Free solution electrophoresis

Hoagland, DA Smisek, DL Chen, DY, Gel and Free Solution Electrophoresis of Variably Charged Polymers, Electrophoresis 17, 1151, 1996. 

Jorgenson reported the use of glass capillaries for free solution electrophoresis 25 years ago (Jorgenson andLukacs, 1981,1983). Aplug of analyte was introduced into a buffer-filled capillary and separated at high electric fields. Capillaries of 75 im inner diameter were employed, and detection of labeled amino acids and peptides was based on fluorescence. 

Karim, M.R., Shinagawa, S., Takagi, T. (1994). Electrophoretic mobilities of the complexes between sodium dodecyl sulfate and various peptides or proteins determined by free solution electrophoresis using coated capillaries. Electrophoresis 15, 1141-1146. 

Free solution electrophoresis (capillary zone electrophoresis). 

Cottet, H., and Gareil, P. (2000). From small charged molecules to oligomers A semiempirical approach to the modeling of actual mobility in free solution. Electrophoresis 21, 1493-1504. 

Electrophoresis experiments in glass tubes were reported as early as in the nineteenth century, but the first real breakthrough occurred in the first half of the twentieth century when the Swedish chemist Arne Tiselius applied free-solution electrophoresis—i.e moving boundary—to serum protein analysis, for which he later received the 1937 Nobel Prize [2], In less than two decades, just after the striking scientific discovery of the double-helical structure of DNA by Watson and Crick in 1953 [3] and the following unveiling of the genetic code, electrophoresis became a standard and indispensable tool in the field of modern... 

High performance capillary electrophoresis is one form of free-solution electrophoresis. CE is useful to researchers and analysts working in areas in which traditional electrophoresis is customarily applied (e.g., biopolymer analysis) and also in disciplines not usually associated with electrophoretic analysis, such as inorganic ion analysis. The potential application areas of CE are vast, because this technique can separate a variety of ligates, from inorganic ions up to intact cells, using the same instrumental hardware designed for separations based on different physical-chemical mechanisms. 

Since the late 1960s, capillaries have been used for free-solution electrophoresis, but instrumental limitations, such as capillary materials, the large internal... 

CZE is high voltage, free-solution electrophoresis carried out in a capillary. The capillary is filled with the running electrolyte (a buffer solution), and the ionic analytes are separated on the basis of the differences in their electrophoretic mobilities. The favorable ratio of surface area to volume allows the dissipation of the Joule heat from the capillary and the application of high electric fields with rapid and efficient separations. Also, the anticonvective characteristic of the capillary limits the process of zone diffusion, maintaining the efficiency of separation without the need of further anticonvective media such as gels. 

Nathakarnkitkool S, Oefner PJ, Bartsch G, Chin MA, Bonn GK (1992) High resolution capillary electrophoretic analysis of DNA in free solution. Electrophoresis 13 18-31. 

The enzymes from malt sorghum and sweet potato were completely homogeneous by both gel and free-solution electrophoresis, and the three individual wheat components and the soya-bean enzyme were essentially homogeneous a mixture of the three wheat enzymes, however, gave three discernible peaks. Sedimentation studies on the wheat enzymes in the presence of iodine and potassium iodate showed no change in the sedimentation coefficient under these conditions, indicating the absence of intermolecular S — S bonding. There was also, from sedimentation, no evidence of dissociation into subunits in the presence of 8 M urea. 

Capillary Electrophoresis in Analytical Biotechnology, Righetti, P.G., Ed. CRC Press Boca Raton, FL, 1996. Capillary Electrophoresis—Theory and Practice. New Directions in Organic and Biological Chemistry Series Camilleri, P., Ed. CRC Press Boca Raton, FL, 1997. Heller, C. Slater, G.W. Mayer, P. Dovichi, N. Pinto, D. Viovy, J.-L. Drouin, G. Free-solution electrophoresis of DNA. J. Chromatogr., A 1998, 806 (1), 113-221. 

This mode of electrophoresis, in which the electrolyte migrates through the capillary, is the most widely used. In this mode, samples are applied as a narrow band that is surrounded by the electrolyte buffer. This electrolyte can be, depending upon the application, acidic (phosphate or citrate) or basic (borate) or an amphoteric substance (a molecule possessing both an acidic and an alkaline function). The electro-osmotic flow increases with the pH of the liquid phase or can be rendered inexistent. This procedure is also called, in contrast to CGE (cf. Section % A3), free solution electrophoresis. 

Size-based separations of homogeneous polyelectrolytes, such as DNA, are not possible in free solution electrophoresis [159]. This is due to the proportionality of the friction hydrodynamic force and total charge of the molecule to its length. The friction hydrodynamic forces exerted on the free-drained polymer coil while it moves as well as the accelerating electrostatic force both increase proportionally with the addition of a nucleotide to the chain. This is why one must typically use a sieving media, such as a gel or an entangled polymer solution, to obtain size-based separations of DNA using electrophoresis. 

From the beginning of free solution electrophoresis, band broadening from thermal effects was foreseen as the main problem restricting its development [6]. The energy generated as heat by application of an electric field to an electrolyte solution in a column raises the solution temperature, but more critically, results in a radial... 

Free-solution electrophoresis of DNA that has been conjugated to a friction-perturbing entity, which we call a drag-tag, was not feasible until after the development of CE in the 1990s. Free-solution electrophoresis generates a significant amount of heat, which must be dissipated. The narrow channels used in CE have diameters in the order of 25-100 p,m, which efficiently remove this Joule heat. Free-solution conjugate electrophoresis (FSCE) was first examined quantitatively in 1994 by Mayer... 

The first iteration of FSCE theory predicted the potential sequencing of up to 2000 bases in less than 1 h under perfect (diffusion-limited) conditions using a drag-tag with an a between 100 and 200. The initial theory, however, assumed that /u.(M) and the diffusion coefficient D M) eould be related using the Nernst-Einstein equation, which it turns out is not a valid assumption under free-solution electrophoresis conditions. This assumption caused Mayer to overestimate the potential performance of DNA sequencing by FSCE. Eurther development of the theory with a eorreetion for this error led to the following equation that estimates the maximum number of bases that ean be sequenced by a drag-tag with a specified a-value ... 

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