Glycosaminoglycan Assay Blyscan™
The Blyscan™ Glycosaminoglycan Assay is a quantitative dye-binding method for the analysis of sulfated proteoglycans and glycosaminoglycans, (sGAG).
Test material can be assayed directly when present in a soluble form, or following papain extraction from biological materials. The assay can be used to measure the total sGAG content and can also be adopted to determine the O- and N-sulfated glycosaminoglycan ratio within test samples.
The dye label used in the assay is 1,9-dimethylmethylene blue and the dye is employed under conditions that provide a specific label for the sulfated polysaccharide component of proteoglycans or the protein free sulfated glycosaminoglycan chains.
The assay is not suitable for small sulfated disaccharide fragments or for the samples containing alginates, as these contain uronic acid.
Blyscan Assay; step-by-step
(a) 0 and 5µg of sGAG and Blyscan Dye, (after 15 minutes mixing).
(b) 30 min mixing and then centrifuged, (note the sGAG-Dye pellet).
(c) The non-sGAG Dye was drained from tubes with pellet retained.
(d) Dye released from sGAG using the Dye Dissociation Reagent.
Soluble extracts from:
- fibrous and hyaline cartilages.
- arteries, heart, lung, skin and other material containing extracellular matrix, (connective tissue) and solid tumours.
- extracellular matrix components that may be released by live cells into the culture medium, some of which can be attached to cell culture plasticware.
- soluble sGAG from synovial fluid, urine and gel chromatography fraction aliquots.
Q. What is the sensitivity of the Blyscan assay?
Q. How long does the Blyscan assay take to run?
A. 1 hour
Q. Can I differentiate between N-sulfated and O-sulfated glycosaminoglycans?
A. A method is given in the manual to differentiate between the glycosaminoglycans present and all necessary reagents are supplied in the kit.
Q. What absorbance values can samples be measured at?
A. The absorbance peak for Blyscan dye in the dissociation reagent is 656nm. This absorbance is suitable for use with most colorimeters and microplate readers with a red filter.
Q. How do I prepare samples for measurement with the Blyscan assay?
A. Preparation details for samples in tissue culture medium, cartilage/soft tissue and urine samples are described in the assay manual.