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Molecular & Cellular Proteomics 1:490-499, 2002.
© 2002 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

High Throughput Peptide Mass Fingerprinting and Protein Macroarray Analysis Using Chemical Printing Strategies^*

Andrew J. Sloane, Janice L. Duff, Nicole L. Wilson, Parag S. Gandhi, Cameron J. Hill, Femia G. Hopwood, Paul E. Smith, Melissa L. Thomas, Robert A. Cole, Nicolle H. Packer, Edmond J. Breen, Patrick W. Cooley, David B. Wallace, Keith L. Williams and Andrew A. Gooley^,¶

Proteome Systems Limited, 1/35–41 Waterloo Rd., North Ryde, Sydney, New South Wales 2113, Australia
Microfab Technologies, Inc., Plano, Texas 75074

We describe a chemical printer that uses piezoelectric pulsing for rapid, accurate, and non-contact microdispensing of fluid for proteomic analysis of immobilized protein macroarrays. We demonstrate protein digestion and peptide mass fingerprinting analysis of human plasma and platelet proteins direct from a membrane surface subsequent to defined microdispensing of trypsin and matrix solutions, hence bypassing multiple liquid-handling steps. Detection of low abundance, alkaline proteins from whole human platelet extracts has been highlighted. Membrane immobilization of protein permits archiving of samples pre-/post-analysis and provides a means for subanalysis using multiple chemistries. This study highlights the ability to increase sequence coverage for protein identification using multiple enzymes and to characterize N-glycosylation modifications using a combination of PNGase F and trypsin. We also demonstrate microdispensing of multiple serum samples in a quantitative microenzyme-linked immunosorbent assay format to rapidly screen protein macroarrays for pathogen-derived antigens. We anticipate the chemical printer will be a major component of proteomic platforms for high throughput protein identification and characterization with widespread applications in biomedical and diagnostic discovery.

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