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Related Concept Videos

Two-dimensional Gel Electrophoresis01:22

Two-dimensional Gel Electrophoresis

Two-dimensional gel electrophoresis is a high-resolution protein separation method first introduced by O' Farrell and Klose in 1975. This method involves protein separation by two dimensions, mass and charge, making it more accurate than one-dimensional gel electrophoresis.
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as  cells...
DNA Agarose Gel Electrophoresis02:35

DNA Agarose Gel Electrophoresis

Agarose gel electrophoresis is a laboratory technique commonly used to separate DNA fragments by size. However, it can also be used to isolate and purify DNA fragments using a gel extraction protocol.
Gel extraction follows five major steps: running gel electrophoresis to separate fragments, isolating the individual bands, extracting DNA from those bands, and removing the dye and salts from the extracted mixture to obtain pure DNA.
In cloning experiments, both the insert and vector DNA...
Tagging and Fusion Proteins01:24

Tagging and Fusion Proteins

Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...
Capillary Electrophoresis: Applications01:30

Capillary Electrophoresis: Applications

Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
SDS-PAGE01:27

SDS-PAGE

Gel electrophoresis is a method that separates biological macromolecules like nucleic acids or proteins by forcing them to pass through a gel matrix under an electric field.
A variation of gel electrophoresis, termed  polyacrylamide gel electrophoresis (PAGE), is commonly used for separating proteins according to their molecular size by passing them through a polyacrylamide gel. Because of the varying charges associated with amino acid side chains, PAGE can be used to separate intact proteins...

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Related Experiment Video

Updated: Jul 6, 2026

Selective Labelling of Cell-surface Proteins using CyDye DIGE Fluor Minimal Dyes
14:43

Selective Labelling of Cell-surface Proteins using CyDye DIGE Fluor Minimal Dyes

Published on: November 26, 2008

Difference gel electrophoresis based on lys/cys tagging.

Reiner Westermeier1, Burghardt Scheibe

  • 1GE Healthcare Europe GmbH, Munich, Germany.

Methods in Molecular Biology (Clifton, N.J.)
|March 29, 2008
PubMed
Summary

This study introduces CyDye DIGE Fluors for multiplexed protein analysis, enabling sensitive detection of protein expression changes with an internal standard for improved reproducibility.

Area of Science:

  • Proteomics
  • Biochemistry
  • Analytical Chemistry

Background:

  • Differential fluorescent labeling of proteins is crucial for comparative proteomics.
  • Existing methods face limitations in sensitivity and reproducibility for complex samples.

Purpose of the Study:

  • To present CyDye DIGE Fluors as a multiplexing technique for enhanced protein expression analysis.
  • To demonstrate the reliability and reproducibility of this method using an internal standard.

Main Methods:

  • Proteins are labeled with spectrally distinct CyDye DIGE Fluors targeting lysine or cysteine residues.
  • Minimal labeling of lysine residues offers sensitivity comparable to silver staining.
  • Cysteine labeling provides high sensitivity but alters protein migration patterns.

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Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A (EYFP-CENP-A)
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Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A (EYFP-CENP-A)

Published on: June 10, 2020

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides
09:58

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides

Published on: November 29, 2016

Related Experiment Videos

Last Updated: Jul 6, 2026

Selective Labelling of Cell-surface Proteins using CyDye DIGE Fluor Minimal Dyes
14:43

Selective Labelling of Cell-surface Proteins using CyDye DIGE Fluor Minimal Dyes

Published on: November 26, 2008

Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A (EYFP-CENP-A)
09:02

Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A (EYFP-CENP-A)

Published on: June 10, 2020

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides
09:58

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides

Published on: November 29, 2016

  • Labeled samples are co-migrated in 2D gels and analyzed using a multifluorescent imager.
  • An internal standard is created by labeling a pooled aliquot of all samples.

    • Main Results:

    • Multiplexing allows simultaneous analysis of up to three samples per gel.
    • Minimal labeling preserves protein spot patterns similar to post-stained gels.
    • Cysteine labeling significantly enhances detection sensitivity.
    • The internal standard facilitates reproducible quantification of protein expression changes.
    • Specialized software enables co-detection and comparison of protein spots across samples.

    Conclusions:

    • CyDye DIGE Fluors enable sensitive and reproducible multiplexed protein expression profiling.
    • The minimal labeling approach balances sensitivity with pattern integrity.
    • The internal standard strategy enhances the reliability of quantitative proteomics.