Electrophoresis convection method

Fig. 1. Effect of pH on the corrected sedimentation constant of conalbumin at ionic strength 0.1, protein concentration 1.1—1.3 g./lOO ml. , conalbumin prepared by electrophoresis—convection O. conalbumin prepared by the acid precipitation method Q, conalbumin prepared by electrophoresis—convection, exposed for 1 hr. to pH 2—3 before sedimentation at pH 5.2 A, conalbumin prepared by the acid precipitation method, exposed for 1 hr. to pH 2.5—3.4 before sedimentation at pH 4.5—7.5. (Arch. Biochem. Biophys. 61, 51 [1956]).

All separation procedures rely on some element of differential transport the separation may be based on differences in phase equilibria, as in chromatography, or on the kinetics of transport, as in electrophoresis and centrifugation. Precipitation procedures, filtration and dialysis are also members of the broad dass of separation methods these have been discussed in an earlier chapter (Sect. 3.4). The transport processes involved in separating components are often opposed by dispersion processes such as diffusion and convection, which have to be minimised to achieve the best separation results. 

The use of tubes of rectangular cross section provides a maximum amount of wall area and so facilitates the removal of heat to the surroundings, and the maintenance of the apparatus at somewhat below 4 , where the variation of the density of the liquid with temperature is very small, greatly diminishes convection effects. With these improvements, and a device for obtaining initially sharp boundaries, the macroscopic method has proved a valuable means for the study of electrophoresis, and for its utilization to separate particles moving with different speeds. 

Instant coagulation also excludes methods that require a stable dispersion, such as electrophoresis. The manufacturers of certain types of zetameters claim that their instruments are suitable to perform measurements in electrolyte solutions up to about 1 M. However, in order to use a zetameter, one has to prepare a stable dispersion first, and this may be problematic. Electro-osmosis does not require stability against sedimentation, but other problems, such as low absolute values of the potential (which may be smaller than the scatter of results) and the production of heat, convective currents, or electrolysis products (acids, bases, and gases), severely limit the application of classical electrokinetic methods (including electrophoresis) in measurements at ionic strengths greater than 0.1 M. Very few publications report potentials obtained by classical electrokinetic methods at higher electrolyte concentrations, and the results are controversial. 

This technique, although described in the literature, has been given little attention. Isoelectric focusing without stabilizing media can be done in an apparatus similar to Tiselius free zone electrophoresis [152]. Separation itself occurs in a horizontal quartz tube that is rotated at 40 rpm to counteract convective forces [153]. The pH gradient is evolved without the addition of ampholytes, however this method leads to either too steep or too shallow gradients and is therefore not practically applicable. Another alternative method for free solution isoelectric focusing is the application of polyethylene coils however this procedure makes use of ampholines [154]. 

When performing a run in practice, the remixing of separated zones, due to undesired convection and other movements in the liquid electrolyte system, has to be prevented. The liquid system must be stabilized against convection in some way. In electrophoresis such stabilization can be achieved in many ways. Valmet (53) has made the following summary on methods used ... 

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. 

Such heat removal almost eliminates convection so that the peaks approach the theoretical limit [24] of being broadened only by diffusion (from high to low concentration). In favorable cases, about 1 million theoretical plates have been realized [43], which makes capillary electrophoresis the most highly efficient electrophoresis method [11]. 

Currently, analytical approaches are still the most preferred tools for model reduction in microfluidic research community. While it is impossible to enumerate all of them in this chapter, we will discuss one particular technique - the Method of Moments, which has been systematically investigated for species dispersion modeling [9, 10]. The Method of Moments was originally proposed to study Taylor dispersion in a circular tube under hydrodynamic flow. Later it was successfully applied to investigate the analyte band dispersion in microfluidic chips (in particular electrophoresis chip). Essentially, the Method of Moments is employed to reduce the transient convection-diffusion equation that contains non-uniform transverse species velocity into a system of simple PDEs governing the spatial moments of the species concentration. Such moments are capable of describing typical characteristics of the species band (such as transverse mass distribution, skew, and variance). 

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