The Histochemical Journal

, Volume 7, Issue 6, pp 529–546 | Cite as

The Masson staining of collagen — an explanation of an apparent paradox

  • Michael H. Flint
  • Mary F. Lyons
  • M. F. Meaney
  • D. E. Williams


Although collagen of either tendon or dermis can be stained equally well with Ponceau 2R/Acid Fuchsin or Light Green SF if the dyes are used independently, tendon collagen retains the red dye mixture and dermal collagen the green counterstain when the dyes are used sequentially in the Masson trichrome procedure. The results of experiments designed to assess differences in the penetration, retention and displacement of these arylmethane dyes have demonstrated that they are retained more firmly by the tensioned collagen of tendon or stretched dermis, and are more easily displaced from the collagen of relaxed tendon or dermis.

Experiments designed to test the basis of these differences in dye retention indicate that more positively-charged amino dye-binding sites are available in the tensioned collagen than in relaxed collagen, where they appear to be closely associated with adjacent carboxyl groups on the collagen fibres. The possibility that the carboxyl groups of associated acid mucopolysaccharides are implicated in the differences in staining propensity has been investigated and discounted. It is suggested that whereas the binding of arylmethane dyes to collagen under tension is through strong ionic linkages to amino groups, the binding of these and other dyes to relaxed collagen is through weaker hydrogen bonds. It is proposed that these differences in charge distribution on the collagen of the two situations is related to the previously described piezo-electric effect demonstrable on stretched collagen.


