Improvement of Quantitative Measurements in Multiplex Proteomics Using High-Field Asymmetric Waveform Spectrometry

Quantitative
proteomics using isobaric reagent tandem mass tags
(TMT) or isobaric tags for relative and absolute quantitation (iTRAQ)
provides a convenient approach to compare changes in protein abundance
across multiple samples. However, the analysis of complex protein
digests by isobaric labeling can be undermined by the relative large
proportion of co-selected peptide ions that lead to distorted reporter
ion ratios and affect the accuracy and precision of quantitative measurements.
Here, we investigated the use of high-field asymmetric waveform ion
mobility spectrometry (FAIMS) in proteomic experiments to reduce sample
complexity and improve protein quantification using TMT isobaric labeling.
LC–FAIMS–MS/MS analyses of human and yeast protein digests
led to significant reductions in interfering ions, which increased
the number of quantifiable peptides by up to 68% while significantly
improving the accuracy of abundance measurements compared to that
with conventional LC–MS/MS. The improvement in quantitative
measurements using FAIMS is further demonstrated for the temporal
profiling of protein abundance of HEK293 cells following heat shock
treatment.