β-O-linked N-acetylglucosamine (O-GlcNAc) is an abundant post-translational modification occuring on serine and threonine residues. The modification has been detected on a variety of nuclear and cytoplasmic proteins involved in diverse cellular functions. Studies to elucidate the role of O-GlcNAc-ylation have been hampered by the lack of a sensitive, specific assay to identify or purify proteins bearing O-GlcNAc.