Thin-layer isoelectric focusing

Kumaratilake, L. M. Thompson, R. C. A. (1979). A standardized technique for the comparison of tapeworm soluble proteins by thin-layer isoelectric focusing in polyacrylamide gels, with particular reference to Echinococcus granulosus. Science Tools, 26 21-4. 

Delincee, H., and Radola, B.J. (1978). Determination of isoelectric points in thin-layer isoelectric focusing The importance of attaining the steady state and the role of C02 interference. Anal. Biochem. 90, 609-623. 

This effect is illustrated in Fig. 17.1. Multidimensional chromatography separations can be done in planar systems or coupled-column systems. Examples of planar systems include two-dimensional thin-layer chromatography (TLC) (Consden et al., 1944 Grinberg et al., 1990), where successive one-dimensional TLC experiments are performed at 90° angles with different solvents, and 2D electrophoresis, where gel electrophoresis is run in the first dimension followed by isoelectric focusing in the second dimension (O Farrell, 1975 Anderson et al., 1981 Celis and... 

Fig. 4.5.2 Actual strategies for CDG diagnosis. Initial investigations on CDG patients are routinely carried out by isoelectric focusing (IEF) of serum transferrin. With a CDG type I pattern, subsequent analysis should imply determination of phosphomannomutase (PMM) and phos-phomannose isomerase (PMI) activities. Further studies, like analysis of the lipid-linked- and protein-bound-oligosaccharides, determination of enzyme or sugar transporter activities and molecular biology studies often have to be performed in more specialised laboratories. HPLC High-performance liquid chromatography, TLC thin-layer chromatography...

Isoelectric focusing and electrophoresis were used for protein mapping and to study protein extraction in horizontal ultra-thin-layer format [138], The 0.12-0.36-mm-thick polyacrylamide gel layer was deposited onto tiny glass plates (e.g., microscope slides). The method enabled the analysis of 1-ng tissue culture specimens. Ultra-thin-layer polyacrylamide isoelectric focusing gel was also employed in two-dimensional analysis of plant and fungal proteins. Marlow et al. [139] reported on the use of 0.2-mm semirigid backing (polyester)-supported... 

The introduction of density gradients allowed several improvements in the analytical and preparative capabilities of centrifugal processes. First, it is possible to pour the sample in the top of a tube, in a more or less thin layer differences in density between sample and separation medium avoids convection and delays the homogenization process. Second, it is possible to diminish convection fluid movements in the tube convection can be produced by a difference in temperature between different parts of the centrifugal rotor. Third, it is possible to focus a particle when its reaches its own density, in a process analogous to isoelectric focusing. 

Isotachophoresis is similar to isoelectric focusing but includes the use of additional ampholines or multiphasic buffer systems to act as spacers to improve the separation and resolution. Each of these techniques can be run using columns, tubes, thin layers, or slabs and are comprehensively dealt with by Andrews (1981). 

Gll. GSrg, A., Postel, W., and Westermeier, R., Ultra-thin-layer horizontal electrophoresis, isoelectric focusing and protein mapping in polyacrylamide gels on cellophane. In Electrophoresis 79 (B. J. Radola, ed.), pp. 67-78. Walter de Gruyter, Berlin, 1980. 

Fluorescence bound to albumin, extractable with charcoal treatment under acidic conditions, soluble in water and polar alcohols, diminished at pH <3 or >12 Gel chromatography, isoelectric focusing, protein electrophoresis, thin-layer chromatography, extraction with charcoal and alcohol... 

Figure 22. Apparatus for isoelectric focusing in thin layer polyacrylamide gels according to Awdeh, Williamson, and Askonas (7). (o) polyacrylamide gel 1 mm thick (b) glass plate (c) carbon electrodes 20 cm apart (d) site of sample application. Each sample, 100/400 Mg of protein in less than 50 m1, was pipetted onto the surface of the gel and spread over a rectangular area about 1X2 cm. (Awdeh el al, 39.)...

Creatinine. Electrophoresis Principles Isoelectric Focusing. Enzymes Enzymes In Physiological Samples Industrial Products and Processes Enzyme Assays. Forensic Sciences Alcohol In Body Fluids DNA Profiling Systematic Drug Identification Thin-Layer Chromatography. Immunoassays Applications Forensic. Microscopy Applications Forensic. Nucleic Acids Electrochemical Methods. Polymerase Chain Reaction. Spectrophotometry Oven/lew Biochemical Applications. 

Multiple forms of the a-L-fucosidase from human liver have been revealed by isoelectric focusing in polyacrylamide gel and in thin layers of Sephadex. The aggregation of various forms of the enzyme was investigated. 

Agarose gels for zone electrophoresis or isoelectric focusing of protein samples are mostly applied in the area of clinical diagnostics. For this purpose the gels are cast on horizontal glass plates or plastic films as thin layers of about 1 mm. 

Isoelectric focusing is most commonly carried out as horizontal thin layer electrophoresis, and is mostly used for separation of proteins, often as one of the separation modes in a two-dimensional separation. In lEF, which is an equilibrium process, the proteins will migrate to the location of their isoelectric point and stay. 

Polyacrylamide Gel Isoelectric Focusing Two well-established techniques that use polyacrylamide are the gel cyhnder and the thin layer. Here we discuss only the gel cylinder technique because it is simpler and easier to understand. Figure 13.15... 

D electrophoresis 2DE Combined application of electrophoresis and isoelectric focusing in a thin gel-based layer. 

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