Web Release Date: January 6,
Integrated Capillary Isoelectric Focusing/Nano-reversed Phase Liquid Chromatography Coupled with ESI-MS for Characterization of Intact Yeast Proteins
and
Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland 20742, Calibrant Biosystems, 7507 Standish Place, Rockville, Maryland 20855, and Department of Mechanical Engineering and Bioengineering Program, University of Maryland, College Park, Maryland 20742
Received July 22, 2004
Abstract:
An integrated protein concentration/separation platform, combining capillary isoelectric focusing (CIEF) with nano-reversed phase liquid chromatography (nano-RPLC), is developed to provide significant protein concentration and high resolving power for the analysis of complex protein mixtures. Upon completion of protein focusing, the proteins are sequentially and hydrodynamically loaded into individual trap columns using a group of microinjection and microselection valves. Repeated pro-tein loadings and injections into trap columns are carried out automatically until the entire CIEF cap-illary content is sampled and fractionated. Each CIEF fraction "parked" in separate trap columns is further resolved using nano-RPLC, and the eluants are analyzed using electrospray ionization-mass spectrometry.
Keywords: capillary isoelectric focusing 
ESI-MS intact proteins proteomics reversed-phase liquid
chromatography
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