Structural Biochemistry/Proteins/Purification/Gel Electrophoresis

Gel Electrophoresis
Gel electrophoresis a method of qualitatively separating proteins by charge or size. It is also usable with DNA or RNA, only it separates the molecules based on length as opposed to molecular weight. These gels are placed loaded into a box that passes an electric current over the samples in solutions and forces them to migrate the negatively charged molecules from one end of the gel to another. While the pore sizes differ between to two main types of gels, agarose and polyacrylamide, the lighter the protein the faster it runs on the gel. The result of an electrophoresis gel is a sorted out list of proteins found in the sample arranged by size.

The main types of gels used in electrophoresis are Agarose (proteins larger than 200 kDa) and polyacrylamide (proteins ranging from 5 to 2000 kDa).

The two types of conditions a gel can be run at is either an SDS denaturing gel, where the native structure is not maintained, the protein is unfolded and denatured, and the protein becomes sorted solely on its molecular weight. The other condition of gel is Native, which maintains the proteins tertiary structure.

Gels not only allow a scientist to track and qualitatively evaluate a desired protein, but it also helps visualize how pure the protein of desire is in solution. Depending on the gel and the specific conditions made, a trace amount of protein can be picked up by a gel.

Picture above is an example of a typical SDS-gel. The bar on the far left is known as a benchmark ladder (molecular marker) that marks different molecular weights allow comparison to the samples run. The darker the band, the higher the concentration of the protein in that given sample.