Electrophoretic mobility buffer

Electroosmotic Mobility When an electric field is applied to a capillary filled with an aqueous buffer, we expect the buffer s ions to migrate in response to their electrophoretic mobility. Because the solvent, H2O, is neutral, we might reasonably expect it to remain stationary. What is observed under normal conditions, however, is that the buffer solution moves toward the cathode. This phenomenon is called the electroosmotic flow. 

In the presence of a buffer with constant composition across the electrophoretic chamber, the angle of deflection (0) of the solute in the electric field is dependent upon the intrinsic electrophoretic mobility of the solute (p. ), the linear velocity of the buffer (v) and the current through the chamber (I) and can be described as [17] ... 

The unknown phosphate ester had the same electrophoretic mobility as 2-deoxy ribitol 5-phosphate and it seemed reasonable to expect that in the conditions used (0.1 M pyridinium acetate buffer of pH 5) 2-deoxy ribitol-4- and -5-phosphates would behave similarly therefore it was considered probable that the unknown phosphate ester is 2-deoxy ribitol 4-phosphate, resulting from the reduction of the periodate resistant 2-deoxy ribose 4-phosphate. However, the possibility that both 2-deoxy ribitol 4-phosphate and 2-deoxy erythritol 3-phosphate (formed from... 

The presence of Individual chains In a hemoglobin variant can also be demonstrated by electrophoresis at alkaline pH after the protein has been dissociated Into Its subunits through exposure to 6 M urea In the presence of 3-mercaptoethanol. The buffer is either a barbital buffer or a tris-EDTA-boric acid buffer, pH 8.0 - 8.6, and contains 6 M urea and 3-niercapto-ethanol. Dissociation of the hemoglobin Into subunits Is best accomplished In a mixture of 1 ml 10 g% Hb (or whole hemolysate), 4 ml 6 M urea barbital or tris-EDTA-boric acid buffer, and 1 to 1.5 ml 3-mercaptoethanol. After 30 minutes to 1 hour the sample Is subjected to cellulose acetate or starch gel electrophoresis. Each chain has a specific mobility and an alteration In electrophoretic mobility easily Identifies the abnormal chain. 

Log k appears to correlate with log P for standards between log P —0.5 to 5.0. One limitation of this method is that solutes must be electrically neutral at the pH of the buffer solution because electrophoretic mobility of the charged solute leads to migration times outside the range of Tm and TEof- Basic samples are therefore run at pH 10, and acidic samples at pH 3, thus ensuring that most weak acids and bases will be in their neutral form. This method has been used in a preclinical discovery environment with a throughput of 100 samples per week [24]. 

Table 16. Electrophoretic Mobility of Various Lignins in Glycine-NaCl Buffer, pH 10-7 (100).

In a solution without chiral selectors, enantiomers cannot be distinguished from each other through their electrophoretic mobility. Separation can, however, be achieved when the buffer solution contains certain chiral compounds. The chiral compounds used to distinguish enantiomers are referred to as selectors. 

Buffer parameters such as type of buffer used, its pH and ionic strength, and organic solvent content can be varied to influence the electrophoretic mobility of target compounds leading to an improved peak resolution. 

The electrophoretic mobility of an ion is inversely related to the ionic strength of the buffer rather than to its molar concentration. The ionic strength (ytt) of a buffer is half the sum of the product of the molar concentration and the valency squared for all the ions present in the solution. The factor of a half is necessary because only half of the total ions present in the buffer carry an opposite charge to the colloid and are capable of modifying its charge ... 

Anions and uncharged analytes tend to spend more time in the buffered solution and as a result their movement relates to this. While these are useful generalizations, various factors contribute to the migration order of the analytes. These include the anionic or cationic nature of the surfactant, the influence of electroendosmosis, the properties of the buffer, the contributions of electrostatic versus hydrophobic interactions and the electrophoretic mobility of the native analyte. In addition, organic modifiers, e.g. methanol, acetonitrile and tetrahydrofuran are used to enhance separations and these increase the affinity of the more hydrophobic analytes for the liquid rather than the micellar phase. The effect of chirality of the analyte on its interaction with the micelles is utilized to separate enantiomers that either are already present in a sample or have been chemically produced. Such pre-capillary derivatization has been used to produce chiral amino acids for capillary electrophoresis. An alternative approach to chiral separations is the incorporation of additives such as cyclodextrins in the buffer solution. 

Figure 4.19. Torsion constant a versus buffer concentration for supercoiled M13mp7 DNA in different buffers. All samples except that in 10 mM Tris contain 10 raW NaCl, so all have between 10 and 12 mM univalent positive ions. The sample in the middle contains only 10 mM NaCl. The numbers (1, 2, and 4) in the sample label refer to the gel electrophoretic mobilities, which reflect different tertiary structures, as described in the text. The a samples all contain varying amounts of Tris, while the b samples all contain citrate.

In CZE, the capillary, inlet reservoir, and outlet reservoir are filled with the same electrolyte solution. This solution is variously termed background electrolyte, analysis buffer, or run buffer. In CZE, the sample is injected at the inlet end of the capillary, and components migrate toward the detection point according to their mass-to-charge ratio by the electrophoretic mobility and separations principles outlined in the preceding text. It is the simplest form of CE and the most widely used, particularly for protein separations. CZE is described in Capillary Zone Electrophoresis. ... 

Although protein behavior in SDS-containing buffers is qualitatively different from small molecules, applications of MEKC-type conditions have been applied to many protein separations. In applications using uncoated capillaries, protein-wall interactions are eliminated because of the anionic character of SDS-protein complexes. In applications using coated capillaries with no EOF, the high electrophoretic mobility of SDS-protein complexes can decrease analysis time. 

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