Rapid N-Glycan Release from Glycoproteins using Immobilized Glycosylase Columns

摘要

N-glycosylation profiling of glycoprotein biotherapeutics is an essential step in each phase of product development in the biopharmaceutical industry. For example, during clone selection hundreds of clones should be analyzed quickly from limited amounts of samples. On the other hand, identification of disease related glycosylation alterations can serve as early indicators for various pathological conditions in the biomedical field. We describe an improved packed bed column PNGase F functionalized column reactor. The reactor may be packed into a pipette tip column. In some embodiments, a second column or mixed stationary phase may be packed into the column to capture and purify the cleaved glycan prior to analysis. Complete N-glycan removal can be obtained in 10 minutes from all major N-linked glycoprotein types. The approach can be readily applied to automated sample preparation systems, such as liquid handling robots.