Electroblotting from Polyacrylamide Gels

Visualization of acid phosphatase activity on nitrocellulose filters following electroblotting of polyacrylamide gels.

A method for visualizing acid phosphatase isoenzymes by activity staining on nitrocellulose filters after electroblotting of proteins fractionated on nondenaturing polyacrylamide gels is described. Reproducible results were obtained when 25 mM Tris-192 mM glycine was used as the transfer buffer instead of 0.7% acetic acid, 50 mM sodium acetate, pH 4, or 0.14 M acetic acid--0.35 M beta-alanine, ...

Electroblotting onto activated glass. High efficiency preparation of proteins from analytical sodium dodecyl sulfate-polyacrylamide gels for direct sequence analysis.

We have developed a new method for the isolation of proteins for microsequencing. It consists of electrophoretic transfer (electroblotting) of proteins or their cleavage fragments onto activated glass filter paper sheets immediately after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The proteins are immobilized on the glass fiber sheets by ionic interactions or by co...

Western blot of stained proteins from dried polyacrylamide gels.

Removal of coomassie blue precipitates from polyacrylamide gels.

During staining of polyacrylamide gels, varying amounts of Coomassie blue may precipitate on the surface of the gels. This background obscures faint bands and is esthetically unappealing. The problem is worse when gels are stained for longer periods of time or when the stain has not been recently filtered. Rinsing a gel with methanol efficiently and quickly removes this background. Basically, a...

Protein purification from polyacrylamide gels by sonication extraction.

A protein purification procedure using sonication extraction from polyacrylamide gels (PAGE), which involves identification of a particular protein band and its excision, homogenization, sonication, and subsequent passage through a Sephadex G-25 minicolumn, is reported. Our results show a high degree of recovery regardless of the nature of the protein (soluble or membrane bound) or the characte...