Cellulose electrophoresis supports

K29. Kohn, J., A cellulose acetate supporting medium for zone electrophoresis. Clin. Chim. Acta 2, 297-303 (1957). 

Traditional electrophoresis paper, cellulose acetate or polymeric gels used as a supporting medium for the electrolyte solution enclosed tank with electrodes and buffer reservoirs dc power supply. 

Electrophoresis has also been employed to separate neomycin from analytically-interfering substances such as proteins. Hence Brammer and Hemsonl82 have determined the neomycin content of blood serum. Neomycin was separated from the serum proteins by electrophoresis on cellulose acetate and assayed colorimetrically following elution from the support. 

Other supports in electrophoresis are agarose gels, paper, or celluloses acetate. 

The connection between electrode chambers of a horizontal electrophoresis apparatus filled with buffer A and the cellulose support is made by wetted paper bridges. To avoid liquid moving, the buffer level must be the same in both chambers. A glass plate, laying on the paper bridges, covers the separation plate cooled to 0-4 °C. 

Zone electrophoresis is normally carried out horizontally in a suitable medium such as paper, polyacrylamide gel, starch gel or cellulose acetate. The sample components can be completely separated and quantitatively and qualitatively identified in much lower quantities than by the moving-boundary method. The procedure consists of saturating the support material with a buffer solution. The ends of the strip of support are immersed in separate reservoirs of buffer solution to maintain the saturation. The sample is then applied as a narrow band near one end of the support strip. A voltage potential is created down the length of the strip causing the sample components to ionize and then migrate at a rate dependent on their charge, molecular size and interactions with the support medium. When the process is complete, the strip is removed and developed for examination of the separated components. Densitometry is normally used for quantitation of the bands after suitable color development. 

Zone electrophoresis is used mainly as an analytical technique and, to a lesser extent, for small-scale preparative separations. The main applications are in the biochemical and clinical fields, particularly in the study of protein mixtures. Like chromatography, zone electrophoresis is mainly a practical subject, and the most important advances have involved improvements in experimental technique and the introduction and development of a range of suitable supporting media. Much of the earlier work involved the use of filter paper as the supporting medium however, in recent years filter paper has been somewhat superseded by other materials, such as cellulose acetate, starch gel and polyacrylamide gel, which permit sharper separations. 

To reduce adsorption of cellulose powder (T2) Flodin introduced ethanol-treated cellulose as a supporting medium for zone electrophoresis columns (FI). The main advantage is low adsorption, so that the column can be eluted and used over and over again. In the large models up to 2 liters of serum are separated, while microcolumns are under development which should give excellent results for clinical work (PI, Tl). 

The electrophoretic separation technique is based on the principle that, under the influence of an applied potential field, different species in solution will migrate at different velocities from one another. When an external electric field is applied to a solution of charged species, each ion moves toward the electrode of opposite charge. The velocities of the migrating species depend not only on the electric field, but also on the shapes of the species and their environmment. Historically, electrophoresis has been performed on a support medium such as a semisolid slab gel or in nongel support media such as paper or cellulose acetate. The support media provide the physical support and mechanical stability for the fluidic buffer system. Capillary electrophoresis (CE) has emerged as an alternative form of electrophoresis, where the capillary wall provides the mechanical stability for the carrier electrolyte. Capillary electrophoresis is the collective term which incorporates all of the electrophoretic modes that are performed within a capillary. 

Most practical applications of electrophoresis in biochemistry employ some form of zonal electrophoresis, in which the aqueous ionic solution is carried in a solid support and samples are applied as spots or bands of material. Paper electrophoresis, cellulose acetate strip and cellulose nitrate strip, and gel electrophoresis are all examples of zonal... 

The support medium provides the matrix in which protein separation takes place. Various types of support media are used in electrophoresis and range from pure buffer solutions in a capfilary to insoluble gels (e.g., sheets, slabs, or columns of starch, agarose, or polyacrylamide), or membranes of cellulose acetate. Gels are cast in a solution of the same buffer to be used in the procedure and may be used in a horizontal or vertical direction. In either case, maximum resolution is achieved if the sample is applied in a very fine starting zone. Separation is based on differences in charge-to-mass ratio of the proteins and, depending on the pore size of the medium, possibly molecular size. 

Electrophoresis in Cellulose. Duraiswami et al. and Saxena and Rathman have used electrophoresis on cellulose columns in the purification of FSH (D5, S4), and Squire et al. have employed the same technique in the purification of LH (S18). These investigators found that considerable losses of gonadotropic activity occurred when cellulose was used to support the medium for electrophoresis e.g., Saxena and Rathman found that 33% of their FSH was lost by this procedure (S4). The fact that LH is adsorbed onto cellulose has been used in the purification of that hormone after it has been labeled with radioactive iodine (S24). The LH is eluted off the cellulose with 50 mg of albumin/100 ml buffer at pH 8.6. 

Serum LDH isoenzymes can be separated by electrophoresis on agarose gel or cellulose acetate membrane, usually at pH 8.6. After separation, their location is determined by incubation of the support medium in a... 

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