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Detection technologies in proteome analysis.

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  • 1Proteomics Section, Biosciences Department, Molecular Probes, Inc., 4849 Pitchford Avenue, Eugene, OR 97402-9165, USA. wayne.patton@probes.com

Abstract

Common strategies employed for general protein detection include organic dye, silver stain, radiolabeling, reverse stain, fluorescent stain, chemiluminescent stain and mass spectrometry-based approaches. Fluorescence-based protein detection methods have recently surpassed conventional technologies such as colloidal Coomassie blue and silver staining in terms of quantitative accuracy, detection sensitivity, and compatibility with modern downstream protein identification and characterization procedures, such as mass spectrometry. Additionally, specific detection methods suitable for revealing protein post-translational modifications have been devised over the years. These include methods for the detection of glycoproteins, phosphoproteins, proteolytic modifications, S-nitrosylation, arginine methylation and ADP-ribosylation. Methods for the detection of a range of reporter enzymes and epitope tags are now available as well, including those for visualizing beta-glucuronidase, beta-galactosidase, oligohistidine tags and green fluorescent protein. Fluorescence-based and mass spectrometry-based methodologies are just beginning to offer unparalleled new capabilities in the field of proteomics through the performance of multiplexed quantitative analysis. The primary objective of differential display proteomics is to increase the information content and throughput of proteomics studies through multiplexed analysis. Currently, three principal approaches to differential display proteomics are being actively pursued, difference gel electrophoresis (DIGE), multiplexed proteomics (MP) and isotope-coded affinity tagging (ICAT). New multiplexing capabilities should greatly enhance the applicability of the two-dimensional gel electrophoresis technique with respect to addressing fundamental questions related to proteome-wide changes in protein expression and post-translational modification.

PMID:
12015990
[PubMed - indexed for MEDLINE]
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