Consistent analysis pipeline. Shown is a schematic presentation of consistent analysis pipeline for the identification of high-quality N-linked glycosites using glycopeptide capture and LC-MS/MS. LC, liquid chromatography; MS/MS, tandem mass spectrometry.

Comparison of number of N-linked glycosites commonly or uniquely detected from plasma and tissues/cells. Shows the overlap of N-linked glycosites identified in plasma with tissues or cells.

Representative output of N-linked glycosites from database using UniPep. UniPep contains all proteins in the International Protein Index (IPI) database (version 2.28) with at least one N-linked glycosite and allows users to view all the predicted and identified N-linked glycosites from a specific protein. For each potential N-linked glycoprotein, a user can see the protein annotation, predicted subcellular location, and sequence(s) of predicted N-linked glycosites(s). The uniqueness of a peptide in the database is also presented as number of hits in the database, and for those peptides present in multiple proteins, linkage to other proteins in the database is provided. If any predicted N-linked glycosite was identified in the dataset from this study, then it is listed as an identified peptide with PeptideProphet score [39] to allow researchers to evaluate the confidence of the identification. The sequence of the proteins queried is overlaid with different sequence features such as the N-linked glycosites, the predicted and identified peptide sequences, signal peptide, and transmembrane segment(s) [21].

Ratio of identified N-linked glycosites identified from proteins predicted as intracellular proteins and extracellular proteins. The extracellular proteins include secreted proteins, cell surface proteins, and transmembrane proteins. The findings are expressed a function of probability stringency.