Examples of metalloproteinase-dependent proteolytic processing identified by TAILS and flow cytometry in stored platelets. Platelets were stored under blood-banking conditions for 7 days in the absence or presence of 10 μM marimastat. Two technical replicates from the same platelet unit were performed. Platelet proteomes from different time points and conditions were analyzed by 8-plex iTRAQ-TAILS analyses. Examples of the identified N termini are shown for glycoproteins 1bα (A), 1bβ (B), and VI (C). Peptides identified by TAILS are underlined; sequence positions of internal neo- and natural N termini are indicated by arrows. Peptide sequences show start and end position of the identified peptides and their N-terminal modifications, sequences, preceding amino acids, and iTRAQ_{DMSO/Marimastat} ratios on specified days. Graph inserts show time-resolved changes in relative peptide levels expressed as % of peptide levels on day 1 in the presence (dashed line) or absence (solid line) of marimastat. Processing of GPIb_α (D) and GPV (E) was confirmed by flow cytometry. Receptor density for platelets stored with 10 µM marimastat (black bars) or DMSO vehicle control (empty bars). Values for individual platelets units and mean values (GPIb_α, n = 7; GPV, n = 5) are shown. *P < .05 (paired t test with Bonferroni correction). Error bars indicate standard deviation of the mean.