Detection of Enzymes

It is generally considered that specific enzyme activities can only be visualized following gel electrophor-esis if native conditions have been used. However, there are several reports demonstrating that SDS-de-natured proteins can also be visualized provided that it is possible to achieve at least partial renaturation of their spatial configuration. Such renaturation is most effective if disulfide bonds are not essential for en-zymic activity and if the native protein is not composed of subunits of different molecular weights. Pre-electrophoresis of gels is usually recommended to remove unreacted acrylamide monomers and catalysts.

Enzyme staining can be achieved by incubating the unfixed gel in a solution of the appropriate reagents using either fluorogenic or chromogenic substrates. This method works well if the final reaction product is insoluble. However, a soluble reaction product will rapidly diffuse resulting in loss of resolution. It is generally preferable to use a print or gel overlay technique. In this approach, the substrates and other reagents are either impregnated into a filter or included in a thin layer of agarose or polyacrylamide gel cast on a glass or plastic support. The overlay is then placed in direct contact with the surface of the separation gel and following a suitable period of incubation, the enzymic activity is visualized on the overlay. Methods are available for the visualization of a large number of enzyme activities following gel electrophoresis.

See also: Ill/Proteins: Capillary Electrophoresis; Electrophoresis. Electrophoresis: Detection Techniques: Staining; Autoradiography and Blotting

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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