Electrophoresis 2-D PAGE

Significant improvements in the technologies of high-resolution two-dimensional Polyacrylamide Gel Electrophoresis (2-D PAGE) and Mass Spectrometry (MS) have marked the start of proteome analysis. Proteomics permits the analysis of thousand of proteins simultaneously, and have the potential to identify markers for early detection, classification and prognosis of diseases, as well as pinpointing targets for improved treatment outcomes [42]. 

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 Rotofor has been incorporated into two-dimensional purification schemes based on the principles of 2-D PAGE. Fractions collected from the Rotofor were further purified by 2-D PAGE62 or by preparative polyacrylamide gel electrophoresis.63-65 Highly purified proteins were obtained by this scheme, even low-abundance proteins, to allow for multiple analyses and for use as antigens. 

An ingenious combination, known as two-dimensional gel electrophoresis (2-D gels), allows for enhanced separation by using isoelectric focusing in one dimension and SDS-PAGE run at 90° to the first (Figure 5.13). 

Combination of mass spectrophotometry with other analytical methods, e.g., 2-D electrophoresis, SDS-PAGE, capillary zone electrophoresis (CZE), HPCL, and FPLC or a combination of spectrophotometers into the so-called MS-MS tandems, provides additional information and enables us to identify compounds at even much lower concentrations. 

Figure 4.2 Schematic overview of two protein identification strategies commonly followed in proteomics. Protein samples are separated by either two-dimensional (2-D) or one-dimensional (1 -D) polyacrylamide gel electrophoresis (PAGE). In both strategic tracks, proteins are converted into a set of peptides by enzymatic digestion (e.g., with trypsin) prior to MS analysis. Peptide mass fingerprinting (PMF) by MALDl MS is predomi-...

D electrophoresis (lEF/SDS PAGE) High resolution separation/fractionation based on pi and m.w., long and complex procedure. Preparative/analytical... 

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