STUDIES ON THE QUANTIFICATION OF PROTEOGLYCANS BY THE DIMETHYLMETHYLENE BLUE DYE-BINDING METHOD - SPECIFICITY, QUANTITATION IN SYNOVIAL LAVAGE FLUID, AND AUTOMATION

The dimethylmethylene blue (DMMB) dye-binding technique is widely used for the quantification of sulfated glycosaminoglycans (sGAG) and proteoglycans. We conducted further studies on this technique in our laboratory and found that concentrations of DNA and RNA in excess of 20 mug/ml interfered negatively with the detection of sGAGs; interference was eliminated by using DNase and RNase. Hyaluronan at 40 mug per ml did not interfere with the detection of sGAG. However, because of the higher concentrations of hyaluronan in synovial lavage fluid, it was necessary to treat this fluid with Streptomyces hyaluronidase in order to quantify sGAG. The DMMB assay was automated with a laboratory work station and compared to the standard method.