2D gel electrophoresis

2016-10-12T20:01:08Z

NickTest: Corrected a reference

It is a powerful method of separating [[Protein|protein]] along a tubular gel using [[Isoelectric focusing|iso-electric focusing]] and [[SDS polyacrylamide-gel electrophoresis|sodium dodecyl sulphate polyacrylamide gel]]. The sample undergoes [[Isoelectric focusing|iso-electric focusing]] first; an electrical current is passed through a gel with a [[PH gradient|pH gradient]], and the proteins stop moving when they reach the pH at which they have no net charge - this is known as the [[Isoelectric point|Iso-electric point]] (pI). The sample is then placed horizontally on top of [[SDS-PAGE|SDS-PAGE]] and spreads across. The sample then moves vertically down again to yield the second dimension based on the size of the protein, producing a 2D pattern. So the sample has been separated by virtue of [[Isoelectric point|Iso-electric point]] (pI), when they move horizontally and by virtue of their size; when they move down vertically. 

Because of the specificity of this method, it is possible to identify over one thousand proteins on the same gel, in one experiment <ref>Stryer, Lubert; Tymoczko, John L.; Berg, Jeremy M., (2012) Biochemistry, 7th Edition, New York: WH Freeman and Co. Page 74</ref>.                                                                        

The concentration of isolated proteins can be determined by measuring the intensity of the spots on the gel. Comparing this to other cells can show differences due to differing physiological conditions <ref>Two- dimensional electrophoresis: (Page 76) Berg J, Tymoczko J, Stryer L. Biochemistry. Seventh Edition. New York: W.H. Freeman; 2012.</ref>.

This method can be used to separate DNA fragments because the phosphate backbone of DNA has a negative charge. The distance a DNA fragment travels isinversely proportional to the log of its molecular weight.

=== Reference ===

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2D gel electrophoresis