A solid-phase assay for quantitative analysis of sulfated glycosaminoglycans at the nanogram level. Application to tissue samples

A sensitive and accurate solid-phase methodology for the quantitative analy sis of glycosaminoglycans is described. Chondroitin-4-sulfate (CSA) was lab elled with biotin hydrazide after the reaction of its carboxyl groups with it in the presence of carbodiimide. Polystyrene plates modified with sequen tial reaction with glutaraldehyde (GH) and spermine to possess amino groups were used to immobilize electrostatically the biotin labelled CSA. Exogeno usly added sulfated glycosaminoglycans (GAGS) [variously sulfated chondroit in sulfates and heparan sulfate (HS)] were found to compete to this immobil ization in a concentration dependent mode, within a concentration range fro m 10 up to 300 ng/ml. Glycosaminoglycan-derived oligosaccharides competed t o a degree similar to that of intact molecules. Hyaluronan (HA) and keratan sulfate (KS) did not compete the immobilization. The procedure was applied for the rapid and reproducible determination of the sulfated glycosaminogl ycans in proteinase digests of small tissue samples or cell cultures with h igh sensitivity and accuracy. (C) 1999 Elsevier Science B.V. All rights res erved.