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Fully Automated Nano-Electrospray Coupled with a Finnigan LCQ Deca XP Plus for Sub-fmol Proteomic Analysis

Dirk Chelius, Terry Zhang, and Ken Miller;
Thermo Electron Corporation, San Jose, CA

Key Words Sensitivity Proteomics Finnigan LCQ Deca XP Plus PepFinder Kit Finnigan Surveyor HPLC LC/MS/MS

Introduction
Detection limits of mass spectrometers have been pushed downward to levels no one would have thought possible only a decade ago. Sensitivity gains are driven by instrument hardware improvements and sophisticated methods of sample introduction to the mass analyzer. Nanoflow ESI is a very popular method in proteomics to achieve maximum sensitivity and separation of peptides. Peptides are separated on a capillary reversed-phase column (75 μm internal diameter [ID]), which is placed directly in front of the mass spectrometer to limit peak broadening after the column. For many reasons, use of these small ID columns can be complex. Engineering an optimal solution to these practical issues can be very time consuming. The optimal flow rate for a 75 μm ID column is between 100 and 200 nL/min, often requiring flow splitting. At such low flow rates, loading of a sample onto the column can take a long time: at a flow rate of 100 nL/min it takes 200 minutes to elute a 20 μL sample loop. Researchers have attempted to overcome this problem by loading the sample onto the column during the packing of the column, but this method is very time consuming and each column can only be used one time.Alternative approaches are the loading of the samples at a higher flow rate or the use of a peptide trap. The peptide trap has several advantages compared to other loading techniques; peptides can be loaded at a higher flow rate, desalted and washed. This combination allows high sample throughput and maximal sensitivity of detection. This plumbing configuration is the basis of the PepFinder Kit. Designed for easy installation and maximum productivity, all PepFinder Kit componen
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