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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...
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...
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...
Electrophoresis: Overview01:20

Electrophoresis: Overview

Electrophoresis is a powerful analytical separation technique that relies on the differential migration of charged species when subjected to an electric field. The core strength of electrophoresis lies in its ability to separate high-molecular-weight species in complex mixtures. It has found widespread use in biochemistry, molecular biology, and analytical chemistry, allowing the separation of compounds like amino acids, nucleotides, carbohydrates, and proteins with excellent resolution.
There...
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,...
Southern Blot02:57

Southern Blot

Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments.
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...

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Related Experiment Video

Updated: Jul 7, 2026

Two-dimensional Gel Electrophoresis Coupled with Mass Spectrometry Methods for an Analysis of Human Pituitary Adenoma Tissue Proteome
12:34

Two-dimensional Gel Electrophoresis Coupled with Mass Spectrometry Methods for an Analysis of Human Pituitary Adenoma Tissue Proteome

Published on: April 2, 2018

Two-dimensional gel electrophoresis.

S Harper1, J Mozdzanowski, D Speicher

  • 1The Wistar Institute, Philadelphia, Pennsylvania, USA.

Current Protocols in Cell Biology
|January 30, 2008
PubMed
Summary

Two-dimensional gel electrophoresis (2D-PAGE) separates proteins by charge and size, resolving thousands of proteins. This unit details methods for characterizing cell lysates using 2D-PAGE, including sample preparation and database utilization.

Area of Science:

  • Proteomics
  • Biochemistry
  • Molecular Biology

Background:

  • One-dimensional SDS-PAGE separates proteins solely by molecular weight.
  • Two-dimensional gel electrophoresis (2D-PAGE) offers enhanced protein resolution by separating proteins based on isoelectric point (charge) in the first dimension and molecular weight (size) in the second dimension.
  • Sensitive detection methods enable 2D-PAGE to resolve 1000 to 2000 distinct proteins.

Purpose of the Study:

  • To describe comprehensive methods for characterizing cell lysates using 2D-PAGE.
  • To provide protocols for various modifications and sample preparations for enhanced protein separation and analysis.
  • To guide researchers in utilizing 2D protein databases for data interpretation.

Main Methods:

  • Two-dimensional gel electrophoresis (2D-PAGE) involving isoelectric focusing (IEF) followed by SDS-PAGE.

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Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers
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Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers

Published on: April 26, 2018

Agarose Gel Electrophoresis for the Separation of DNA Fragments
07:10

Agarose Gel Electrophoresis for the Separation of DNA Fragments

Published on: April 20, 2012

Related Experiment Videos

Last Updated: Jul 7, 2026

Two-dimensional Gel Electrophoresis Coupled with Mass Spectrometry Methods for an Analysis of Human Pituitary Adenoma Tissue Proteome
12:34

Two-dimensional Gel Electrophoresis Coupled with Mass Spectrometry Methods for an Analysis of Human Pituitary Adenoma Tissue Proteome

Published on: April 2, 2018

Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers
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Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers

Published on: April 26, 2018

Agarose Gel Electrophoresis for the Separation of DNA Fragments
07:10

Agarose Gel Electrophoresis for the Separation of DNA Fragments

Published on: April 20, 2012

  • Modifications for analyzing acidic and basic proteins.
  • Utilizing immobilized pH gradients (IPGs) for precise first-dimension separation.
  • Performing nonreducing and reducing electrophoretic separations.
  • Support protocols for gel casting, sample preparation, molecular weight standard preparation, and pH profiling.

    • Main Results:

    • Detailed methodologies for performing 2D-PAGE on cell lysates are presented.
    • Optimized protocols for handling diverse protein types (acidic, basic) and conditions (reducing/nonreducing) are described.
    • The capability of resolving 1000-2000 proteins with sensitive detection is highlighted.

    Conclusions:

    • 2D-PAGE is a powerful technique for high-resolution protein separation and characterization of complex biological samples like cell lysates.
    • The described methods and protocols facilitate robust and versatile proteomic analysis.
    • Integration with protein databases aids in the identification and interpretation of separated protein spots.