alcian-blue and Mucopolysaccharidoses

The dimethylmethylene blue (DMB)-based screening procedure for mucopolysaccharidosis and the alcian blue (AB)-based procedure both measure glycosaminoglycan (GAG) content directly in urine. We compared the two procedures. Absorbance per microgram of GAG for DMB was 25 times that obtained with AB, resulting in a requirement for 10 times smaller sample volumes for the former. Recoveries of added heparan sulphate for the AB assay in the absence and presence of added protein (2.5 g/L) were comparable (78-95% and 75-111%), as was the case with the DMB assay. Here, recoveries were generally better (94-103% and 96-100%). The coefficient of correlation (CC) between the two methods measured on 86 reference urine samples was 0.861. For mucopolysaccharidosis urines the CC was higher (0.928). Age-dependent reference values were comparable. Performance of the assays was compared by measurement of GAG content in 24 mucopolysaccharidosis urines. Standard deviation scores (measured value minus mean of reference values divided by standard deviation in reference values) and probabilities for classification in the patient group were calculated and compared. Especially for urines from patients with less distinctly increased GAG content (Sanfilippo, Scheie), scores and probabilities were higher with the DMB assay.

Topics: Adolescent; Adult; Alcian Blue; Child; Child, Preschool; Glycosaminoglycans; Humans; Infant; Methylene Blue; Middle Aged; Mucopolysaccharidoses; Sensitivity and Specificity

The trypanocidal drug suramin was previously reported to induce mucopolysaccharidosis in rats; apart from the biochemical demonstration of increased tissue concentrations of sulfated glycosaminoglycans (GAGs), a strongly positive staining reaction with the cationic dye Alcian Blue was taken as indicating GAG-storage (Constantopoulos et al. 1983). The purpose of the present report is to point out a methodical pitfall. In model experiments it was found that suramin itself, being a polysulfated compound, gives a strongly positive reaction with Alcian Blue at pH 1. It is known that suramin is accumulated in the lysosomes and that high drug concentrations are retained in the tissues for weeks. Therefore a positive staining reaction with Alcian Blue observed in a given cell cannot be conclusively attributed to the storage of sulfated GAGs as has been done in the past. The present report may be a warning that, in the case of the suramin-induced animal model of mucopolysaccharidosis, the usual histochemical strategy, i.e. staining with cationic dyes, is not suitable for analysing the cellular distribution pattern of GAG-storage, since the inducing drug by itself reacts with the indicator dye.

Topics: Alcian Blue; Animals; Female; Glycosaminoglycans; Indoles; Molecular Conformation; Mucopolysaccharidoses; Rats; Rats, Inbred Strains; Staining and Labeling; Suramin

A method is presented for the determination of free glycosaminoglycan (GAG) concentration in as little as 30 microliter serum. By filtration of the serum through DEAE-cellulose paper, the free GAG fraction is selectively adsorbed and concentrated on a circular area of 15.9 mm2; these GAG spots are stained with Alcian Blue. The relationship between the amounts of adsorbed GAG and the optical density of the Alcian Blue spots is linear with a certain range; e.g. for chondroitin sulfate (mixed isomers), from 0.25 to 1.00 micrograms. With this method--which we will refer to as the "DEAE Alcian Blue" method--we estimated the free GAG concentration in sera of individuals of various ages including newborns.

Topics: Adolescent; Adult; Aging; Alcian Blue; Child; Child, Preschool; Female; Glycosaminoglycans; Humans; Infant; Infant, Newborn; Infant, Premature; Male; Microchemistry; Middle Aged; Mucopolysaccharidoses; Mucopolysaccharidosis I; Mucopolysaccharidosis III; Spectrophotometry