Collagen Fibre Fuchsin Mucopolysaccharide Ponceau Weak Hydrogen Bond 
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  1. Baker, J. R. (1958).Principles of Biological Microtechnique. London: Methuen.Google Scholar
  2. Bassett, C. A. L. (1971). Effect of force on skeletal tissues. In:Physiological Basis of Rehabilitation Medicine (eds J. Downey and R. E. Darling), Ch. 16, pp. 283–316. Philadelphia: Saunders.Google Scholar
  3. Bassett, C. A. L. &Becker, R. O. (1962). Generation of electric potentials by bone in response to mechanical stress.Science 137, 1063–4.Google Scholar
  4. Constantine, V. S. &Mowry, R. W. (1968). Selective staining of human dermal collagen.J. invest. Derm. 50, 414–18.Google Scholar
  5. Craik, J. E. &McNeil, I. R. R. (1965). Histological studies on stressed skin. In:Biomechanics and Related Bio-engineering Topics (ed. R. M. Kenedi), pp. 159–64. Oxford: Pergamon Press.Google Scholar
  6. Craik, J. E. &McNeil, I. R. R. (1966). Micro-architecture of skin and its behaviour under stress.Nature, Lond. 209, 931–2.Google Scholar
  7. Crossmon, G. (1937). A modification of Mallory's connective tissue stain with a discussion of the principles involved.Anat. Rec. 69, 33–8.Google Scholar
  8. Dreyer, C. J. (1961). Properties of stressed bone.Nature, Lond. 189, 594–5.Google Scholar
  9. Flint, M. H. (1972). Interrelationships of mucopolysaccharide and collagen in connective tissue remodelling.J. Embryol. exp. Morph. 27, 481–95.Google Scholar
  10. Flint, M. H. (1973a). The biological basis of Langer's lines. In:The Ultrastructure of Collagen (ed. J. Longacre), Ch. 8. Springfield, Illinois: Charles Thomas.Google Scholar
  11. Flint, M. H. (1973b). The basis of the histological demonstration of tension in collagen. In:The Ultrastructure of Collagen (ed. J. Longacre), Ch. 4. Springfield, Illinois: Charles Thomas.Google Scholar
  12. Fukada, E. &Yasuda, I. (1957). On the piezoelectric effect of bone.J. phys. Soc. Japan 12, 1158–62.Google Scholar
  13. Fukada, E. &Yasuda, I. (1964). Piezoelectric effects in collagen.Jap. J. appl. Phys. 3, 117–21.Google Scholar
  14. Gibson, T. (1965). Biomechanics in plastic surgery. In:Biomechanics and Related Bio-engineering Topics (ed. R. M. Kenedi), pp. 129–34. Oxford: Pergamon Press.Google Scholar
  15. Gurr, E. (1971).Synthetic Dyes in Biology, Medicine and Chemistry, p. 395. London and New York: Academic Press.Google Scholar
  16. Harkness, R. D. (1968). Mechanical properties of collagenous tissues. In:Treatise on Collagen (ed. G. N. Ramachandran), Vol. 2, pp. 247–310. London: Academic Press.Google Scholar
  17. Lendrum, A. C. &McFarlane, D. (1940). A controllable modification of Mallory's trichrome staining method.J. Path. 50, 381–4.Google Scholar
  18. Liisberg, M. F. (1959). A new differential staining method for connective and muscular tissue.Acta anat. 36, 93–100.Google Scholar
  19. Lillie, R. D. (1940). Further experiments with the Masson trichrome modification of Mallory's connective tissue stain.Stain Technol. 15, 17–22.Google Scholar
  20. Lillie, R. D. (1945). Studies on selective staining of collagen with acid anilin dyes.J. tech. Meth. 25, 1–47.Google Scholar
  21. Lillie, R. D. (1964). Histochemical acylation of hydroxl and amine groups.J. Histochem. Cytochem. 12, 821–41.Google Scholar
  22. Lillie, R. D. (1965).Histopathologic Technic and Practical Histochemistry, 3rd Ed., pp. 539–50. New York: McGraw-Hill.Google Scholar
  23. Lillie, R. D. (1969).H. J. Conn's Biological Stains, pp. 44–5. Baltimore: Williams & Wilkins.Google Scholar
  24. Macconaill, M. A. (1961). Staining properties of stressed bone.Nature, Lond. 192, 368–9.Google Scholar
  25. Masson, P. (1929). Some histological methods. Trichrome stainings and their preliminary technique.J. tech. Meth. 12, 75–90.Google Scholar
  26. Millington, P. F., Gibson, T., Evans, J. H. &Barbenel, J. C. (1971). Structural and mechanical aspects of connective tissue. In:Advances in Biomedical Engineering, Vol. 1 (ed. R. M. Kenedi), pp. 189–248. London: New York: Academic Press.Google Scholar
  27. Mowry, R. W. (1963). The special value of methods that color both acidic and vicinal hydroxyl groups in the histochemical study of mucins. With revised directions for the colloidal iron stain, the use of alcian blue G8X and their combinations with the periodic acid-schiff reaction.Ann. N.Y. Acad. Sci. 106, 402–23.Google Scholar
  28. Parks, L. R. &Bartlett, P. G. (1935). Dyeing with acid dyes.Am. Dyestuff Reptr. 24, 476–8, 495.Google Scholar
  29. Pearse, A. G. E. (1968).Histochemistry: Theoretical and Applied, 3rd Ed., Vol. 1, pp. 73, 604. London: Churchill.Google Scholar
  30. Puchtler, H. &Isler, H. (1958). The effect of phosphomolybdic acid on the stainability of connective tissues by various dyes.J. Histochem. Cytochem. 6, 265–70.Google Scholar
  31. Ridge, M. D. &Wright, V. (1965). A rheological study of skin. In:Biomechanics and Related Bio-engineering Topics (ed. R. M. Kenedi), pp. 165–75. Oxford: Pergamon Press.Google Scholar
  32. Stearn, A. E. &Stearn, Esther W. (1929). The mechanism of staining explained on a chemical basis. I. The reaction between dyes, proteins and nucleic acid.Stain Technol. 4, 111–19.Google Scholar
  33. Stearn, A. E. &Stearn, Esther W. (1930). The mechanism of staining explained on a chemical basis. II. General presentation.Stain Technol. 5, 17–24.Google Scholar
  34. Sweat, Faye, Meloan, Susan N. &Fuchtler, H. (1968). A modified one-step trichrome stain for demonstration of fine connective tissue fibers.Stain Technol. 43, 227–31.Google Scholar
  35. Wilkes, G. L., Brown, I. A. &Wildnauer, R. H. (1973). The biomechanical properties of skin.CRC Crit. Rev. Bioeng. 1, 453–95.Google Scholar
  36. Wright, V. (1971). Elasticity and deformation of skin. In:Biophysical Properties of the Skin (ed. H. R. Elden), pp. 437–49. New York, London: InterscienceGoogle Scholar

Copyright information

© Chapman and Hall Ltd 1975

Authors and Affiliations

  • Michael H. Flint
    • 1
  • Mary F. Lyons
    • 1
  • M. F. Meaney
    • 1
  • D. E. Williams
    • 1
  1. 1.Department of Surgery, School of MedicineUniversity of AucklandAucklandNew Zealand

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