Protein Chemistry. Structure, Properties, Research Methods - Shendryk A.N. 2022

Methods for Experimental Study of Protein Structure
Protein Separation Methods
Experimental Electrophoresis Methods - Disc Electrophoresis

This type of Electrophoresis is a variant of gel electrophoresis. It is performed in gradients of medium pH, gel concentration, and voltage magnitude. The schematic diagrams of Disc electrophoresis and the apparatus used for it are shown in Fig. 4.6 and 4.7.

Class="center">

Fig. 4.6 Schematic diagram of disc electrophoresis in a Glass tube

The gel is contained within a vertical glass Column and divided into three regions. The upper region is the sample or stacking gel, the middle is the spacer or concentration gel, and the lower is the resolving gel. The stacking and concentration gels have larger pore sizes than the resolving gel and are prepared in a buffer solution with low Ionic strength and a lower pH value.

The lower ionic strength provides greater electrical resistance, resulting in a steeper electric potential gradient in the upper part of the column.

Fig. 4.7 Schematic diagram of the disc electrophoresis apparatus.

1 - lid; 2 - test sample; 3 - stacking gel (spacer gel); 4 - resolving gel; 5 - upper reservoir; 6 - buffer; 7 - lower reservoir; 8 - glass tube.

The large pores and potential gradient at the top of the column allow the sample to move rapidly through its upper layers. As a result, sample concentration occurs at the boundary between the spacer and the resolving layer. In the lower gel, a molecular sieving effect is created due to the higher ionic strength, higher pH value, and smaller pore size. Passing through this gel, molecules of the same species form narrower and better-resolved zones than in standard gel electrophoresis (see figure).

Following electrophoresis and gel staining, distinct zones appear for individual components that differ in mobility — Q/r (zones 1–6).