Title: Learn: Stacking in SDS-PAGE
Course: BTEC-2020-301-302-303-F21 > Section: Protein Gels

Module2 Stacking in SDS-PAGE

This video shows a pre-stained protein ladder being run on an SDS-PAGE gel. As the proteins move through the top layer of gel, called the stacking gel, they are pressed into a tight band.

Stacking - The compression of a sample in an electrophoresis well into a tight band; in SDS-PAGE, the top layer of gel is called the stacking gel.

The stacking gel’s pores are very large so very little separation by mass occurs here. Because of stacking, the proteins all hit the “starting line” of the separating gel at the same time. As they move through the more dense matrix of the separating gel, the proteins are separated by mass, with smaller proteins moving through the gel more rapidly than large ones. What happens in the separating gel is analogous to DNA separation by size in agarose gel electrophoresis.

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The goal of SDS-PAGE is generally to obtain information about the molecular mass of a protein, by comparing its migration to that of known standards (the ladder). Stacking helps ensure bands are sharp, rather than smeared, so that a more accurate mass can be determined.