Journal
NATURE PROTOCOLS
Volume 7, Issue 5, Pages 882-890Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nprot.2012.036
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Funding
- Boehringer Ingelheim
- Christian Doppler Research Association
- Austrian Proteomics Platform within the Austrian GenomeResearch program (GEN-AU)
- Austrian Science Fund via the Special Research Program Chromosome Dynamics [SFB-F3402]
- European Commission
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The majority of proteome-wide studies rely on the high separation power of two-dimensional liquid chromatography-tandem mass spectrometry (2D LCLC-MS/MS), often combined with protein prefractionation. Alternative approaches would be advantageous in order to reduce the analysis time and the amount of sample required. On the basis of the recent advances in chromatographic and mass spectrometric instrumentation, thousands of proteins can be identified in a single-run LCLC-MS/MS experiment using ultralong gradients. Consequently, the analysis of simple proteomes or clinical samples in adequate depth becomes possible by performing single-run LCLC-MS/MS experiments. Here we present a generally applicable protocol for protein analysis from unseparated whole-cell extracts and discuss its potential and limitations. Demonstrating the practical applicability of the method, we identified 2,761 proteins from a HeLa cell lysate, requiring around 10 h of nanoLCLC-MS/MS measurement time.
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