Gemperline Erin, Li Lingjun
Department of Chemistry, University of Wisconsin- Madison.
Department of Chemistry, University of Wisconsin- Madison; School of Pharmacy, University of Wisconsin- Madison;
J Vis Exp. 2014 Mar 5(85):51434. doi: 10.3791/51434.
Most techniques used to study small molecules, such as pharmaceutical drugs or endogenous metabolites, employ tissue extracts which require the homogenization of the tissue of interest that could potentially cause changes in the metabolic pathways being studied(1). Mass spectrometric imaging (MSI) is a powerful analytical tool that can provide spatial information of analytes within intact slices of biological tissue samples(1-5). This technique has been used extensively to study various types of compounds including proteins, peptides, lipids, and small molecules such as endogenous metabolites. With matrix-assisted laser desorption/ionization (MALDI)-MSI, spatial distributions of multiple metabolites can be simultaneously detected. Herein, a method developed specifically for conducting untargeted metabolomics MSI experiments on legume roots and root nodules is presented which could reveal insights into the biological processes taking place. The method presented here shows a typical MSI workflow, from sample preparation to image acquisition, and focuses on the matrix application step, demonstrating several matrix application techniques that are useful for detecting small molecules. Once the MS images are generated, the analysis and identification of metabolites of interest is discussed and demonstrated. The standard workflow presented here can be easily modified for different tissue types, molecular species, and instrumentation.
大多数用于研究小分子（如药物或内源性代谢物）的技术都采用组织提取物，这需要对感兴趣的组织进行匀浆处理，而这可能会潜在地导致正在研究的代谢途径发生变化(1)。质谱成像（MSI）是一种强大的分析工具，能够提供生物组织样本完整切片内分析物的空间信息(1 - 5)。该技术已被广泛用于研究各种类型的化合物，包括蛋白质、肽、脂质以及内源性代谢物等小分子。通过基质辅助激光解吸/电离（MALDI）-MSI，可以同时检测多种代谢物的空间分布。在此，介绍一种专门为在豆科植物根和根瘤上进行非靶向代谢组学MSI实验而开发的方法，该方法可以揭示正在发生的生物过程的见解。这里介绍的方法展示了一个典型的MSI工作流程，从样品制备到图像采集，并重点关注基质应用步骤，展示了几种对检测小分子有用的基质应用技术。一旦生成MS图像，将讨论并展示对感兴趣的代谢物的分析和鉴定。这里介绍的标准工作流程可以很容易地针对不同的组织类型、分子种类和仪器进行修改。
