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Cryo-electron Microscopy

Cryo-electron Microscopy

Conventional electron microscopy (EM) involves dehydration, fixation, and staining of biological samples, which distorts the native state of biological molecules and results in several artifacts. Also, the high-energy electron beam damages the sample and makes it difficult to obtain high-resolution images. These issues can be addressed using cryo-EM, which uses frozen samples and gentler electron beams. The technique was developed by Jacques Dubochet, Joachim Frank, and Richard Henderson, for...

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Updated: Nov 2, 2025

Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells

Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells

Published on: May 28, 2021

Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells.

Nina Vyas¹, Nina Perry², Chidinma A Okolo²

¹Harwell Science and Innovation Campus, Beamline B24, Diamond Light Source; nina.vyas@diamond.ac.uk.

Journal of Visualized Experiments : Jove

|June 14, 2021

Summary

This summary is machine-generated.

Three-dimensional structured illumination microscopy (3D SIM) offers enhanced resolution for imaging cellular structures. This protocol details using a cryoSIM microscope for high-resolution 3D imaging of biological samples at cryogenic temperatures.

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Last Updated: Nov 2, 2025

Cryo-Structured Illumination Microscopic Data Collection from Cryogenically Preserved Cells

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Published on: May 28, 2021

Electron Cryotomography of Bacterial Cells

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Published on: May 6, 2010

A 3D Cartographic Description of the Cell by Cryo Soft X-ray Tomography

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Published on: March 15, 2021

Area of Science:

Biophysics
Cell Biology
Microscopy

Background:

Conventional fluorescence microscopy has resolution limitations for visualizing molecular processes.
Super-resolution microscopy techniques, like 3D SIM, overcome these limitations.
Correlative imaging requires compatible sample preparation for multiple microscopy techniques.

Purpose of the Study:

To provide detailed methods and protocols for operating a cryoSIM microscope.
To enable high-resolution 3D imaging of cryogenically preserved biological samples.
To facilitate correlative imaging with X-ray microscopy techniques.

Main Methods:

Commissioning of a 3D SIM microscope for cryogenic samples (cryoSIM) at the UK synchrotron B24 beamline.
Development of protocols for sample preparation, fluorophore selection, and parameter settings for cryoSIM.
Demonstration of the protocol using fluorescently labelled U2OS cells (mitochondria and tubulin).

Main Results:

Successful implementation of 3D SIM for imaging biological samples at cryogenic temperatures.
Demonstrated compatibility of cryoSIM with subsequent X-ray microscopy methods.
Provided practical guidelines for optimizing cryoSIM imaging parameters and sample choices.

Conclusions:

The cryoSIM microscope enables high-resolution 3D imaging of biological specimens under cryogenic conditions.
This technique is valuable for correlative studies, bridging light and X-ray microscopy.
The detailed protocol facilitates the adoption and application of cryoSIM in biological research.