Zone Convection Electrofocusing

A great advance in the equipment for isoelectric focusing has recently been made by Valmet (40). He has described a new principle for electrofocusing called zone convection electrofocusing. In this technique the stabilizing against convection does not call for any capillary system, density gradient or membranes. 

There are no foreign additives, such as sugar or gel, used to stabilize against convection. 

If a protein becomes insoluble at its isoelectric point, the resulting precipitate does not disturb the separation to the same extent as during electrofocusing in a column. 

Rose (41) reported an apparatus based on zone convection electro-focusing. Its purpose was preparatory separation or treatment prior to column electrofocusing. 

Stabilization by using rotating tubes has been described by Hjert n (42). His apparatus for free electrophoresis was used. Too little is known about the capacity and resolution of his method to make an evaluation possible. He has, however, reported good results. 

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The stabilization techniques of importance today in electrofocusing are (a) density gradient electrofocusing, (b) gel electrofocusing, and (c) zone convection electrofocusing. These methods are described in detail later in this article. 

Zone convection electrofocusing equipment, as created by Valmet (40). 

LKB-Produkter AB is reported to be planning the manufacture and sale of apparatus for zone convection electrofocusing. 

Figure 32. Zone convection electrofocusing apparatus according to Valmet. The water-cooled lid is lifted from the trough. (Valmet, 78,53.)...

Figure 33. Schematic drawing of the zone convection electrofocusing apparatus. (A) The filled trough (T), before or after the experiment. The separated protein zones are illustrated by the dark areas. The lid (L), lifted up as before or after the run. (B) Both parts put together, as during the experiment. Notice the different positions of the ampholyte solution level. Both parts are water-cooled. The arrows show the circulation of cooling water. (Pt) indicates the electrodes. (C) Details of the electrode Pt—platinum wire, G—sintered glass filter, a—ampholyte solution, C—cooling water. (Valmet, 53.)...

Figure 35. Density gradient electrofocusing of part of defatted human serum albumin. The pH 5.2-S.9 gradient was prepared by the method of zone convection electrofocusing. (Valmet, 78.)...

E. Valmet, Zone Convection Electrofocusing Presides Biol. Fluids, Brugge, 1969b. 

Of the techniques mentioned, the density gradient technique is used in most electrofocusing experiments reported to date, although gel electro-focusing has recently had wide applications. Valmet s zone convection eUcirofocusing is a promising new approach to the stabilization problem. There also, no added stabilizing medium is necessary. 

Isoelectric focusing has undei ne extensive developments in recent years. The most important are self-stabilizing zone convection electro-focusing according to Valmet (40), and the relatively new branch of gel electrofocusing. 

Figure 34. Zone convection electrofocumi of a 40% Ampholine solution, pH 4-6. The electrofocusing time was 5 days and the voltage 800-1200 V. The contents of two different compartments were taken and diluted to 0.5%. They were electrofocused by density gradient electrofocuaing method. Each fraction covered about 0.5 pH units. pH-gradients with specified shapes can also be constructed by adding or removing different fractions (Valmet, 53).

The method of stabilization against convection determines the character of the equipment. Gels, density gradients or zone convection indeed give the equipment different images. Many of the techniques used in electrophoresis for stabilization today have not yet been studied and tested for electrofocusing. Further developments based on those techniques can be expected. 

Electrofocusing as a preparative technique will have a future. The recent development of zone convection is a demonstration of that. 

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