MS 分析。 单探针 SCMS 技术可对细胞内化合物进行高度灵敏的检测， 包括活单细胞内的细胞代谢物、脂质和抗癌化合物。单探头有 也已成功用于 MSI 实验，以重建物种的空间分布（例如脂质 和细胞代谢物）来自动物组织切片。我们的长期研究目标是全面建立Singleprobe 基于 SCMS 和 MSI 的技术在生物分析中得到广泛应用。此应用程序的目的 旨在开发单探针 MS 技术作为细胞定量 SCMS (qSCMS) 的一流工具 以及基于 MSI 的肿瘤研究。该研究目标将通过完成 以下三个具体目标：1) 创建化合物的定量单细胞质谱 (qSCMS) 来自培养细胞系的单细胞内部； 2) 使用单探针质谱仪进行体外单细胞分析 肿瘤； 3) 对体外和动物源性肿瘤进行单探针 MSI 测量。这 单探针质谱是一项创新技术，为单细胞提供了新的实验能力 分析和组织成像。所提出的研究意义重大，因为单探针 MS 实验 应用程序（即 qSCMS 和 MSI）一旦开发出来，将有助于对单细胞和肿瘤有新的认识 目前不可能的生物学。