Analysis of 8-aminonaphthalene-1,3,6-trisulfonic acid labelled N-glycans by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

Fluorophore-assisted carbohydrate electrophoresis (FACE) is a fast and effi cient analytical method which is now widely used in glycobiology for the se paration and quantification of free or glycoprotein-released oligosaccharid es. However, since identification by FACE of N-glycan structures is only ba sed on their electrophoretic mobility after labelling with 8-aminonaphthale ne-1,3,6-trisulfonic acid (ANTS), comigration of derived glycans on gel cou ld occur which may result in erroneous structural assignments. As a consequ ence, a protocol was developed for the fast and efficient matrix-assisted l aser desorption/Ionisation Time-of-flight (MALDI-TOF) mass spectrometric an alysis of ANTS-labelled N-glycans, N-Glycans were isolated from plant and m ammalian glycoproteins, reductively aminated with the charged fluorophore 8 -aminonaphthalene-1, 3, 6-trisulfonic acid (ANTS) and separated using high resolution polyacrylamide gel electrophoresis, The ANTS-labelled glycans we re eluted from FACE gel slices and then analysed by MALDI-TOF mass spectrom etry in negative ion mode. Using 3-aminoquinoline containing 2.5 mM citrate NH4+ as matrix, neutral N-linked N-glycans, as well as labelled sialylated oligosaccharides, were found to be easily detected in the 2-10 picomole ra nge giving rise to [M - H](-) ions, Copyright (C) 2000 John Whey & Sons, Lt d.