Microfluidic Liquid Chromatography System for Proteomic Applications and Biomarker Screening

A microfluidic liquid chromatography (LC) system for proteomic investigations that integrates all the necessary components for stand-alone operation, i.e., pump, valve, separation column, and electrospray interface, is described in this paper. The overall size of the LC device is small enough to ena...

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Bibliographic Details
Published in:	Analytical chemistry (Washington) Vol. 78; no. 15; pp. 5513 - 5524
Main Authors:	Lazar, Iulia M, Trisiripisal, Phichet, Sarvaiya, Hetal A
Format:	Journal Article
Language:	English
Published:	Washington, DC American Chemical Society 01-08-2006
Subjects:	Analytical chemistry Biochemistry Biomarkers, Tumor - analysis Breast cancer Cell Line, Tumor Chemistry Chromatographic methods and physical methods associated with chromatography Chromatography Chromatography, Liquid - instrumentation Chromatography, Liquid - methods Exact sciences and technology Fluid dynamics General, instrumentation Humans Mass Spectrometry - instrumentation Mass Spectrometry - methods Medical screening Microfluidic Analytical Techniques - instrumentation Microfluidic Analytical Techniques - methods Other chromatographic methods Particle Size Proteomics Proteomics - instrumentation Scientific apparatus & instruments Semiconductors Sensitivity and Specificity Time Factors Cation exchange Performance evaluation Filtering Peptides Glass Biological marker Instrumentation Liquid chromatography System on a chip Digestion Electrospray Protein Ion exchange chromatography Separation column Sample preparation Electroosmosis Proteomics Microfluidics
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Summary:	A microfluidic liquid chromatography (LC) system for proteomic investigations that integrates all the necessary components for stand-alone operation, i.e., pump, valve, separation column, and electrospray interface, is described in this paper. The overall size of the LC device is small enough to enable the integration of two fully functional separation systems on a 3 in. × 1 in. glass microchip. A multichannel architecture that uses electroosmotic pumping principles provides the necessary functionality for eluent propulsion and sample valving. The flow rates generated within these chips are fully consistent with the requirements of nano-LC platforms that are routinely used in proteomic applications. The microfluidic device was evaluated for the analysis of a protein digest obtained from the MCF7 breast cancer cell line. The cytosolic protein extract was processed according to a shotgun protocol, and after tryptic digestion and prefractionation using strong cation exchange chromatography (SCX), selected sample subfractions were analyzed with conventional and microfluidic LC platforms. Using similar experimental conditions, the performance of the microchip LC was comparable to that obtained with benchtop instrumentation, providing an overlap of 75% in proteins that were identified by more than two unique peptides. The microfluidic LC analysis of a protein-rich SCX fraction enabled the confident identification of 77 proteins by using conventional data filtering parameters, of 39 proteins with p < 0.001, and of 5 proteins that are known to be cancer-specific biomarkers, demonstrating thus the potential applicability of these chips for future high-throughput biomarker screening applications.
Bibliography:	istex:63A2FF064697DB60CA649ABF3A7B6EA1E2509A57 ark:/67375/TPS-R8WC5L1M-3 ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	0003-2700 1520-6882
DOI:	10.1021/ac060434y