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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...

Scanning Electron Microscopy

Scanning Electron Microscopy

A scanning electron microscope (SEM) is used to study the surface features of a sample by using an electron beam that scans the sample surface in a two-dimensional manner. Typically, areas between ~1 centimeter to 5 micrometers in width can be imaged. SEM can be used to image bacteria, viruses, tissues as well as larger samples like insects. Conventional SEM gives a magnification ranging from 20X to 30,000X and spatial resolution of 50 to 100 nanometers.
Fundamental Principles
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Overview of Electron Microscopy

Overview of Electron Microscopy

The wavelengths of visible light ultimately limit the maximum theoretical resolution of images created by light microscopes. Most light microscopes can only magnify 1000X, and a few can magnify up to 1500X. Electrons, like electromagnetic radiation, can behave like waves, but with wavelengths of 0.005 nm, they produce significantly greater resolution up to 0.05 nm as compared to 500 nm for visible light. An electron microscope (EM) can create a sharp image that is magnified up to 2,000,000X.

Transmission Electron Microscopy

Transmission Electron Microscopy

In 1931, physicist Ernst Ruska—building on the idea that magnetic fields can direct an electron beam just as lenses can direct a beam of light in an optical microscope—developed the first prototype of the electron microscope. This development led to the development of the field of electron microscopy. In the transmission electron microscope (TEM), electrons are produced by a hot tungsten element and accelerated by a potential difference in an electron gun, which gives them up to 400...

Immunogold Electron Microscopy

Immunogold Electron Microscopy

Immunoelectron microscopy utilizes immunogold labeling of endogenous proteins with specific antibodies to detect and localize these proteins in cells and tissues. The procedure provides insights into the distribution and quantification of protein under different stimulation conditions offering clues about their functions. Conjugating highly electron-dense gold particles with primary or secondary antibodies allow antigen detection on and within cells, with high resolution and specificity.

Preparation of Samples for Electron Microscopy

Preparation of Samples for Electron Microscopy

To be visualized by an electron microscope, either transmission or scanning, biological samples need to be fixed (stabilized) so the electron beam does not destroy them and dried thoroughly (desiccated/dehydrated) so the vacuum does not affect them. Fixation needs to be done as quickly as possible because the sample properties will start changing as soon as it is removed from its natural environment. For example, in a tissue sample, the oxygen levels begin decreasing, causing an altered...

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Related Experiment Video

Updated: Jan 28, 2026

Micropatterning Transmission Electron Microscopy Grids to Direct Cell Positioning within Whole-Cell Cryo-Electron Tomography Workflows

Micropatterning Transmission Electron Microscopy Grids to Direct Cell Positioning within Whole-Cell Cryo-Electron Tomography Workflows

Published on: September 13, 2021

An improved specimen loader for cryo-scanning electron microscopy.

Rajgopal Iyer¹, M A Arunagirinathan, C S Prabhu

¹Department of Chemical Engineering, IIT Bombay, Powai, Mumbai, India.

|June 7, 2005

Summary

A new specimen loader for cryo-electron microscopy (cryo-EM) simplifies sample handling. This innovation reduces inadvertent exposure to atmospheric moisture, minimizing frost deposition on cryofixed samples.

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Cryo-electron Microscopy Specimen Preparation By Means Of a Focused Ion Beam

Cryo-electron Microscopy Specimen Preparation By Means Of a Focused Ion Beam

Published on: July 26, 2014

Area of Science:

* Structural Biology
* Microscopy Techniques

Background:

* Cryo-electron microscopy (cryo-EM) is crucial for visualizing hydrated biological samples.
* Specimen handling during cryo-EM workflow, particularly loading onto sample holders, is prone to atmospheric moisture exposure.
* This moisture exposure leads to frost deposition, compromising sample integrity and image quality.

Purpose of the Study:

* To develop an improved specimen loader for cryo-EM.
* To streamline the process of loading specimens onto sample holders.
* To minimize frost formation during sample handling.

Main Methods:

* Design and implementation of a novel specimen loader with modified components.
* Key modifications include a movable liquid freon cup and an improved stage holder shape.
* Elimination of the need for a lock-screw to simplify the mounting procedure.

Main Results:

* The new loader facilitates easier and smoother specimen loading.
* Reduced handling of the specimen holder significantly lowers exposure to atmospheric moisture.
* Consequent reduction in frost deposition on cryofixed samples was observed.

Conclusions:

* The redesigned specimen loader enhances the efficiency and reliability of cryo-EM sample preparation.
* Minimizing frost deposition is critical for obtaining high-resolution cryo-EM data.
* This improved handling technique supports better structural biology research through cryo-EM.