Journal of Endocrinological Investigation

, Volume 17, Issue 8, pp 625–629 | Cite as

Comparison of immuno- and HPLC-assays for the measurement of urinary collagen cross-links

  • E. Calabresi
  • L. Lasagni
  • F. Franceschelli
  • V. de Leonardis
  • A. Becorpi
  • M. Serio
  • Maria Luisa Brandi


Pyridinoline (Pyr) and deoxypyridinoline (D-Pyr) are two cross-links of collagen molecules, that are present in the extracellular matrix and released during its degradation. Pyr is present in bone and cartilage, but not in significant amounts in other connective tissues and D-Pyr appears to be specific for bone tissue. Therefore, the urinary excretion of Pyr and D-Pyr might be a sensitive marker of bone matrix degradation. For the determination of urinary Pyr and D-Pyr two methods are available: a chromatographic method (HPLC) by which it is possible to measure separately Pyr and D-Pyr, and a new immunoassay which measures total free and low molecular weight pyridinoline released in the urine. We compared the results obtained by HPLC analysis of 205 urinary samples from normal subjects and patients affected by various bone disorders with those obtained by the immunoassay. The overall correlation coefficient between the results obtained by the two methods was 0.34. When calculated in a range of pyridinoline concentrations from 0 to 30, 30 to 60, and over 60 pmol/μmol creatinine the correlation coefficient was respectively − 0.094, 0.38, and 0.12. The two methods yielded variable profiles in the detection of circadian rhythms and these differences did not segregate with normal or pathological conditions. We conclude that the immunoassay proposed for the determination of urinary collagen cross-links is not immediately applicable to clinical use. The improvement of the antibody specificity will probably contribute to replace the HPLC method with the immunoassay.


Cross-links HPLC immunoassay 


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  1. 1.
    Fujimoto D., Moriguchi T., Ishida T., Hayashi H. The structure of pyridinoline. A collagen crosslink. Biochem. Biophys. Res. Commun. 84: 52, 1978.PubMedCrossRefGoogle Scholar
  2. 2.
    Robins S.P., Duncan A. Cross-linking of collagen. Location of pyridinoline in bone articular cartilage at two sites of the molecule. Biochem. J. 215: 175, 1983.PubMedCentralPubMedGoogle Scholar
  3. 3.
    Robins S.P., Duncan A. Pyridinium crosslinks of bone collagen and their location in peptides isolated from rat femur. Biochem. Biophys. Acta 114: 233, 1987.Google Scholar
  4. 4.
    Eyre D.R., Paz M.A., Gallop P.M. Crosslinking in collagen and elastin. Annu. Rev. Biochem. 53: 717, 1984.PubMedCrossRefGoogle Scholar
  5. 5.
    Eastell R., Colwell A., Assiri A.M.A. Prediction of response to estrogen therapy using urine deoxypyridinoline. J. Bone Miner. Res. 5:, 1990.Google Scholar
  6. 6.
    Delmas P.D., Schlemmer A., Gineyts E. Urinary excretion of pyridinoline correlated with bone turnover measured on iliac crest biopsy in patients with vertebral osteoporosis. J. Bone Miner. Res. 6: 639, 1991.PubMedCrossRefGoogle Scholar
  7. 7.
    Colwell A., Eastell R., Assiri A.M.A., Russel R.G.G. Effect of diet on deoxypyridinoline excretion. In: Christiansen C., Overgaard K. (Eds.), Osteoporosis 1990. Proceedings of the third international symposium on osteoporosis. Osteopress, Aalborg, 1990, p. 590.Google Scholar
  8. 8.
    Eyre D.E., Koob T.J., Van Ness K.P. Quantitation of hydroxypyridinium cross-links in collagen by high performance liquid chromatography. Anal. Biochem. 137: 380, 1984.PubMedCrossRefGoogle Scholar
  9. 9.
    Calabresi E., Lasagni L., Bartolini L., Franceschelli F., de Leonardis V., Brandi M.L. Comparison of immuno- and HPLC-assays for the measurement of urinary collagen cross-links. Fourth International Symposium on Osteoporosis. Hong Kong 1993, (abstract 125).Google Scholar
  10. 10.
    Black D., Marabani M., Sturrock R.D., Robins S.P. Urinary excretion of hydroxypyridinium crosslinks of collagen in patients with rheumatoid arthritis. Ann. Rheum. Dis. 48: 641, 1989.PubMedCentralPubMedCrossRefGoogle Scholar
  11. 11.
    Body J.J., Delmas P.D. Urinary pyridinium crosslinks as markers of bone resorption in tumor-associated hypercalcemia. J. Clin. Endocrinol. Metab. 74: 471, 1992.PubMedGoogle Scholar
  12. 12.
    Robins S.P., Black D., Paterson R.C., Reid D.M., Duncan A., Seibel M.J. Evaluation of urinary hydroxypyridinium crosslink measurement as resorption markers in metabolic bone disease. Eur. J.Clin. Invest. 21: 310, 1991.PubMedCrossRefGoogle Scholar
  13. 13.
    Seibel M.J., Gartenberg F., Silverberg S.J. Urinary hydroxypyridinium crosslinks of collagen in primary hyperparathyroidism. J. Clin. Endocrinol. Metab. 74: 481, 1992.PubMedGoogle Scholar
  14. 14.
    Uebelhardt D., Gineyts E.C., Chapuy M.C., Delmas P.D. Urinary excretion of pyridinium crosslinks: a new marker of bone resorption in metabolic bone disease. Bone Miner. 8: 87, 1990.CrossRefGoogle Scholar
  15. 15.
    Robins S.P. An enzyme-linked immunoassay for the collagen crosslink, pyridinoline. Biochem. J. 207: 617, 1982.PubMedCentralPubMedGoogle Scholar
  16. 16.
    Seyedin S., Zuk R., Kung V., Danilov Y., Shepard K. An immunoassay to urinary collagen crosslinks. Bone Miner. (suppl.) 17, 1992, (abstract 534).Google Scholar
  17. 17.
    Delmas P.D. Biochemical markers of bone turnover in osteoporosis. Fourth International Symposium on Osteoporosis. Hong Kong, 1993, (abstract 48).Google Scholar
  18. 18.
    Gineyts E., Bertholin A., Garnero P., Marchand F., Delmas P.D. Immunoassay (ELISA) of urinary pyridinoline crosslink excretion. Fourth International Symposium on Osteoporosis. Hong Kong 1993, (abstract 149).Google Scholar

Copyright information

© Italian Society of Endocrinology (SIE) 1994

Authors and Affiliations

  • E. Calabresi
    • 1
  • L. Lasagni
    • 1
  • F. Franceschelli
    • 3
  • V. de Leonardis
    • 4
  • A. Becorpi
    • 2
  • M. Serio
    • 1
  • Maria Luisa Brandi
    • 1
  1. 1.Dipartimento di Fisiopatologia ClinicaUniversità di FirenzeFirenzeItaly
  2. 2.Dipartimento di GinecologiaUniversità di FirenzeFirenzeItaly
  3. 3.III Istituto di Medicina InternaUniversità di FirenzeFirenzeItaly
  4. 4.IV Istituto di Medicina InternaUniversità di FirenzeFirenzeItaly

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