Web Release Date: December 23,
Membrane Proteome Analysis of Microdissected Ovarian Tumor Tissues Using Capillary Isoelectric Focusing/Reversed-Phase Liquid Chromatography-Tandem MS
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Calibrant Biosystems, 910 Clopper Road, Suite 220N, Gaithersburg, Maryland 20878, Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland 20742, Molecular Pathogenesis Unit, Surgical Neurology Branch, National Institute of Neurological Disorders and Stroke, Bethesda, Maryland 20892, Department of Pathology, University of Arizona, Tucson, Arizona 85724, and Department of Mechanical Engineering and Bioengineering Program, University of Maryland, College Park, Maryland 20742
Received for review August 29, 2006. Accepted November 16, 2006.
Abstract:
This work expands our tissue proteome capabilities from the analysis of soluble proteins in previous studies to the examination of membrane proteins within the pellets of enriched and selectively isolated tumor cells procured from microdissected tissue specimens. The pellets of targeted ovarian tumor cells are treated by two different membrane protein extraction methods, including the use of detergent and organic solvent. The detergent-based membrane protein preparation protocol not only extracts proteins effectively from cell pellets but also is compatible with subsequent proteome analysis using combined capillary isoelctric focusing/nano reversed-phase liquid chromatography separations coupled with nano electrospray ionization mass spectrometry. Among proteins identified from an amount of pellet equivalent to 20 000 cells, 773 proteins are predicted to contain one or more transmembrane domains, corresponding to 22% membrane proteome coverage within the SwissProt Human protein sequence entries.
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