FIGURE 1.

Overview of Isobaric Tag Workflow. The standard protocol involves proteins that undergo reduction, alkylation, and digestion to generate peptides. For the case of DiLeu 4‐plex, up to four samples are labeled with different channels, and are combined at equal concentrations prior to clean up steps. DiLeu‐labeled samples are analyzed via LC‐MS/MS where at the MS1 level, peptides from the pooled samples will appear as a single composite peak, which after fragmentation will show distinct reporter ion masses between m/z 114 and 118. The intensity of the reporter ion will indicate the relative amount of peptide in the mixture. All LC‐MS/MS data will undergo further data processing for downstream analysis