Advertisement

The Histochemical Journal

, Volume 19, Issue 12, pp 671–675 | Cite as

The impact of different fixation procedures on staining of macromolecules in a microtiter system

  • M. R. Barer
  • H. O. Lyon
Papers

Summary

The effects of formaldehyde, glutaraldehyde, methanol, Clarke's fixative and microwave irradiation on the quantitative staining of proteins (Naphthol Yellow S) and nucleic acids (Ethyl Green—Pyronin) in a cell culture system have been investigated. Overall, glutaraldehyde rapidly yielded the highest and most consistent levels of staining when compared to all other chemical fixatives. Although microwave irradiation was found to be uneven, 4 min exposure to 700W was found to give higher levels of protein staining than those achieveable with glutaraldehyde. Time-dependent processes were observed with all procedures. In addition, dissociations in the trends of protein and nucleic acid staining were observed. It is suggested that these results domonstrate fixation events that have not previously been resolved from the effects of reagent penetration into tissue blocks.

Keywords

Nucleic Acid Fixation Event Macromolecule Glutaraldehyde Culture System 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. BAKER, J. R. (1966)Cytological Technique — The Principles Underlying Routine Methods. 5th edn. pp. 39–54. London: Methuen.Google Scholar
  2. BANCROFT, J. D. & STEVENS, A. (1982)Theory and Practice of Histological Techniques. 2nd edn. Churchill-Livingstone: Edinburgh.Google Scholar
  3. BARER, M. R., LYON, H. O. & DRASAR, B. S. (1986a) Quantitative of Dye binding to cell monolayers in a microtiter system.Histochem. J. 18, 122–8.Google Scholar
  4. BARER, M. R., MANN, G. F. & DRASAR, B. S. (1986b) A semi-automated system for the assessment of toxicity to cultured mammalian cells based on quantitation of staining properties.Dev. Biol. Stand. 64, 251–9.Google Scholar
  5. BRACHET, J. (1940) La detection histochimique des acides pentose-nucleiques.Compete. Rend. Soc. Biol. 133, 88–90.Google Scholar
  6. BRACHET, J. (1953) The use of basic dyes and ribonuclease for the cytochemical detection of ribonucleic acid.Quart. J. Micr. Sci. 94, 1–10.Google Scholar
  7. HAYAT, M. A. (1970)Principles and Techniques of Electron Microscopy: Biological Applications. Vol. 1, pp. 5–107. New York: Van Nostrand Reinhold Company.Google Scholar
  8. HOPWOOD, D. (1975) The reactions of glutaraldehyde with nucleic acids.Histochem. J. 7, 267–75.Google Scholar
  9. HOPWOOD, D. (1985) Cell and tissue fixation, 1972–1982.Histochem J. 17, 389–442.Google Scholar
  10. HOPWOOD, D., COGHILL, G. RAMSAY, J., MILNE, G. & KERR, M. (1984) Microwave fixation: its potential for routine techniques, histochemistry, immunocytochemistry and electron microscopy.Histochem. J. 16, 1171–91.Google Scholar
  11. HOROBIN, R. W. (1982)Histochemistry. pp. 19–55. Gustav Fischer: Stuttgart. Butterworths: London.Google Scholar
  12. JAMES, J. & TAS, J. (1984)Histochemical Protein Staining Methods. p. 28 Royal Microscopical Society: Oxford University Press.Google Scholar
  13. LYON, H., JAKOBSEN, P., CLAUSEN, P. P. & ANDERSON, A. P. (1983) Methyl Green-Pyronin staining with pure pyronin Y.Histochem J. 15, 605–6.Google Scholar
  14. PATTERSON, M. K., JR. & BULARD, R. (1980) Microwave fixation of cells in tissue culture.Stain Technol. 55, 71–5.Google Scholar
  15. PEARSE, A. G. E. (1980)Histochemistry: Theoretical and Applied. 4th edn. Vol. 1 pp. 97–158. Edinburgh: Churchill Livingstone.Google Scholar
  16. PEARSE, A. G. E. (1985)Histochemistry: Theoretical and Applied. 4th edn. Vol. 2 pp. 611–74. Edinburgh: Churchill Livingstone.Google Scholar
  17. VANHA-PERTULA, T. & GRIMLEY, P. M. (1976) Loss of proteins and other macromolecules during preparation of cell culture for high resolution autoradiography. Quantitation by a micromethod.J. Histochem. Cytochem. 18, 565–73.Google Scholar

Copyright information

© Chapman and Hall Ltd. 1987

Authors and Affiliations

  • M. R. Barer
    • 1
  • H. O. Lyon
    • 2
  1. 1.Department of Clinical Tropical MedicineThe London School of Hygiene and Tropical MedicineLondonUK
  2. 2.Department of Pathology, Hvidovre HospitalUniversity of CopenhagenHvidovreDenmark

Personalised recommendations