Gel Electrophoresis

Electrophoretic separations can also be successfully carried out using an appropriate gel that usually takes the form of a cross-linked polymer. This technique, called gel electrophoresis, is widely used in microbiology, biochemistry, genetic studies and for forensic purposes. The composition and porosity of the gel is arranged by modification of the synthetic process (by choosing different concentrations of acrylamide and cross-linking agent) to compliment the composition and molecular weight of the substances to be separated. The polymer must take the form of a solid yet be appropriately porous to permit electrophoretic migration. If the molecules to be separated are relatively small such as, proteins or small nucleic acid (e.g. deoxyribonucleic acid, ribonucleic acid and oligonucleotides) the gel is usually made from different concentrations suitably cross-linked polyacrylimide. However, if the size of the molecules to be separated is much larger, then a form of pure agarose might be more suitable. Agarose is a carbohydrate with long side chains that can be made to produce a gel with very large pores that will permit the migration of very large molecules. 

When completed the separation can be examined using a number of different procedures. If the components are radioactive, the gel plate can be scanned and counted; if some or all of the components fluoresce, then the plate can be scanned for fluorescence. Alternatively the materials separated can be stained with suitable reagents such as ethidium bromide, silver, or some other appropriate dye. If samples are placed next to each other they will travel down individual lanes each component appearing at a specific position in its respective lane. The position of the spot indicates the nature of the substance and the intensity of the spot allows quantitative assessment. An example of one two such separations are shown in figure 12.

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Figure 12 The Separation of Mixture Containing Proteins on a Gel Plate.

Gel electrophoresis can be carried out in the two dimensional mode but before this technique id discuss the isoelectric focussing processes needs to be discussed.