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Profiling Approach for Biomarker Discovery using an Agilent HPLC-Chip Coupled with an Accurate-Mass Q-TOF LC/MS

Abstract: A profile-directed biomarker discovery approach involves initial differential expression analysis followed by targeted identification of differentially expressed proteins. In the present study, a very small amount (100 femtomole and 10 femtomole) of horseradish peroxidase was spiked into human plasma and a profile-directed approach was used for the identification of the spiked protein. Excellent reproducibility in retention time and intensity measurement, and outstanding accuracy in mass measurement and sensitivity over a wide dynamic range, were achieved using various Agilent platforms such as a Multiple Affinity Removal (MARS) column, a 1200 Series HPLC-Chip/MS and a 6520 Accurate-Mass Quadrupole Time-of-Flight (Q-TOF) LC/MS. Differential analyses using MassHunter Mass Profiler and MassHunter Mass Profiler Professional (MPP) software enabled the comparison between spiked and non-spiked plasma samples. Subsequent targeted MS/MS analysis on a Q-TOF LC/MS followed by a database search using Spectrum Mill helped identify the spiked protein.

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