Sample Preparation of Isolated Mitochondria for Cryoelectron Tomography and In Situ Studies of Translation

Lena Thärichen, Robert Englmeier, Friedrich Förster*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterAcademic

Abstract

Cryoelectron tomography is a method to image biological samples three-dimensionally at molecular resolution. This modality provides insights into intracellular processes in their physiological settings. Obtaining a high-quality sample for cryoelectron tomography on mitochondria, however, can be challenging. In this chapter, we describe the crucial steps from sample preparation to data acquisition enabling studies of mitochondrial translation in situ by cryoelectron tomography. We provide detailed protocols for yeast and human mitochondria preparations yielding a high concentration of intact mitochondrial vesicles on cryo-EM grids. In addition, we describe a workflow for particle identification and spatial mapping in context of the organelle.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
Subtitle of host publicationMethods and Protocols
EditorsAntoni Barrientos, Flavia Fontanesi
Place of PublicationNew York
PublisherHumana Press
Chapter5
Pages75-88
Number of pages14
Edition1
ISBN (Electronic)978-1-0716-3173-7
ISBN (Print)978-1-0716-3170-6
DOIs
Publication statusPublished - 12 May 2023

Publication series

NameMethods in Molecular Biology
Volume2661
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • Cryoelectron tomography
  • Mitochondria
  • Ribosome
  • Translation

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