Resolving power electrophoresis

Two-dimensional gel electrophoresis (2DE) is a two-dimensional technique for protein separation, which combines isoelectric focusing and sodium dodecyl sulphate (SDS) electrophoresis. The high resolving power results from separation according to charge (isoelectric point) in the first dimension and size (mobility in a porous gel) in the second dimension. Depending on the gel size, from several hundred to more than 5,000 proteins can be separated. 

Capillary electrophoresis has also been combined with other analytical methods like mass spectrometry, NMR, Raman, and infrared spectroscopy in order to combine the separation speed, high resolving power and minimum sample consumption of capillary electrophoresis with the selectivity and structural information provided by the other techniques [6]. 

The choice of HPLC as a separation technique in biopharmaceutical development may depend upon its scalability, even if its resolving power is less than other techniques. For example, HPLC may be used in preference to gel electrophoresis or capillary electrophoresis, where scaleup is difficult or impossible. 

CE provides analysis based on orthogonal separation principles compared to other techniques as well as high resolving power. Like slab gel electrophoresis, CE is a family of techniques that resolve sample components by differences in intrinsic molecular characteristics such as size, mass, charge, differential interaction, and isoelectric point (pi). 

Capillary electrophoresis is a new technique that combines the high resolving power of electrophoresis with the speed, versatility, and automation of... 

Reaction mixtures from restriction enzyme digestion may be analyzed directly by agarose gel electrophoresis. This technique combines high resolving power and sensitive detection to allow the analysis of minute amounts of DNA fragments. 

Capillary electrophoresis has found use in the biotechnology industry for structural analysis of recombinant proteins. The high resolving power of CE for charged analytes makes it a powerful tool for the analysis of tryptic digests. Therefore, many of the techniques given here, such as the determination of thiols, carbohydrates, and amino acids, will be employed for this purpose. 

As an analytical tool, electrophoresis is simple, relatively rapid, and has unparalleled resolving power. It is used chiefly for analysis and purification of very large molecules such as proteins and nucleic acids. Highly sensitive detection methods have been developed to monitor and analyze electrophoretic separations. 

Some residues require derivatization to enhance the extractability, clean-up, or subsequent chromatographic resolution and determination steps. Instead of chromatography, capillary electrophoresis with a high resolving power may be considered as well. 

The particularly high resolving power of moderately concentrated gel media is to a large extent a consequence of molecular sieving acting as an additional separative factor. For example, blood serum can be separated into about 25 components in polyacrylamide gel, but only into 5 components on filter paper or by moving boundary electrophoresis. 

One of the most useful techniques for visualization of the proteome is two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2-D SDS-PAGE). This technique possesses unmatched resolving power for separation of proteins [2-4] and has been used extensively to analyze proteins [5-8], their regulation [9-18], and posttranslational modifications [19-22], Several tech-... 

The advancement of modern biochemistry and developments in micro- and macromolecular separations have been intimately linked. Capillary electrophoresis offers major advantages over other separation techniques, including speed and resolving power. The potential of capillary electrophoresis seems so vast that it will significantly complement the technology of high-performance liquid chromatography. However, because of unique characteristics of capillary electrophoresis, it will also... 

With V— 102-5 x 104 and z = 1-10, as above, we see that from 10 to 800 distinct peaks can theoretically be resolved in electrophoresis. (In practice the number of resolvable components is much less than nc due to statistical peak overlap as explained in Section 6.7). This enormous resolving power is consistent with observation (see Figures 8.2 and 8.4). Theory thus provides an explanation of the unusual power of electrophoresis and an insight into the variables (z and V) that must be manipulated for increased performance. 

From a strict biochemical point of view a clear-cut definition of the role of the liver in the biosynthesis of any particular plasma protein can be made only when the particular protein has been clearly and cleanly isolated, as in the case of fibrinogen. The practical difficulties of effecting such isolations on a small scale from isotopic labeling studies of the plasma proteins, such as we have described, seriously militate against such a detailed demonstration at present. The use of fractionation techniques with greater resolving power such as acrylamide gel electrophoresis already show some promise in our laboratory toward affording a more definitive picture of the biosynthetic role of the liver and the nonhepatic tissues in plasma protein production. 

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