immunoblotting Protocols

Category: Protein Protocols: Western Blot Protocol

Nice Western Blotting Protocol using some eBioscience Products. - [Read Immunoblotting Using eBioscience Products]

Category: Western Blot Protocols: Antigen Detection Methods Protocols

The blot is blocked to prevent nonspecific adsorption of the immunological reagents.  Antibodies are then bound to the proteins immobilized on the membrane, and the antigen is detected by labeling the antibodies with conveniently identified tags.  Common labeling methods for chemiluminescent detection include anti-immunoglobulin antibody-coupled enzymes such as horseradish peroxidase, which catalyzes the oxidation of luminol and in turn releases light. - [Read Immunoblotting: Antigen Detection Using Chemiluminescence Protocol]

Category: Western Blot Protocols: Antigen Detection Methods Protocols

The blot is blocked to prevent nonspecific adsorption of the immunological reagents.  Antibodies are then bound to the proteins immobilized on the membrane, and the antigen is detected by labeling the antibodies with conveniently identified tags. - [Read Immunoblotting: Antigen Detection Using Chromogenic Methods Protocol]

Category: Western Blot Protocols: General Western Blot Methods Protocols

For immunoblotting experiments, it is often important to compare the total amount of an antigen from many different sources or to learn if a particular source has the antigen under study.  In the approach described here, tissue cultures, bacteria, yeast cells, tissues, and other sources of antigens are disrupted directly in an electrophoresis sample . - [Read Immunoblotting: Preparing Cell Lysates Protocol]

Category: Western Blot Protocols

Protein immunoprecipitation can be a useful preparative step for immunoblotting.  For very rare proteins, the protein of interest can be purified and concentrated by standard immunoprecipitation techniques before immunoblotting.  In addition, protein-protein interactions can be tested with an immunoprecipitating antibody that is specific for one protein of a complex and an immunoblotting antibody that is specific for a second member of the complex. - [Read Immunoblotting: Preparing Immunoprecipitated Proteins Protocol]

Category: Western Blot Protocols: General Western Blot Methods Protocols

Protocol describes how proteins are mixed with a sample buffer and prepared for electrophoresis on standard Tris/glycine SDS-polyacrylamide gels. - [Read Immunoblotting: Preparing Protein Solutions Protocol]

Category: Western Blot Protocols: General Western Blot Methods Protocols

The transfer of proteins from a Tris/glycine SDS-polyacrylamide gel to a membrane using a semi-dry method is achieved by placing the gel next to a piece of nitrocellulose filter.  This sandwich is placed directly between two plate electrodes, and the proteins are then transferred from the gel onto the filter. - [Read Immunoblotting: Semi-Dry Electrophoretic Transfer of Proteins from Gels to Membranes Protocol]

Category: Western Blot Protocols: General Western Blot Methods Protocols

Transfer of proteins from a Tris/glycine SDS-polyacrylamide gel to a membrane using the submerged method is achieved by placing the gel next to a piece of nitrocellulose filter, submerging this sandwich in a large volume of transfer buffer in a transfer tank, and running current from one side of the transfer tank to another.  The proteins are then eluted by transferring them from the gel onto the filter. - [Read Immunoblotting: Submerged Electrophoretic Transfer of Proteins from Gels to Membranes Protocol]

Category: Transfection Protocols: Stable Transfection Protocols

Stably transfected cells, generated in the first two stages of the procedure, are induced for expression of the target gene. After harvesting and lysis, the lysates are analyzed by SDS-PAGE and immunoblotting. - [Read Tetracycline as Regulator of Inducible Gene Expression III]

Category: Immunology: Immunoassays

3-step process of Western immunoblotting for detection of particular sites of phosphorylation on specific proteins recognized by modification-sensitive antibodies. - [Read Western Blot Analysis—Phosphoprotein-Specific Antibody Mixture Protocol]

Category: Protein Protocols: Western Blot Protocol

Theory and Protocol for Western Blotting / immunoblotting. R&D Systems. - [Read Western Blot Protocol and Theory]