细胞器和囊泡的电子断层扫描在SNARE功能与定位研究中的应用

Electron Tomography of Organelles and Vesicles in the Investigation of SNARE Function and Localization.

作者信息

Bright Nicholas A, Luzio J Paul

机构信息

Cambridge Institute for Medical Research (CIMR) and Department of Clinical Biochemistry, University of Cambridge School of Clinical Medicine, Cambridge, UK.

出版信息

Methods Mol Biol. 2025;2887:237-247. doi: 10.1007/978-1-0716-4314-3_17.

DOI:10.1007/978-1-0716-4314-3_17

PMID:39806159

Abstract

Electron tomography can provide additional morphological information not easily obtained by conventional transmission electron microscopy of thin sections. It uses a goniometer stage in the electron microscope to tilt the specimen and collect a series of 2D images from different orientations, which are combined to provide a 3D volume tomogram and a colored reconstruction of the morphological feature(s) of interest. Here we describe the protocols for its use in visualizing changes in organelle morphology after depletion of the SNARE proteins VAMP7 and VAMP8 and to study VAMP7 localization on endolysosomes/lysosomes.

摘要

电子断层扫描能够提供一些额外的形态学信息，而这些信息通过传统的薄切片透射电子显微镜不易获得。它在电子显微镜中使用测角仪载物台来倾斜样本，并从不同方向收集一系列二维图像，这些图像被组合起来以提供一个三维体积断层图像以及对感兴趣的形态特征进行彩色重建。在这里，我们描述了使用该技术来可视化SNARE蛋白VAMP7和VAMP8缺失后细胞器形态变化以及研究VAMP7在内溶酶体/溶酶体上定位的实验方案。