Characterization of Glycosylated Proteins at Subunit Level by HILIC/MS

Hydrophilic interaction chromatography (HILIC) coupled to mass spectrometry (MS) is considered as the reference analytical technique for glycans profiling, especially for the characterization of glycosylated protein therapeutics such as monoclonal antibodies (mAbs) and mAbs-related products. Although HILIC/MS is mainly known to profile enzymatically released and fluorescently labeled N-glycans, the recent commercialization of new widepore HILIC amide bonded stationary phases packed with sub-2 μm particles has allowed for remarkable separations also at the subunit level. Here, we describe a simple protocol to perform the mAb glycans profiling at subunit level by HILIC/MS.