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线粒体转位的STED超分辨率显微镜

Sarah V Schweighofer¹, Kaushik Inamdar², Daniel C Jans²

¹Department of NanoBiophotonics, Max Planck Institute for Multidisciplinary Sciences, Göttingen, Germany; Translational Neuroinflammation and Automated Microscopy TNM, Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, Göttingen, Germany.

Methods in enzymology

|November 2, 2024

概括

本章详细介绍了刺激排放枯竭 (STED) 超分辨率显微镜的协议,以可视化线粒体蛋白质进口复合体,特别是TOM20,以及固定细胞中的线粒体DNA.

科学领域:

细胞生物学细胞生物学
线粒体生物学线粒体生物学
显微镜技术显微镜技术

背景情况:

外膜 (TOM) 和内膜 (TIM) 的线粒体转位对于蛋白质进口到线粒体至关重要.
了解这些复杂的空间组织对于阐明线粒体功能至关重要.
刺激发射衰减 (STED) 超分辨率显微镜为现场细胞成像提供亚衍射分辨率.

研究的目的:

为STED显微镜TOM20和固定哺乳动物细胞中的线粒体DNA提供详细的协议.
为了使线粒体蛋白质进口机械在其原生细胞环境中以高分辨率可视化.
提供一种可适应的方法,使用STED显微镜对其他细胞点进行成像.

主要方法:

详细的样本准备用于免疫标记,包括细胞固定,透,阻断和抗体标记.
用于STED显微镜的样品安装的具体说明.
评估样品和显微镜性能以获得最佳图像的建议.

主要成果:

使用STED超分辨率显微镜成功对TOM20和线粒体DNA进行现场成像.
展示STED显微镜能够在没有复杂的图像处理的情况下提供超高分辨率的原始数据的能力.
验证常规样本准备技术用于线粒体组件的STED成像.

结论:

提供的协议使线粒体蛋白质进口复合体和DNA的高分辨率成像成为可能.
STED显微镜是一种强大的,可访问的技术,用于研究线粒体超结构和动力学.
该协议很容易适应用于研究各种细胞结构和过程.

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