Analytical Separations

At various stages, but more particularly at the end, one needs to know how close to purity the protein fraction is. Of many different analytical techniques that have been used, one is universally employed. This is known as SDS-PAGE, which stands for sodium dodecyl sulfate-polyacrylamide gel elec-trophoresis. The gel (polyacrylamide) has pores that are similar in size to the protein molecules, and the mixture of proteins is forced through the gel by an electric field. First the proteins are fully denatured by treatment with the anionic detergent sodium dodecyl sulfate, which binds to all the proteins and confers on them a negative charge proportional to their size. The smaller proteins move through the gel relatively easily, but the larger ones are retarded by the lack of pores large enough to move through. Consequently there is a separation of components according to size, in a logarithmic way similar to that observed for gel filtration (see Figure 3). The individual components remain in tight bands; as a consequence proteins that differ in size by only 3-5% can be completely separated. This sharp resolution is thus able to detect small amounts of impurity, and a judgement can be made on the absolute purity of the sample. One further feature is that, as with gel filtration, the distance moved by an unknown protein can be compared with known standards and the unknown molecular weight determined to within about 5% accuracy. The amount of protein required is only about 1 |g or less, depending on the detection (staining) method. Capillary electrophoresis systems can be even more sensitive and accurate for analytical work.

See also: II/Affinity Separation. Theory and Development of Affinity Chromatography. Chromatography: Protein Separation. Chromatography: Liquid: Column Technology; Large-Scale Liquid Chromatography; Mechanisms: Ion Chromatography. Electrophoresis: One-dimensional Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis; Proteins, Detection of; Two-dimensional Polyacrylamide Gel Electrophoresis.

Solar Panel Basics

Solar Panel Basics

Global warming is a huge problem which will significantly affect every country in the world. Many people all over the world are trying to do whatever they can to help combat the effects of global warming. One of the ways that people can fight global warming is to reduce their dependence on non-renewable energy sources like oil and petroleum based products.

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