The quantitative measurement of Alcian Blue–glycosaminoglycan complexes

Abstract

1. A simple new quantitative micro method was developed to study the interaction of the cationic dye Alcian Blue 8GX and acid glycosaminoglycans under different conditions. After washing with ethanol the precipitated Alcian Blue–glycosaminoglycan complex was dissociated in Manoxol IB solution and the amount of bound dye measured spectrophotometrically. 2. Reaction profiles of complex-formation were determined in the presence of different concentrations of MgCl2 at pH5.8, and could be used to study the critical electrolyte concentrations of glycosaminoglycans. At least 50mm-MgCl2 was required to produce maximum precipitation of, and maximum uptake of, Alcian Blue by standard glycosaminoglycans. Maximum uptake of Alcian Blue by glycosaminoglycans in the urine of a patient with Hurler's syndrome required the presence of 25–50mm-MgCl2. 3. Under standard conditions of maximum interaction, calibration curves for the quantitative determination of a series of standard glycosaminoglycans in 20μl volumes were nearly linear over the range 1–10μg. 4. The technique was used to determine the molecular binding ratios of Alcian Blue to glycosaminoglycans under controlled conditions.