An integrated stage-tip-based glycomic and glycoproteomic approach for simple and rapid N-glycosylation profiling of glycoproteins

Glycosylation is a post-translational modification that plays an active role in many cellular events. It also regulates many functions of proteins. Monoclonal antibody-derived drugs are used to treat many diseases, and glycosylation affects the activity of such drugs developed. On the other hand, N-glycans may change in certain diseases. Therefore, rapid and efficient bioanalytical methods are needed for N-glycosylation profiling. The study aimed to develop an integrated stage-tip application for the simple and rapid N-glycosylation profiling of glycoproteins. A fast and inexpensive N-glycosylation profiling was achieved by integrating all glycoproteomic and glycomic sample preparation steps into a stage-tip. The glycomic approach of the integrated stage-tip reduces the N-glycan profiling time from 2 days to approximately 2.5 hours. It also allows the profiling of immunoglobulin G (IgG) N-glycopeptides directly from human plasma. In addition, N-glycosylation profiling can be done in the developed method without sorbents, C₁₈ or others, such as strong-cation exchange, at the glycopeptide level.