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. 2017 Jan 6;8(8):13413–13427. doi: 10.18632/oncotarget.14542

Figure 1. Workflow for the integrated analysis of global protein expression and glycosite-containing peptides.

Figure 1

In forward labeling experiments, equal amounts of protein obtained from “heavy labeled” NCI/ADR-RES and “light labeled” OVCAR8 cells were combined and followed by in-solution tryptic digestion. Mixed labeled peptide samples were then separated into two aliquots. One aliquot was enriched for glycosite-containing peptides using HILIC and the other aliquot was used for high pH reverse phase C18 fractionation followed by the analysis of global protein expression. Finally, LC-MS/MS analysis of peptides were performed using a LC-MS/MS system. In reverse labeling experiments, “heavy labeled” OVCAR8 and “light labeled” NCI/ADR-RES cells were treated in parallel using procedures same to those described above. Forward labeling and reverse labeling experiments were conducted twice using independent biological replicates.