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Quantitative validation of different protein precipitation methods in proteome analysis of blood platelets
Author(s) -
Zellner Maria,
Winkler Wolfgang,
Hayden Hubert,
Diestinger Michael,
Eliasen Maja,
Gesslbauer Bernd,
Miller Ingrid,
Chang Martina,
Kungl Andreas,
Roth Erich,
Oehler Rudolf
Publication year - 2005
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200410262
Subject(s) - trichloroacetic acid , chemistry , proteome , protein precipitation , precipitation , isoelectric point , chromatography , proteomics , ethanol precipitation , isoelectric focusing , platelet , gel electrophoresis , electrophoresis , blood proteins , two dimensional gel electrophoresis , biochemistry , mass spectrometry , extraction (chemistry) , biology , enzyme , immunology , physics , meteorology , gene
Abstract For the preparation of proteins for proteome analysis, precipitation is frequently used to concentrate proteins and to remove interfering compounds. Various methods for protein precipitation are applied, which rely on different chemical principles. This study compares the changes in the protein composition of human blood platelet extracts after precipitation with ethanol (EtOH) or trichloroacetic acid (TCA). Both methods yielded the same amount of proteins from the platelet preparations. However, the EtOH‐precipitated samples had to be dialyzed because of the considerable salt content. To characterize single platelet proteins, samples were analyzed by two‐dimensional fluorescence differential gel electrophoresis. More than 90% of all the spots were equally present in the EtOH‐ and TCA‐precipitated samples. However, both precipitation methods showed a smaller correlation with nonprecipitated samples (EtOH 74.9%, TCA 79.2%). Several proteins were either reduced or relatively enriched in the precipitated samples. The proteins varied randomly in molecular weight and isoelectric point. This study shows that protein precipitation leads to specific changes in the protein composition of proteomics samples. This depends more on the specific structure of the protein than on the precipitating agent used in the experiment.