Studies on the in vitro incubation procedure for [35S]sulphate labelling of articular cartilage glycosaminoglycans.

Articular cartilage from cow and calf femoral condyles was incubated in Tyrodes solution containing [35S]sulphate for different periods up to 80 min. Glycosaminoglycans from the cartilage tissue and incubation medium were fractionated on Cetylpyridinium chloride and ECTEOLA cellulose microcolumns. The incorporation of [35S]sulphate into all individual fractions of chondroitin sulphate and keratan sulphate was found to be linear from 20 to 80 min incubation time. As a rule the total specific activities of keratan sulphate and chondroitin sulphate were similar for both calves and cows. The proteoglycan material recovered from the medium amounted to about 1% of the tissue dry weight and was found to have a higher chondroitin sulphate: keratan sulphate ratio than the corresponding cartilage tissue for both calf and cow. The solubility profiles for the newly synthesised glycosaminoglycans, obtained from determination of the radioactivity in the individual fractions, were compared with those of glycosaminoglycans already present. These curves indicated that newly synthesised chondroitin sulphate had a higher average molecular size than that present in the tissue whereas the newly synthesised keratan sulphate had a smaller average molecular size. These newly synthesised components were also detected in the proteoglycans recovered from the incubation medium.