The Tn antigen, a simple glycoconjugate composed of GalNAc alpha-O-linked to Ser or Thr, is expressed in nearly every carcinoma. In cancer cells, MUC-1 is the most common protein that bears the Tn epitope and is the second most common tumor-associated antigen for cancer vaccine targets. The glycosylation pattern of MUC-1 diverges significantly in cancer cells, showing an increase in the frequency of the Tn antigen motif. Understanding normal patterns of glycosylation can assist in the determination of glycosites altered in tumor settings and help identify predictive biomarkers. Thus, O-glycosylation of MUC-1 was evaluated after SmE digestion, similar to the strategy used for MUC-16. Figure compares the glycan compositions identified based on analysis of HCD, EThcD, and UVPD spectra. MUC-1 demonstrates more varied O-glycosylation than MUC-16 or TIM-1, revealing double the number of glycan compositions. For the SmE digests of TIM-1 and MUC-16, there were some glycopeptides suspected to contain N-glycolylneuraminic acid (Neu5Gc), but the MS/MS spectra did not meet the criteria to be Table . Glycoforms for MUC-16 identified by UVPD. Glycoforms are grouped by their composition, the glycopeptide for which they were identified, and then listed in order of glycosite. Multiple glycoforms were identified at each site. Repeated identification of identical glycopeptides is denoted by the number in the right-most column. Core linkages were assumed (β1,3 for Gal to GalNAc, β1,6 for GlcNAc to GalNAc) and α2,6 linkage was assumed for Neu5Ac to GalNAc. A check mark refers to the presence of that linkage in the glycan and an x means that that linkage was not detected for the glycan. In the "Linkages" column, A refers to sialic acid linkages, H refers to linkages of galactose, and N refers to GlcNAc linkages. a glycan position relative to complete protein sequence for MUC-16. Full table is given as Table . considered valid hits. However, in the MS/MS analysis of MUC-1 there were four glycan types containing one or two Neu5Gc moieties identified. Owing to the more extensive fragmentation produced by UVPD and EThcD, these MS/MS methods outperform HCD for identification of these glycans. Additionally, more abundant fucosylated glycopeptides were observed for MUC-1 compared to abundances found for TIM-1 and MUC-16. The number of glycopeptides identified by UVPD, HCD, and EThcD for MUC-1 are compared in Figure where a significant number of unique identifications are found for UVPD.