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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.

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...

Immunofluorescence Microscopy

Immunofluorescence Microscopy

A fluorescence microscope uses fluorescent chromophores called fluorochromes, which can absorb energy from a light source and then emit this energy as visible light. Fluorochromes include naturally fluorescent substances (such as chlorophylls) and fluorescent stains that are added to the specimen to create contrast. Dyes such as Texas red and FITC are examples of fluorochromes. Other examples include the nucleic acid dyes 4’,6’-diamidino-2-phenylindole (DAPI), and acridine orange.

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.

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Related Experiment Video

Updated: Apr 29, 2026

4D Microscopy of Yeast

4D Microscopy of Yeast

Published on: April 28, 2019

Imaging yeast NPCs: from classical electron microscopy to Immuno-SEM.

Elena Kiseleva¹, A Christine Richardson², Jindriska Fiserova²

¹Laboratory of Morphology and Function of Cell Structure, Institute of Cytology and Genetics, Russian Academy of Science, Novosibirsk, Russia.

Methods in Cell Biology

|May 27, 2014

Summary

We optimized electron microscopy (EM) protocols for yeast Saccharomyces cerevisiae, overcoming challenges like cell walls and vacuoles. This allows detailed study of nuclear pore complex (NPC) structure and protein localization.

Keywords:

Electron microscopy Immunogold labeling Sacharomyces cerevisiae Scanning Transmission Yeast

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Area of Science:

Cell Biology
Microscopy Techniques

Background:

Electron microscopy (EM) is crucial for studying nuclear transport and the nuclear pore complex (NPC).
Yeast (Saccharomyces cerevisiae) presents unique challenges for EM due to its cell wall, vacuoles, and dense cytoplasm.
Existing EM methods struggle with yeast sample preparation, impacting fixation, embedding, and imaging.

Purpose of the Study:

To present optimized protocols for Transmission EM (TEM) and Scanning EM (SEM) in yeast.
To overcome challenges in preparing yeast samples for ultrastructural analysis.
To enable detailed investigation of yeast NPC structure and nucleoporin/transport factor localization.

Main Methods:

Developed specific sample preparation protocols for TEM and SEM in yeast.
Utilized immunogold labeling for visualizing proteins within the NPC ultrastructure.
Focused on overcoming fixation, embedding, and imaging difficulties specific to yeast.

Main Results:

Successfully prepared yeast samples for high-resolution TEM and SEM.
Achieved detailed ultrastructural information of the yeast NPC.
Localized nucleoporins and transport factors within the yeast NPC structure using immunogold labeling.

Conclusions:

Optimized EM protocols facilitate detailed ultrastructural analysis of yeast NPCs.
These methods enable precise localization of key proteins involved in nuclear transport.
The presented protocols advance the study of nuclear transport mechanisms in Saccharomyces cerevisiae.