Protein identification platform utilizing micro dispensing technology interfaced to matrix-assisted laser desorption ionization time-of-flight mass spectrometry

Research output: Contribution to journalArticle

Abstract

An integrated protein microcharacterization/identification platform has been developed. The system has been designed to allow a high flexibility in order to tackle challenging analytical problems. The platform comprises a cooled microautosampler, an integrated system for microcolumn HPLC, and a capillary reversed-phase column that is interfaced to matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) system via a low internal volume flow-through microdispenser. The chromatographic separation is continuously transferred onto a MALDI target plate as discrete spots as the dispenser ejects bursts of droplets of the column effluent in a precise array pattern. A refrigerated microfraction collector was coupled to the outlet of the flow-through microdispenser enabling enrichment and re-analysis of interesting fractions. The use of target plates pre-coated with matrix simplified and increased the robustness of the system. By including a separation step prior to the MALDI-TOF-MS analysis and hereby minimizing suppression effects allowed us to obtain higher sequence coverage of proteins compared to conventional MALDI sample preparation methodology. Additionally, synthetic peptides corresponding to autophosphorylated forms of the tryptic fragment 485-496 (ALGADDSYYTAR) of tyrosine kinase ZAP-70 were identified at sensitivities reaching 150 amol. Copyright (C) 2000 Elsevier Science B.V.

Details

Authors
Organisations
External organisations
  • AstraZeneca
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Chemical Sciences

Keywords

  • Instrumentation, Interfaces, LC-MS, Matrix-assisted laser desorption ionization mass spectrometry, Proteins
Original languageEnglish
Pages (from-to)99-110
Number of pages12
JournalJournal of Chromatography A
Volume886
Issue number1-2
StatePublished - 2000 Jul 21
Publication categoryResearch
Peer-reviewedYes