Continuous electrophoretic separations

Svensson, H., and Brattsten, I., An apparatus for continuous electrophoretic separation in flowing liquids. Arkiv Kemi 1, 401 (1949). 

Many novel electrophoretic devices and techniques have been proposed for continuous electrophoretic separations such as the velocity-stabilized Biostream/Harwell device (9-11), the recycle continuous-flow electrophoresis device (12-14), Bier s isoelectric focusing technique (15),... 

Mesaros, J. M., Luo, G, Roeraade, J., and Ewing, A. G, Continuous electrophoretic separations in narrow channels coupled to small-bore capillaries. Anal. Chem., 65, 3313,1993. 

Fre- flow electrophoresis is the only continuous electrophoretic separation method. The electrical field is applied perpendicular to a continuous stream of buffer film, which flows through a 0.5-1.0 mm wide cuvette. At one end the sample is injected at a defined location, and at the other end, the fractions are collected by an array of tubes. The different electrophoretic mobilities of the sample components lead to differently strong but constant deviations inside the stream... 

Fig, 7.24 (a) The principle of the Biostream process for continuous electrophoretic separation of proteins. (Courtesy Harwell Laboratory, UK Atomic Energy Authority,)... 

A new and ingenious principle for preparative electrophoresis was published from the Tiselius laboratory, in 1949, by Svensson and Brattsten (72). They described a method for continuous electrophoretic separation in liquids flowing through a packed channel. The current was applied at right angles to the flowing liquid and the separated fractions were col-... 

Several workers, Grassman and Hannig (30), Durrum (16), Strain (70), and Brattsten and Nilsson (3), have described methods of continuous electrophoretic separation in flowing liquids, employing a broad sheet of filter... 

Ultimately, however, it should be noted that these examples of classical gel electrophoretic separations are batch processes and therefore limited in sample throughput. To achieve true preparative-scale separations by electrophoresis, it becomes necessary to convert to continuous processes. 

The development of electrophoretic techniques afforded possibilities for fractionations based on charge density differences. Duxbury (1989) has reviewed applications of different electrophoretic separation methods, including zone electrophoresis, moving boundary electrophoresis, isotachophoresis, and isoelectric focusing (IEF). Preparative column electrophoresis (Clapp, 1957) and continuous flow paper electrophoresis (Hayes, 1960 summarized by Hayes et al., 1985) methods have been used to separate components isolated from sapric histosol soils. These techniques allowed separation of polysaccharides from the colored components the electrophoretograms of the colored components were diffuse, showing a continuum of components of different charge densities. 

Two-dimensional (2D) electrophoretic methods are important variants of electrophoresis. In Section 6.4 we noted that electrophoresis could combine with flow in a 2D system providing continuous (and thus preparative) separation. Fundamentally, this is simply zonal electrophoresis converted into a continuous form by nonselective flow (see Section 7.5). If we observe the separation at different positions along the flow axis (as illustrated for one position in Figure 7.3), we have essentially a series of snapshots of the zones evolving with time. Each component zone is deflected from the flow axis at a unique angle as a consequence of the evolution of the electrophoretic separation. 

Elaboration of new electrophoretic methods for PolyP separation is continuing. For example, capillary electrophoretic separations of sodium PolyPs with chain lengths of 5 to 44 has been reported. In this work, a buffer containing pyromellitic acid, triethanolamine and hexamethonium hydroxide gives high-resolution separation of linear and cyclic PolyPs (Stover, 1997 Wang and Li, 1998). 

Gavin PF, Ewing AG. Characterization of electrochemical array detection for continuous channel electrophoretic separations in micrometer and submicrometer channels. Anal Chem 1997 69 3838-3845. 

I) Zone electrophoresis, an electrophoretic separation technique, conducted In a continuous buffer system. Samples are Injected and move under the applied field they separate into distinguishable zones If their mobilities are sufficiently apart. 

Proteins can ba fractionated by electrophoretic techniques on the basis of one or a combination of their three major properties size, net charge and relative hydrophobicity. Electrophoresis under native conditions is ideal for soluble proteins, where biological properties can often be retained. In contrast, more vigorous and often denaturing conditions must be used for analysis of less soluble proteins. Electrophoretic separations can be carried out using either a continuous or discontinuous (Multiphasic) buffer system. The techniques are referred to as continuous zone electrophoresis (CZE) or discontinuous ("disc") electrophoresis (also known as multiphasic zone electrophoresis, MZE). 

This procedure represents a preparative version of zone electrophoresis. The apparatus and technique has been previously elaborated [294,295] separation itself is carried out either in a stream of electrolytic solution or on a sheet of cardboard (curtain electrophoresis). In the free flow version the separation is carried out in a cell formed by two parallel glass plates (50 x 50 cm) situated 0.5-1.0 mm apart. It is necessary to ensure an equal and laminar flow of the electrolyte, which is carried out by feeding the buffer through a multichannel peristaltic pump. The sample is continuously applied in the middle near the upper edge of the cuvette (or paper sheet). The electrophoretic separation occurs transversally between vertical electrodes located on the right and left hand side of the separation cuvette (Fig. 6.33). The separated fractions are collected at the lower end of the cell by a system of small communicating vessels or by a multi-channel pump. In the version using paper the... 

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