Fast MS/MS acquisition without dynamic exclusion enables precise and accurate quantification of proteome by MS/MS fragment intensity

Abstract

Most currently proteomic studies use data-dependent acquisition with dynamic exclusion to identify and quantify the peptides generated by the digestion of biological sample. Although dynamic exclusion permits more identifications and higher possibility to find low abundant proteins, stochastic and irreproducible precursor ion selection caused by dynamic exclusion limit the quantification capabilities, especially for MS/MS based quantification. This is because a peptide is usually triggered for fragmentation only once due to dynamic exclusion. Therefore the fragment ions used for quantification only reflect the peptide abundances at that given time point. Here, we propose a strategy of fast MS/MS acquisition without dynamic exclusion to enable precise and accurate quantification of proteome by MS/MS fragment intensity. The results showed comparable proteome identification efficiency compared to the traditional data-dependent acquisition with dynamic exclusion, better quantitative accuracy and reproducibility regardless of label-free based quantification or isobaric labeling based quantification. It provides us with new insights to fully explore the potential of modern mass spectrometers. This strategy was applied to the relative quantification of two human disease cell lines, showing great promises for quantitative proteomic applications.