A Simpler and Faster Version of Two-Dimensional Gel Electrophoresis Using Vertical, Mini SDS-PAGE Apparatus

Abstract

We have modified one of the most useful methods of protein separation; namely, two dimensional gel electrophoresis (2-DE). This modified version of 2-DE is not only simpler and easier but also faster than all the currently available methods. In this method, isoelectric focusing is carried out in the first dimension using a vertical sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) apparatus. Following focusing, each individual lane is excised from this gel and after a 90° rotation, is inserted into a vertical specially fabricated SDS-PAGE gel for the second dimension run, which can be stained with Coomassie Brilliant Blue for protein analysis or immunoblotted for further analysis. This modified version of IEF can be run in less than 2 hours compared to the overnight run required by the O’Farrel method. Difficult tube gel casting and gel extrusion as well as tube gel distortion are eliminated in our method. This method is simpler, faster and inexpensive. Both dimensions can be done on the same SDS-PAGE apparatus and up to 10 samples can be run simultaneously using one gel. We have analyzed human sera, animal saliva, eye and liver tissue samples to verify this method.

Keywords

• 2d gel electrophoresis
• isoelectric focusing
• sodium dodecyl sulfate polyacrylamide
• gel electrophoresis