Identification of carbohydrate peripheral epitopes important for recognition by positive-ion MALDI multistage mass spectrometry

[Display omitted] •Positive-ion MALDI-MSn of permethylated glycans for analysis of carbohydrate epitopes.•The peripheral epitopes assigned include the Lewis and blood-group related antigens.•Epitope ion and epitope spectrum were proposed for use in the assignment.•It can be used for both reducing and reduced sugars different from published methods.•It was used in conjunction with MALDI-MS profiling of N-glycan and O-glycan alditol. Carbohydrate sequences are important for various biological processes. It has recently been estimated to have 100,000–500,000 carbohydrate structures in mammalian glycome. However, the peripheral carbohydrate determinants on N- and O-glycoproteins, glycolipids, polysaccharides and secreted free sugars are limited in numbers. Among these blood-group-related antigens the ABO(H)- and Lewis-types are particularly important. Negative-ion MS/MS has been successfully used in assignment of these epitopes on free reducing sugars but cannot be applied to reduced sugars, e.g. O-glycans typically released from mucins as alditols, or in positive-ion detection of either reducing or reduced oligosaccharides. In the present study, we investigate the fragmentation features of permethylated reducing and reduced sugars under positive-ion conditions of multi-stage MALDI-MS, and propose the concept of epitope ion and epitope spectrum for determination of peripheral blood-group related epitopes on secreted human milk oligosaccharides and N-glycans as reducing sugars and O-glycans as reduced alditols in conjunction with MALDI-MS glycan profiling.