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Electrophoresis, Gel, Two-Dimensional: Electrophoresis in which a second perpendicular electrophoretic transport is performed on the separate components resulting from the first electrophoresis. This technique is usually performed on polyacrylamide gels.
 JoVE Immunology and Infection

Total Protein Extraction and 2-D Gel Electrophoresis Methods for Burkholderia Species

1Department of Pediatrics, Centre for Understanding and Preventing Infection in Children, University of British Columbia


JoVE 50730

Members of the Burkholderia genus are pathogens of clinical importance. We describe a method for total bacterial protein extraction, using mechanical disruption, and 2-D gel electrophoresis for subsequent proteomic analysis.

 JoVE Biology

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

1Research and Development, GE Healthcare Bio-Sciences AB


JoVE 945

A simple and specific method was demonstrated for fluorescent labeling and enhanced detection of cell surface proteins without a fractionation step. Differential abundance in cell surface proteins was analyzed using two-dimensional (2-D) electrophoresis and Ettan™ DIGE technology.

 JoVE Biology

Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels

1Biological Medical Research Center (BMFZ), University Duesseldorf


JoVE 1431

This video shall popularize a colloidal Coomassie G-250 staining protocol according to Kang et al. for the detection of average 4 ng protein in gels. The staining is completed within 2 hours and without any effort. We routinely use Kang's protocol for analytical purposes in gel-based proteomics.

 JoVE Medicine

Skeletal Muscle Gender Dimorphism from Proteomics

1Center for Proteomics, Smith College, 2Department of Molecular Biophysics and Biochemistry, Yale University, 3Department of Chemistry, Smith College, 4Department of Biological Sciences and Center for Proteomics, Smith College


JoVE 3536

A straight-forward set of methods to isolate and determine the identity of the most abundant proteins expressed in skeletal muscle. About 800 spots are discerned on a two-dimensional gel from 10 mg muscle; this allows for the determination of gender-specific protein expression. These methods will give equivalent results in most tissues.

 JoVE Immunology and Infection

Purification of Pathogen Vacuoles from Legionella-infected Phagocytes

1Max von Pettenkofer-Institut, Ludwig-Maximilians-Universität


JoVE 4118

This article describes a method for the isolation and purification of intact Legionella-containing vacuoles (LCVs) from amoeba and macrophages. The two-step protocol comprises LCV enrichment by immuno-magnetic separation using an antibody against a bacterial LCV marker and further purification by density gradient centrifugation.

 Science Education: Basic Methods in Cellular and Molecular Biology

Separating Protein with SDS-PAGE

JoVE Science Education

Sodium Dodecyl Sulfate Poly-Acrylamide Gel Electrophoresis , or SDS-PAGE, is a widely-used technique for separating mixtures of proteins based on their size and nothing else. SDS, an anionic detergent, is used to produce an even charge across the length of proteins that have been linearized. By first loading them into a gel made of polyacrylamide and then applying an electric field to the gel., SDS-coated proteins are then separated. The electric field acts as the driving force, drawing the SDS coated proteins towards the anode with larger proteins moving more slowly than small proteins. In order to identify proteins by size, protein standards of a known size are loaded along with samples and run under the same conditions. This video presents an introduction to SDS-PAGE by first explaining the theory behind it and later demonstrating its step-by-step procedure. Various experimental parameters, such as the polyacrylamide concentration and voltage applied to the gel are discussed. Downstream staining methods like Coomassie and silver stains are introduced, and variations of the method, like 2D gel electrophoresis are presented.

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