Segmentation of Precursor Mass Range Using Tiling Approach Increases Peptide Identifications for MS1-Based Label-Free Quantification

Label-free quantification is a powerful tool for the measurement of protein abundances by mass spectrometric methods. To maximize quantifiable identifications, MS1-based methods must balance the collection of survey scans and fragmentation spectra while maintaining reproducible extracted ion chromatograms (XIC). Here we present a method which increases the depth of proteome coverage over replicate data-dependent experiments without the requirement of additional instrument time or sample prefractionation. Sampling depth is increased by restricting precursor selection to a fraction of the full MS1 mass range for each replicate; collectively, the m/z segments of all replicates encompass the full MS1 range. Although selection windows are narrowed, full MS1 spectra are obtained throughout the method, enabling the collection of full mass range MS1 chromatograms such that label-free quantitation can be performed for any peptide in any experiment. We term this approach binning or tiling depending on the type of m/z window utilized. By combining the data obtained from each segment, we find that this approach increases the number of quantifiable yeast peptides and proteins by 31% and 52%, respectively, when compared to normal data-dependent experiments performed in replicate.