A monoclonal antibody that recognizes 2,3-, 2,6-, and 4,6-disulphate ester ring substitution in pyranose-containing polysaccharides. Its production, characterization and application for the quantitation of pentosan polysulphate, dextran sulphate, glycosaminoglycan polysulphate and chondroitin sulphate E.
A method is described for the preparation of a monoclonal antibody (MAb) which binds specifically to polysaccharides which contain 2,3-, 2,6- and 4,6-disulphate ester pyranose ring substitution. Such molecules include the semisynthetic heparinoids, pentosan polysulphate (PPS), dextran sulphate (DS) and glycosaminoglycan polysulphates (GAGPS), as well as the naturally occurring polysulphated polysaccharides, chondroitin sulphate E, and the 2,6-disulphated galactoses of carrageenans. The antibody (MAb 5-B-10) did not significantly cross-react with other sulphated polysaccharides such as heparin, heparan sulphate, the chondroitin sulphates, A, B, C or D, or keratan sulphate. No cross-reactivity was found with non-sulphated polysaccharides or polyanions including hyaluronic acid, xylan, or DNA. MAb 5-B-10 was characterized as IgM and kappa light chains, and was to develop an amplified enzyme-linked immunosorbent inhibition assay (ELISIA) to detect and quantitate some of the polysulphated polysaccharides in biological fluids. Using this assay, the lower limits of detection of these compounds in serum were in the order of 50 ng/ml; however 50% inhibition was obtained between 200-500 ng/ml. The intra- and inter-assay coefficients of variation for PPS were 4.2 +/- 2.8 and 16.7 +/- 13.8% respectively. The MAb 5-B-10 and the ELISIA were used to determine the levels of PPS in plasma of three healthy non-fasted volunteers for up to 120 min post-intravenous infusion (1.0 mg/kg). The results obtained compared favourably with kinetic data reported by others using a competitive binding assay method for this drug.
Raymond Purves Research Laboratories (University of Sydney), Royal North Shore Hospital, St. Leonards, N.S.W., Australia.
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Kongtawelert
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Ghosh
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000302:isolation & purification; D002807:Chondroitin / Q000031:analogs & derivatives*; D002809:Chondroitin Sulfates / Q000032:analysis* / Q000276:immunology; D016264:Dextran Sulfate; D003911:Dextrans / Q000032:analysis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006025:Glycosaminoglycans / Q000032:analysis* / Q000276:immunology; D006825:Hybridomas / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D010426:Pentosan Sulfuric Polyester / Q000032:analysis* / Q000276:immunology; D011134:Polysaccharides / Q000032:analysis*
D000911:Antibodies, Monoclonal; D003911:Dextrans; D006025:Glycosaminoglycans; D011134:Polysaccharides; C013786:A73025; D010426:Pentosan Sulfuric Polyester; D002807:Chondroitin; D002809:Chondroitin Sulfates; D016264:Dextran Sulfate