A turning point in proteome analysis: Sample prefractionation via multicompartment electrolyzers with isoelectric membranes

Ben Herbert, Pier Giorgio Righetti*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    201 Citations (Scopus)

    Abstract

    Sample prefractionation, as obtained via multicompartment electrolyzers with isoelectric membranes, greatly enhanced the load ability, resolution and detection sensitivity of two-dimensional (2-D) maps in proteome analysis. This was demonstrated with different samples. In an Escherichia coli total cell extract, analysis by a 2-D map run in a pH 4-5 gradient showed many more spots when prefractionated, as compared with standard maps available in databases such as SWISS-2DPAGE. Analysis of human plasma in the pH 3-6 range showed an increase in the number of highly acidic proteins in the fractionated sample compared to whole plasma. With both samples no protein precipitation or smears occurred and much larger sample amounts could be loaded upon prefractionation, so that a large number of spots could be visualized by Coomassie staining, which is fully compatible with subsequent matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) analysis.

    Original languageEnglish
    Pages (from-to)3639-3648
    Number of pages10
    JournalElectrophoresis
    Volume21
    Issue number17
    DOIs
    Publication statusPublished - 2000

    Keywords

    • Isoelectric membranes
    • Prefractionation
    • Proteome
    • Two-dimensional maps

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