CD24 is a glycosyl-phosphatidyl-inositol linked glycoprotein expressed in a broad range of cell types and is heavily glycosylated. It has been found to be over expressed in cancers and tumors and is also a costimulatory molecule. Therefore, this study was carried out to define the structures of the carbohydrates associated with the CD24 recombinant protein. The CD24 glycoprotein's oligosaccharides were released by chemical and enzymatic means prior to being analyzed by MALDI-TOF-MS. The results obtained showed that CD24 is both N- and O-glycosylated. The major oligosaccharides were found to be Neu5Acα-2,3/6Galβ-1,3GalNAc, NeuAc(2)Gal β-1,3GalNAc(1) (O-glycans), GalNAc(2)GlcNAc(2)Man(3)Fuc(1), Gal(1)GalNAc(2)GlcNAc(2)Man(3)Fuc(1), and Gal(2)GalNAc(2)GlcNAc(2)Man(3)Fuc(1) (N-glycans). The results showed that Neu5Acα-2,3/6Galβ-1,3GalNAc (sialyl-tumor antigen, sT), a cancer-associated carbohydrate, was the most abundant glycan associated with CD24. This result raised the intriguing possibility that CD24 may be a major carrier of the sialyl-T abundantly found in cancer cells.