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Identification of Protein Spots in Silver-stained Two-dimensional Gels by MALDI-TOF Mass Peptide Map Analysis.
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2000
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Silver-stained Two-dimensional GelsMolecular BiologyPathologyPeptide ScienceProtein PurificationProteomic TechnologyBioanalysisVenomicsAnalytical ChemistryPipette TipClinical ChemistryAnalytical BiotechnologyProteomicsChromatographyBiochemistryProtein SpotsNatural SciencesPeptide LibraryMass SpectrometryProtein Mass SpectrometryProtein EngineeringProtein DatabaseMedicine
Protein spots in silver-stained two dimensional gels were analyzed and identified by employing an improved procedure of mass spectrometric peptide mapping, including i) In-gel reduction, alkylation and enzymatic digestion ii) Extraction and desalting by using a pipette tip containing a small C18 micro-column (ZipTip(TM)) iii) Direct MALDI-TOF mass analysis and protein database searching. The results demonstrate that single silver-stained protein spots in a 2-DE-gel could be identified rapidly by this procedure and the use of the ZipTip(TM) pipette tip could increase evidently the sensitivity of the MALDI-TOF analysis. By using this the procedure, 10 protein spots in a silver-stained gel of 2-DE of crude venom of the spider S.Huwena were analyzed and identified.