Conventional Immunoelectrophoresis

The scheme in Figure 1A shows the principles of the techniques, such as it was originally introduced and generally used by many workers. The first step consists of electrophoresis in 1% agar-agar or agarose gels, prepared in different buffers at pH ranging from 8.2 to 8.6 (Veronal 0.025 molL"1, pH 8.2 is one of the buffers frequently used). The sample to be analysed, usually a complex mixture of proteins, is applied in small wells located in the middle of agar-agar gels, or partially displaced towards the extreme nearest to the cathode when agarose gels are used. Most of the proteins, for example in blood sera or plasma, have a negative net charge in buffers with pH higher than 7.0. Thus, when a continuous electrical field is applied to the gel, the tendency of these proteins is to move towards the anode with a migration rate which mainly depends on charge-to-size ratio. However, the migration rate is in part reduced by the electroendosmotic effect due to the negative charges in polymeric molecules of the gel. This elec-troendosmotic effect is greater in agar-agar than in agarose gels. The extension of the electrophoretic run can be precisely fixed by controlling the migration of a marker. One of the markers routinely used is the protein stain Amido black. The second step is a double immunodiffusion performed in the same gel plate (Figure 1A). For this, longitudinal channels are cut parallel to the direction of the electrical flow and separated 4 mm from the original wells where the samples were applied. The channels are filled with the

1. Electrophoresis in agarose gel

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.

Get My Free Ebook


Post a comment