Electrophoresis and PAGE

Question

What is Electrophoresis? And how is PAGE used in protein separation?

Answer

When molecules, such as proteins, encounter an electric field, their polarised nature results in a net migration towards an electrode of opposite charge. This phenomenon is known as electrophoresis and can be utilised in a number of lab based protocol, including Polyacrylamide Gel Electrophoresis (PAGE) (Hames and Hooper, 2006). Poly Acrylamide Gel electrophoresis (PAGE) utilises a polyacrylamide gel to electrophoretically separate proteins based on their molecular weight. The gel acts as a sieve which, based on the variability of pore sizes within the gel, separates proteins as they migrate through the medium towards the electrode of opposite charge. A protein colour marker, such as Bromophenol blue, is added to protein samples in order to visualise the migration of the molecule through the gel. A molecular weight marker is generally added to the first lane of any gel. This marker typically contains a purified protein of known molecular weight (measured in kilo Daltons (kDa)). These markers are ran alongside the investigated protein samples in order to draw comparison and enable the quantification of the protein samples under investigation, based on the molecular weight of the molecules separation, in relation to the known values within the marker (Spurway and Wackerhage, 2006).

References

Hames, D. Hooper, N. (2006) Instant Notes: Biochemistry, 3rd Edition, Abingdon: Taylor and Francis group.

Spurway, N. Wackerhage, H. (2006) Genetics and Molecular Biology of Muscle Adaptation, London: Elsevier Limited.