The future is cold: cryo-preparation methods for transmission electron microscopy of cells
- 1 September 2011
- journal article
- review article
- Published by Wiley in Biology of the Cell
- Vol. 103 (9), 405-420
- https://doi.org/10.1042/bc20110015
Abstract
Our knowledge of the organization of the cell is linked, to a great extent, to light and electron microscopy. Choosing either photons or electrons for imaging has many consequences on the image obtained, as well as on the experiment required in order to generate the image. One apparent effect on the experimental side is in the sample preparation, which can be quite elaborate for electron microscopy. In recent years, rapid freezing, cryo-preparation and cryoelectron microscopy have been more widely used because they introduce fewer artefacts during preparation when compared with chemical fixation and room temperature processing. In addition, cryo-electron microscopy allows the visualization of the hydrated specimens. In the present review, we give an introduction to the rapid freezing of biological samples and describe the preparation steps. We focus on bulk samples that are too big to be directly viewed under the electron microscope. Furthermore, we discuss the advantages and limitations of freeze substitution and cryo-electron microscopy of vitreous sections and compare their application to the study of bacteria and mammalian cells and to tomography.Keywords
This publication has 87 references indexed in Scilit:
- Exploring vitreous cryo-section-induced compression at the macromolecular level using electron cryo-tomography; 80S yeast ribosomes appear unaffectedJournal of Structural Biology, 2011
- CEMOVIS on a pathogen: analysis of Bacillus anthracis sporesBiology of the Cell, 2010
- Cryo-electron tomography of microtubule–kinesin motor complexesJournal of Structural Biology, 2010
- Improving the technique of vitreous cryo-sectioning for cryo-electron tomography: Electrostatic charging for section attachment and implementation of an anti-contamination glove boxJournal of Structural Biology, 2010
- Preservation of high resolution protein structure by cryo-electron microscopy of vitreous sectionsUltramicroscopy, 2009
- Classification and 3D averaging with missing wedge correction in biological electron tomographyJournal of Structural Biology, 2008
- Markov random field based automatic image alignment for electron tomographyJournal of Structural Biology, 2008
- Disclosure of the mycobacterial outer membrane: Cryo-electron tomography and vitreous sections reveal the lipid bilayer structureProceedings of the National Academy of Sciences of the United States of America, 2008
- Dual-Axis Tomography: An Approach with Alignment Methods That Preserve ResolutionJournal of Structural Biology, 1997
- Cryo-electron microscopy of vitrified specimensQuarterly Reviews of Biophysics, 1988