96 A. TISELIUS [6 



of its simplicity and good resolution, and has found wide application as 

 a micro-method. To a certain extent its relation to other types of zone electro- 

 phoresis is similar to that of paper chromatography to column chromato- 

 graphy, but the heat generated by the current may give rise to evaporation 

 disturbances which make the method less suitable for quantitative work. 



Recent experiments in Uppsala with very small columns show that quan- 

 tities as small as those commonly used in paper strip electrophoresis may 

 also be studied in columns under more rigidly controlled conditions than 

 in the paper strip method. 



In the discussion which is to follow Dr Porath will give some examples of 

 the application of the zone column technique. I shall limit myself to a few 

 comments on the advantages and limitations of the zone methods in general. 



The advantages of the zone methods over the classical boundary pro- 

 cedure are obvious, especially that in the former it is possible to achieve 

 complete separation of the migrating components and to isolate them for 

 further investigation. In the boundary method the zones overlap partially 

 and conclusions are drawn only from optical observation of their migra- 

 tion. A further study of the properties and diiference in the characteristics 

 of components is difficult or (with closely similar substances) impossible. 

 In addition the zone methods are much easier to apply to low-molecular 

 weight compounds, like amino acids and peptides. Boundary electrophoresis 

 of such substances has proved very difficult because of their considerable 

 contribution to the conductivity of the system, which results in boundary 

 anomalies. In zone electrophoresis these anomalies are apparently much less 

 pronounced, probably because of the easy interchange of ions by diffusion 

 between the front and the rear of a zone and by a gradual automatic adjust- 

 ment of the concentration to lower values. A further advantage, which has 

 been demonstrated by Porath (1957 b), is that the reduction in diffusion in 

 powdered supporting medium is much greater than the reduction in cur- 

 rent; the heat generated by the current being carried away by the stabilizer 

 almost as efficiently as in free solution. This contributes very much to the 

 separation efficiency, particularly with low-molecular weight substances. 



One has to pay a price, however, for all these advantages. Most materials 

 suitable for use as supporting media show some interaction with proteins 

 and other substances. There is of course a mechanical hindrance but, above 

 all, specific interaction by adsorption is very common, leading to a spread- 

 ing of the zones by taihng and influencing their mobilities in a way which 

 is difficult to predict. This is the key-problem of the zone method and much 

 work has already been done to find more inert supporting materials which 

 would show as small interaction as possible with the migrating substances. 

 Flodin and Kupke (1956) have prepared a partially esterified cellulose by 

 ethanolysis, which has proved much superior to ordinary cellulose for zone 

 electrophoresis columns and is now generally used in the Uppsala labora- 

 tory for protein and peptide separations. No doubt other possibilities will 



