Maastricht MultiModal Molecular Imaging institute (M4I), Maastricht University, Maastricht, the Netherlands.
Department of Surgery, NUTRIM School of Nutrition and Translational Research in Metabolism, Maastricht University, Maastricht, the Netherlands.
Nat Protoc. 2022 Apr;17(4):962-979. doi: 10.1038/s41596-021-00661-8. Epub 2022 Feb 18.
Organoid culture systems are self-renewing, three-dimensional (3D) models derived from pluripotent stem cells, adult derived stem cells or cancer cells that recapitulate key molecular and structural characteristics of their tissue of origin. They generally form into hollow structures with apical-basolateral polarization. Mass spectrometry imaging (MSI) is a powerful analytical method for detecting a wide variety of molecules in a single experiment while retaining their spatiotemporal distribution. Here we describe a protocol for preparing organoids for MSI that (1) preserves the 3D morphological structure of hollow organoids, (2) retains the spatiotemporal distribution of a vast array of molecules (3) and enables accurate molecular identification based on tandem mass spectrometry. The protocol specifically focuses on the collection and embedding of the organoids in gelatin, and gives recommendations for MSI-specific sample preparation, data acquisition and molecular identification by tandem mass spectrometry. This method is applicable to a wide range of organoids from different origins, and takes 1 d from organoid collection to MSI data acquisition.
类器官培养系统是源自多能干细胞、成体来源干细胞或癌细胞的自我更新的三维（3D）模型，可重现其起源组织的关键分子和结构特征。它们通常形成具有顶-基底极性的中空结构。质谱成像（MSI）是一种强大的分析方法，可在单个实验中同时检测多种分子，同时保留其时空分布。在这里，我们描述了一种用于 MSI 的类器官制备方案，该方案（1）保留中空类器官的 3D 形态结构，（2）保留大量分子的时空分布，（3）并能够基于串联质谱进行准确的分子鉴定。该方案特别侧重于在明胶中收集和包埋类器官，并针对 MSI 特定的样品制备、数据采集和串联质谱的分子鉴定提出建议。该方法适用于来自不同来源的广泛的类器官，从类器官收集到 MSI 数据采集需要 1 天时间。
