Posts tagged ‘SDS-PAGE’
SDS-PAGE stands for sodium dodecyl (lauryl) sulfate-polyacrylamide gel electrophoresis. It sepearates proteins accroding to their sizes. Therefore, this technique can be used to identify and quantify proteins and to check sample purity.
SDS is a detergent that has a negative charge (from the sulfate) attached to it. It will denature protein samples, leaving the protein in its primary structure, and give it negative charges. This ensure that the protein samples migrate through the gel based on their size alone, not other properites like secondary, tertiary or quarternary structures or electric charges.
Polyacrylamide is a polymer consisting of acrylamide monomers. It forms a gel with “tunnels” of different sizes. When the protein samples are loaded into the gel and placed in an electric field, they will move toward the positive charge (Recall that the SDS has given the proteins negative charges). The smaller proteins will make their way through the gel faster, so the proteins samples are separated based on their sizes. The samples contains many copies of the same protein. They could also be a mixture of proteins, but the proteins with the same size always migrate to the same position in a defined period of time.
The intensity of the bands depends on the amount of protein in the samples. To see individual bands clearly, only a small quantity is required. While running the gel, the proteins are colourless and invisible. The gel needs to be stained to see bands on the gel. Usually, the gel is never run so long that the bands totally migrate to the other side.
This fact that SDS-PAGE only separates proteins according to their sizes deserves a caveat. Different proteins with the same size will move to the same position and end up in the same band. In this case, individual proteins can not be identified.