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. 1983 Aug 1;213(2):371–378. doi: 10.1042/bj2130371

Cartilage proteoglycan binding region and link protein. Radioimmunoassays and the detection of masked determinants in aggregates.

A Ratcliffe, T Hardingham
PMCID: PMC1152137  PMID: 6193777

Abstract

Antibodies have been raised in rabbits to the hyaluronate-binding region and link-protein components of aggregated proteoglycans from pig laryngeal cartilage. The anti-(binding region) antibodies did not bind 125I-labelled link protein, nor was 125I-labelled binding region bound by the anti-(link protein) antibodies. The antisera were applied in sensitive inhibition radioimmunoassays to determine binding region and link protein in purified proteoglycan preparations. With intact proteoglycan aggregates, the antigenic sites of link protein, and to a lesser extent binding region, were masked. Heat treatment in the presence of sodium dodecyl sulphate (0.025%, w/v) was found to overcome this masking, thereby allowing the determination of link protein and binding region in aggregated proteoglycan preparations in pure and impure samples.

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Selected References

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  1. BITTER T., MUIR H. M. A modified uronic acid carbazole reaction. Anal Biochem. 1962 Oct;4:330–334. doi: 10.1016/0003-2697(62)90095-7. [DOI] [PubMed] [Google Scholar]
  2. Buckwalter J. A., Rosenberg L. C. Electron microscopic studies of cartilage proteoglycans. Direct evidence for the variable length of the chondroitin sulfate-rich region of proteoglycan subunit core protein. J Biol Chem. 1982 Aug 25;257(16):9830–9839. [PubMed] [Google Scholar]
  3. Caterson B., Baker J. R., Levitt D., Paslay J. W. Radioimmunoassay of the link proteins associated with bovine nasal cartilage proteoglycan. J Biol Chem. 1979 Oct 10;254(19):9369–9372. [PubMed] [Google Scholar]
  4. Dimitriadis G. J. Effect of detergents on antibody-antigen interaction. Anal Biochem. 1979 Oct 1;98(2):445–451. doi: 10.1016/0003-2697(79)90165-9. [DOI] [PubMed] [Google Scholar]
  5. Franzén A., Björnsson S., Heinegård D. Cartilage proteoglycan aggregate formation. Role of link protein. Biochem J. 1981 Sep 1;197(3):669–674. doi: 10.1042/bj1970669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hardingham T. E., Ewins R. J., Muir H. Cartilage proteoglycans. Structure and heterogeneity of the protein core and the effects of specific protein modifications on the binding to hyaluronate. Biochem J. 1976 Jul 1;157(1):127–143. doi: 10.1042/bj1570127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hardingham T. E., Muir H. Hyaluronic acid in cartilage and proteoglycan aggregation. Biochem J. 1974 Jun;139(3):565–581. doi: 10.1042/bj1390565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hardingham T. E., Muir H. The specific interaction of hyaluronic acid with cartillage proteoglycans. Biochim Biophys Acta. 1972 Sep 15;279(2):401–405. doi: 10.1016/0304-4165(72)90160-2. [DOI] [PubMed] [Google Scholar]
  10. Hardingham T. E. The role of link-protein in the structure of cartilage proteoglycan aggregates. Biochem J. 1979 Jan 1;177(1):237–247. doi: 10.1042/bj1770237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hardingham T. Proteoglycans: their structure, interactions and molecular organization in cartilage. Biochem Soc Trans. 1981 Dec;9(6):489–497. doi: 10.1042/bst0090489. [DOI] [PubMed] [Google Scholar]
  12. Heinegård D. K., Hascall V. C. The effects of dansylation and acetylation on the interaction between hyaluronic acid and the hyaluronic acid-binding region of cartilage proteoglycans. J Biol Chem. 1979 Feb 10;254(3):921–926. [PubMed] [Google Scholar]
  13. Heinegård D., Axelsson I. Distribution of keratan sulfate in cartilage proteoglycans. J Biol Chem. 1977 Mar 25;252(6):1971–1979. [PubMed] [Google Scholar]
  14. Heinegård D., Hascall V. C. Aggregation of cartilage proteoglycans. 3. Characteristics of the proteins isolated from trypsin digests of aggregates. J Biol Chem. 1974 Jul 10;249(13):4250–4256. [PubMed] [Google Scholar]
  15. Heinegård D. Polydispersity of cartilage proteoglycans. Structural variations with size and buoyant density of the molecules. J Biol Chem. 1977 Mar 25;252(6):1980–1989. [PubMed] [Google Scholar]
  16. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  17. Koziarz J. J., Köhler H., Steck T. L. A system for preparative polyacrylamide gel electrophoresis in sodium dodecyl sulfate. Anal Biochem. 1978 May;86(1):78–89. doi: 10.1016/0003-2697(78)90320-2. [DOI] [PubMed] [Google Scholar]
  18. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  19. Paulsson M., Heinegård D. Radioimmunoassay of the 148-kilodalton cartilage protein. Distribution of the protein among bovine tissues. Biochem J. 1982 Nov 1;207(2):207–213. doi: 10.1042/bj2070207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Perkins S. J., Miller A., Hardingham T. E., Muir H. Physical properties of the hyaluronate binding region of proteoglycan from pig laryngeal cartilage. Densitometric and small-angle neutron scattering studies of carbohydrates and carbohydrate-protein macromolecules. J Mol Biol. 1981 Jul 25;150(1):69–95. doi: 10.1016/0022-2836(81)90325-9. [DOI] [PubMed] [Google Scholar]
  21. Poole A. R., Reiner A., Tang L. H., Rosenberg L. Proteoglycans from bovine nasal cartilage. Immunochemical studies of link protein. J Biol Chem. 1980 Oct 10;255(19):9295–9305. [PubMed] [Google Scholar]
  22. Tang L. H., Rosenberg L., Reiner A., Poole A. R. Proteoglycans from bovine nasal cartilage. Properties of a soluble form of link protein. J Biol Chem. 1979 Oct 25;254(20):10523–10531. [PubMed] [Google Scholar]
  23. Thonar E. J., Kimura J. H., Hascall V. C., Poole A. R. Enzyme-linked immunosorbent assay analyses of the hyaluronate-binding region and the link protein of proteoglycan aggregate. J Biol Chem. 1982 Dec 10;257(23):14173–14180. [PubMed] [Google Scholar]
  24. Wieslander J., Heinegárd D. Immunochemical analysis of cartilage proteoglycans. Radioimmunoassay of the molecules and the substructures. Biochem J. 1980 Jun 1;187(3):687–694. doi: 10.1042/bj1870687. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wieslander J., Heinegård D. Immunochemical analysis of cartilage proteoglycans. Antigenic determinants of substructures. Biochem J. 1979 Apr 1;179(1):35–45. doi: 10.1042/bj1790035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wieslander J., Heinegård D. Immunochemical analysis of cartilage proteoglycans. Cross-reactivity of molecules isolated from different species. Biochem J. 1981 Oct 1;199(1):81–87. doi: 10.1042/bj1990081. [DOI] [PMC free article] [PubMed] [Google Scholar]

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