Free electrophoresis

Seidel A, Wiener M, Kruger E, et al. 1986. Studies on the lysosomal binding of 141Ce, 239Np, 239Pu and 241 Am in rat and Syrian hamster liver using carrier-free electrophoresis. Int J Rad Appl Instrum B13(5) 515-518. 

By use of a crude preparation obtained after the cultivation of Aspergillus niger,104 pectinesterase was purified by repeated chromatography on DEAE-cellulose, using gradient elution. The homogeneity of the product was checked by free electrophoresis, sedimentation analysis, and determination of the N-terminal amino acid (phenylalanine). 

The Electrodes. In most commercial and homemade apparatus the electrodes are of platinum wire or platinum-coated silver. Carbon electrodes are just as satisfactory as long as the same buffer continues to be used, but washing out carbon electrodes when changing to new buffers is difficult. Silver electrodes connected with AgCl/KCl bridges give excellent results (F7), as they do in free electrophoresis. 

Dialysis is not currently performed on samples intended to be applied on buffered supporting media. Theoretically, however, this precaution should be taken because increased conductivity, increased ionic strength, and modification of pH cause several disturbances at the application spot, the most important being local depression of the electric field intensity (G10). Some substitute for dialysis is realized if the sample is diluted with a solution of buffer and serum salts, with volume and concentration calculated to give a final salt concentration corresponding to the expected result of combined dialysis and dilution (S31). This method was successful for free electrophoresis of serum and should also be useful in zone electrophoresis on stabilizing media. 

L9. Levin, B., and Oberholzer, V. G., Paper electrophoresis of serum proteins with micro Kjeldahl nitrogen analysis of the protein fractions A comparison with free electrophoresis and salt fractionation methods. Am. J. Clin. Pathol. 23, 205 (1953). 

M3. Mackay, I. R., Volwiler, W., and Goldsworth, P. D., Paper electrophoresis of serum proteins Photometric quantitation and comparison with free electrophoresis. J. Clin. Invest. 33, 855 (1954). 

In the separation of proteins by electrophoresis, the sample is submitted to an electrical field, causing the electrically charged proteins to move in the direction of the applied current. If the experiment is carried out in solution (free electrophoresis) and the proteins have different charge densities, they will move with different velocities, allowing their separation. In practice, the process is normally carried out in a gel matrix, instead of in solution, and the gel can act not only as an inert support for the electrophoresis buffer, but also, if desired, as an active material that interacts with the proteins. 

Example 40 Scale-up of a continuous, carrier-free electrophoresis... 

Fig. 79 Sketch of a continuous, carrier—free electrophoresis cell...

This heat must be dissipated by cooling, which can be done but only to a limited extent The ability to dissipate heat efficiently is usually the factor that limits the speed of electrophoresis, since excess heat leads to non-uniform electrophoresis and a decrease in resolution. The main reason for this is convection in matrix-free electrophoresis in solution, and the effect of temperature on viscosity and diffusion. High temperatures can also lead to denaturation of proteins and nucleic acids. The thinner the layer used for electrophoresis, the more readily is the heat dissipated, and the higher the voltages that can be used. The thickness of the layer will be a compromise between a desire to have a thin layer to minimise heat problems whilst maintaining sufficient capacity to ran samples that can be detected easily. Consis-... 

After two chromatographic runs, porcine chymotrypsinogen A behaves as a homogeneous substance during free electrophoresis at various pH s and... 

The Retina Foundation was a unique place in the late 1950s. The five-story, old tenement house was rebuilt and filled with the most modern biomedical research equipment available. We had analytical ultra-centrifuge, free electrophoresis machine, nuclear magnetic resonance machine, equipment to determine C-14 in the gas phase (before scintillation counters existed), electron microscope, tissue culture equipment with microcinematography, and a superb instrument shop to make new research tools. 

Using this method, some investigators have obtained albumin yields which compare well with those obtained by 1.8 ilf sodium sulfate fractionation and by separation by free electrophoresis (M13). However, several other reports suggest that difficulties may be encountered with the procedure of Pillemer and Hutchinson, which probably arise from variable and uncontrollable factors owing to the high concentration of serum in the... 

Since 1958 when paper electrophoresis of serum proteins was reviewed in this series (P3), there have been no major technical developments in the art. The incorporation of nonionic detergents (Tween 80, Span 20, etc.) in the conventional barbiturate buffer (D14, G18, L14) and the use of tris -borate as buffer (A14, S27) have led to the separation of 7 to 11 globulin fractions. After zone electrophoresis in borate-containing buffer, an increased number of bands can be visualized by staining, but these are not revealed by the schlieren scanning technique after free electrophoresis in similar buffers (S27) the effects of this buffer may be due to interaction between borate and proteins having a high carbohydrate content. 

Hoagland DA, Smisek DL, Chen DY. Gel and free electrophoresis of variably charged polymers. Electrophoresis 1996 17 1151-1160. 

Studies by equilibrium dialysis (83), free electrophoresis (86), isoelectric focusing (87), and DEAE-cellulose chromatography (88) have suggested that the two metal-binding sites on the protein molecule are... 

Parallel with the technical improvement and analytical refinement of measurements grows the difficulty of the interpretation of results. Whereas the identification of the 4-5 peaks achieved with free electrophoresis of a normal serum was comparatively simple, the more complex... 

None of these principal gravity-dependent and gravity-independent processes have been investigated in multistage electrophoresis, which is designed to minimize the impact of these processes on separation quality. All of these processes have been studied in the past in applications to other forms of free electrophoresis as reviewed by Todd [34,41]. 

Based on research experience and needs identified for free electrophoresis in various fields of bioprocessing and analysis such as virology, endocrinology, enzymology, hematology, and mammalian cell culture [64-69], in the present study an attempt is made to overcome the major bottlenecks of free electrophoresis. 

The multistage electrophoretic method has been developed by combining free electrophoresis and multistage extraction and is explored as an improved alternate method for the purification and concentration of cells or macromolecules. The present discussion is restricted to cells and particles. The present design is based on a derivative of the thin-layer multistage extractor design of Albertsson [70] and Treffrey et al. [71] and called ADvanced SEParation apparatus (ADSEP), designed by SHOT, Inc. [42]. 

The particles move in a solvent, typically aqueous salt solutions, in free electrophoresis or Tiselius electrophoresis. In carrier electrophoresis, the particles move in a swollen carrier [for example, paper, starch gels, cross-linked poly(acrylamide)]. 

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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