Trace LC/MS quantitative analysis of polypeptide biomarkers: impact of 1-D and 2-D chromatography on matrix effects and sensitivity.
We investigated the impact of one dimension (single reverse phase (RP) column) and two dimension (two different RP columns) chromatographic methods on SIM (MS) and multiple reaction monitoring (MRM; MS/MS) performance from human plasma. We find that MRM analysis is clearly preferable for 1-D applications; however, implementation of SIM detection in conjunction with 2-D separation technique resulted in an over 60-fold increase in analyte peak area and improved S/N compared to MRM for our analyte, human C-peptide. Implementation of a 2-D RP-RP technique with SIM detection is capable of eliminating matrix effects and greatly increases signal response and data quality. For two large peptides in complex biological samples, we found that a 2-D approach performed better than high quality sample preparation together with 1-D chromatography and MRM, even on a high-end mass spectrometer.
Rogatsky
Eduard
E
Core Analytical Laboratory at General Clinical Research Center, Albert Einstein College of Medicine, Yeshiva University, Bronx, NY 10461, USA. rogatsky@aecom.yu.edu
Tomuta
Vlad
V
Jayatillake
Harsha
H
Cruikshank
Greg
G
Vele
Louis
L
Stein
Daniel T
DT
D015415:Biomarkers / Q000032:analysis / Q000737:chemistry; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods; D005934:Glucagon / Q000737:chemistry; D006801:Humans; D010455:Peptides / Q000032:analysis* / Q000737:chemistry*; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000295:instrumentation* / Q000379:methods*
D015415:Biomarkers; D010455:Peptides; D005934:Glucagon