title: Demonstration of immunogenic keratan sulphate in commercial chondroitin 6-sulphate from shark cartilage. Implications for ELISA assays.
abstract: The prototype monoclonal keratan sulphate (KS) antibody 5D4 that is widely used for detection of KS in tissues and biological fluids reacts strongly with commercial low grade shark cartilage chondroitin 6-sulphate. Characterization of the immunogenic material by chondroitinase ABC digestion, ELISA inhibition studies, immunoblotting and HPLC analyses confirmed the presence of substantial amounts of KS, probably as a large proteoglycan (> 120 kDa). Commercial and heterogenic glycosaminoglycan preparations therefore must be used with great caution in immunological analyses. On the other hand the shark cartilage chondroitin 6-sulphate is an easy accessible source of immunogenic KS that can be used as a reference standard and as coating antigen in KS-ELISAs. The concentration of immunogenic KS in synovial fluid measured with an ELISA based solely on reagents of shark cartilage chondroitin 6-sulphate correlated well (r = 0.90) with the concentrations obtained with a traditional KS-ELISA that uses purified aggrecan as standard and coating antigen, and KS in both serum and synovial fluid could be measured with sufficient linearity.
authors.affiliation: Department of Clinical Biochemistry, K.H. University Hospital, Aarhus, Denmark.
authors.forename: H J
authors.initials: HJ
authors.lastname: Møller
authors.forename: T
authors.initials: T
authors.lastname: Møller-Pedersen
authors.forename: T E
authors.initials: TE
authors.lastname: Damsgaard
authors.forename: J H
authors.initials: JH
authors.lastname: Poulsen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002240:Carbohydrate Sequence; D002356:Cartilage / Q000276:immunology*; D002808:Chondroitin Lyases / Q000378:metabolism; D002809:Chondroitin Sulfates / Q000276:immunology*; D003315:Cornea; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006025:Glycosaminoglycans; D006801:Humans; D007632:Keratan Sulfate / Q000276:immunology*; D008969:Molecular Sequence Data; D012754:Sharks; D014907:Whales
chemical_list: D000911:Antibodies, Monoclonal; D006025:Glycosaminoglycans; D002809:Chondroitin Sulfates; D007632:Keratan Sulfate; D002808:Chondroitin Lyases