visualization-of-organelles-in-situ-by-cryo-stem-tomography

Visualization of Organelles <em>In Situ</em> by Cryo-STEM Tomography

Cryogenic electron microscopy (cryo-EM) relies on the imaging of biological or organic specimens embedded in their native aqueous medium; water is ...

Weizmann Institute of Sciences

Electron cryo-tomography using the scanning transmission modality (STEM) enables 3D reconstruction of unstained, vitrified specimens as thick as 1μm or more. Contrast is related to mass/thickness and atomic number, providing quantifiable chemical characterization and mass mapping of intact prokaryotic and eukaryotic cells. Energy dispersive X-ray spectroscopy by STEM provides a simple, on-the-spot chemical identification of the elemental composition in sub-cellular organic bodies or mineral deposits. This chapter provides basic background and practical information for performing cryo-STEM tomography on vitrified biological cells.

CryoSTEM tomography in biology.

In this issue of Structure, breakthroughs in cryo-EM/ET research are presented. Klebl et al. (2020) demonstrate how speed in sample vitrification impacts the quality of macromolecular particles in resultant cryo-EM grids. Wu et al. (2020) combine fluorescence, ion beam milling, and tomography to unravel unique features in vitrified yeast cells.

Maintaining Context in Ice: Cryo-EM/ET Workflow Optimizations.

Electron microscopy in three dimensions (3D) of cells and tissues can be essential for understanding the ultrastructural aspects of biological processes. The quest for 3D information reveals challenges at many stages of the workflow, from sample preparation, to imaging, data analysis and segmentation. Here, we outline several available methods, including volume SEM imaging, cryo-TEM and cryo-STEM tomography, each one occupying a different domain in the basic tradeoff between field-of-view and resolution. We discuss the considerations for choosing a suitable method depending on research needs and highlight recent developments that are essential for making 3D volume imaging of cells and tissues a standard tool for cellular and structural biologists.

Article	Year
CryoSTEM tomography in biology. Methods in cell biology\| PubMed ID: 31326021	2019
Maintaining Context in Ice: Cryo-EM/ET Workflow Optimizations. Structure (London, England : 1993)\| PubMed ID: 33147474	2020
Electron microscopy of cellular ultrastructure in three dimensions. Current opinion in structural biology\| PubMed ID: 36041268	2022

Sharon Grayer Wolf

Biography

JoVE Journal Publications

Other Publications