Glycosylation is important role in signal transduction between cells, and various biological processes. Biosimilar is a clone of a biotherapeutic products that have been developed by leveraging biotechnology, such as gene recombination and cell culture techniques. Therefore, glycosylation analysis is necessary during various stages of a biotherapeutic product’s lifecycle and one of the most critical tools in the characterization of biotherapeutic proteins.
In this study, N-glycans are enzymatically released by N-glycosidase F(PNGase F). O-glycans are chemically released by β-elimination. they are purified by SPE(solid-phase extraction)-PGC. The permethylation of sialylated glycans results in the conversion of all hydroxyl groups into methoxyl groups and serves to render glycans hydrophobic which may increase the signal intensity in mass measurement. We revealed the characterization of N- and O-glycans released from glycoproteins by using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and tandem mass spectrometry (MALDI-MS/MS) with 2,5-dihydroxybenzoic acid (2,5-DHB) as the matrix.