Preparative Electrophoresis Using Polyacrylamide Gels

Gels can be synthesized that have pore sizes of the same order of magnitude as protein molecules. Such gels were originally formed from starch but more recently from polyacrylamide. These high-density gels readily permit the passage of small molecules but selectively retard larger molecules according to their dimensions. This process is a form of molecular sieving as employed in exclusion chromatography. Original work on this gel media was carried out using specially hydrated starch carrying between 12 and 16% of the hydrated starch. Subsequently, polyacrylamide was found more effective and had clarity and complete transparency. Unfortunately, such gels have very limited stability and, therefore must be prepared as required. The concentration of acrylamide in the gel can vary widely from 5 to 35%. Cross-linking is achieved by the use of peroxides and is activated photochemically. The gels can be formed in vertical troughs but more frequently in glass tubes, each tube containing a single sample. The separated bands of the different proteins appear as stacked discs and as a consequence this technique is sometimes referred to as disc electrophoresis.