Optimal conditions to radiolabel (3H or 14C) aminosugar-containing glycosphingolipids by de-N-acetylation and re-N-acetylation.

A Rebbaa, J Portoukalian

Index: Biochim. Biophys. Acta 1244(2-3) , 418-22, (1995)

Full Text: HTML

Abstract

The optimal conditions were examined for selective re-N-acetylation with 14C or 3H.acetic anhydride of de-N-acetylated aminosugar-containing glycosphingolipids. Re-N-acetylation, which is nearly quantitative within 10 minutes in methanol, occurs selectively up to a maximal 100% yield when using a molar ratio of 5 mol of acetic anhydride per mole of aminosugar present in the glycosphingolipid. Above this molar ratio, it was observed some O-acetylation of carbohydrates which could be removed by mild alkali treatment. The method allows the choice of 14C- or 3H-labeling of glycosphingolipids with a final specific radioactivity which depends solely on the one of acetic anhydride. The binding of specific antibodies to glycosphingolipids, which was abolished upon de-N-acetylation, was again detectable after re-N-acetylation with radioactive acetic anhydride, suggesting that the native structures were recovered. This procedure of radiolabeling offers safety, rapidity and broad applicability to alkali-stable aminosugar-containing glycosphingolipids.