Continuous Elution SDS-PAGE with a Modified Standard Gel Apparatus to Separate and Isolate an Array of Proteins from Complex Mixtures.

Abstract

Sodium dodecyl sulfate polyacrylamide gel electrophoresis is a powerful tool to separate proteins according to their relative sizes. The technique provides information about both the size of a number of proteins and potentially the comparative amounts of each protein. To confirm the identity of proteins, proteins can be eluted from the gel and transferred electrophoretically to nitrocellulose for antibody-based detection. During electrophoresis, if the current is not stopped, proteins continue to pass down the gel and elute from the bottom of the gel. The standard electrophoresis gel apparatus can be employed with the addition of some tubing and alterations to the separating gel to collect proteins separated by size as they elute from the base of the gel as described in this chapter. Complex protein mixtures can be separated into multiple fractions containing single proteins in a few hours. Small amounts (<500μg of protein) of protein sample can be fractionated.