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Glycobiology Advance Access published online on May 28, 2003
Glycobiology, doi:10.1093/glycob/cwg082
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Submitted on February 21, 2003
Revised on April 14, 2003
Accepted on May 8, 2003

© 2003 Oxford University Press

ORIGINAL ARTICLES

Improved and simple micro assay for sulfated glycosaminoglycans quantification in biological extracts and its use in skin and muscle tissue studies

Isabelle Barbosa 1, Stéphanie Garcia 1, Véronique Barbier-Chassefière 2, Jean-Pierre Caruelle 1, Isabelle Martelly 1, Dulce Papy-García 1*

1 Laboratoire de Recherche sur la Croissance, la Réparation et la Régénération Tissulaires (CRRET), CNRS FRE-2412, Université Paris 12-Val de Marne, 61 Avenue du Général de Gaulle, 94010 Créteil cedex, France
2 Société OTR-3 SARL, 33, avenue Pierre Brossolette, 94000 Créteil cedex, France

* To whom correspondence should be addressed. E-mail: papy{at}univ-paris12.fr.

Abstract

This report describes a simple and selective procedure used for direct measurement of sulfated glycosaminoglycans in biological samples and its application to the determination of glycosaminoglycans during tissue regeneration and myogenic differentiation. We describe a modified procedure of previous glycosaminoglycan assays that has improved specificity, reproducibility and sensibility. The assay is based on the ability of sulfated glycosaminoglycans to bind the cationic dye 1,9-dimethylmethylene blue. We describe conditions that allow isolation of the glycosaminoglycan/dye complex. This complex was dissociated and the optical density measurement of the dissociated dye permitted quantification of glycosaminoglycans in biological samples. Applied to the study of myogenic cell differentiation in vitro, muscle repair and skin ulceration, this method revealed significant modifications in the patterns of expression of different sulfated glycosaminoglycans in these tissues. In particular, application of the method after nitrous acid treatment revealed that heparan sulfate and chondroitin sulfate ratio changed during muscle regeneration process.


1,9-dimethylmethylene blue, glycosaminoglycans, cell culture, skeletal muscle, skin
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