Advertisement

Histochemistry

, Volume 95, Issue 5, pp 491–494 | Cite as

Ultrastructural detection of DNA-incorporated bromodeoxyuridine in resin embedded brain tissue

  • A. Migheli
  • C. Mocellini
  • M. T. Giordana
Article

Summary

We describe a protocol for the ultrastructural detection of DNA-incorporated bromodeoxyuridine (BUdR) in resin embedded tissue by means of post-embedding immunogold labeling. The paraventricular zone of rat embryos brains was dissected, fixed either in paraformaldehyde or glutaraldehyde, and embedded in LR White. BUdR gold labeling was only found when thin sections were pretreated with 4 N HCl. Other DNA denaturing agents, such as Na ethoxide, formamide, formic acid, heat or HCl at lower concentrations were ineffective. Very little difference in the degree of labeling was found depending on the fixation. This method can be applied to investigate the fine structure of replicating cells in other in vivo conditions, such as human tumors.

Keywords

Gold Brain Tissue Formic Acid Paraformaldehyde Fine Structure 
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. Apte SS, Puddle B (1990) Bromodeoxyuridine (BrdUrd) immunohistochemistry in undecalcified plastic-embedded tissue. Elimination of the DNA denaturation step. Histochemistry 93:631–635Google Scholar
  2. Gratzner HG (1982) Monoclonal antibody to 5-bromo and 5-iododeoxyuridine: a new reagent for detection of DNA replication. Science 218:474–475Google Scholar
  3. Harms G, Goor H van, Koudstaal J, Ley L de, Hardonk MJ (1986) Immunohistochemical demonstration of DNA-incorporated 5-bromodeoxyuridine in frozen and plastic embedded sections. Histochemistry 85:139–143Google Scholar
  4. Hayashi Y, Koike M, Matsutani M, Hoshino T (1988) Effects of fixation time and enzymatic digestion on immunohistochemical demonstration of bromodeoxyuridine in formalin-fixed, paraffin-embedded tissue. J Histochem Cytochem 36:511–514Google Scholar
  5. Hoshino T, Prados M, Wilson CB, Cho Kg, Lee K-S, Davis RL (1989) Prognostic implications of the bromodeoxyuridine labeling index of human gliomas. J Neurosurg 71:335–341Google Scholar
  6. Magaud J-P, Sargent I, Clarke PJ, Ffrench M, Rimokh R, Mason DY (1989) Double immunocytochemical labeling of cell and tissue samples with monoclonal anti-bromodeoxyuridine. J Histochem Cytochem 37:1517–1527Google Scholar
  7. Mazzotti G, Rizzoli R, Galanzi A, Papa S, Vitale M, Falconi M, Neri LM, Zini N, Maraldi NM (1990) High-resolution detection of newly synthesized DNA by anti-bromodeoxyuridine antibodies identifies specific chromatin domains. J Histochem Cytochem 38:13–22Google Scholar
  8. Meyer JS, Nauert J, Koehm S, Hughes J (1989) Cell kinetics of human tumors by in vitro bromodeoxyuridine labeling. J Histochem Cytochem 37:1449–1454Google Scholar
  9. Moran R, Darzynkiewicz Z, Staiano-Coico L, Melamed MR (1985) Detection of 5-bromodeoxyuridine (BrdUrd) incorporation by monoclonal antibodies: role of the DNA denaturation step. J Histochem Cytochem 33:821–827Google Scholar
  10. Newman GR, Jasani B, Williams ED (1983) A simple post-embedding system for the rapid demonstration of tissue antigens under the electron microscope. Histochem J 15:543–555Google Scholar
  11. Schutte B, Reynders MMJ, Bosman FT, Blijham GH (1987) Effect of tissue fixation on anti-bromodeoxyuridine immunohistochemistry. J Histochem Cytochem 35:1343–1345Google Scholar
  12. Thiry M, Dombrowicz D (1988) Anti-bromodeoxyuridine monoclonal antibody: an alternative tool for the identification of replicated DNA at the electron microscope level. Biol Cell 62:99–102Google Scholar
  13. Vogel W, Mehnert K, Pentz S (1990) Demonstration of chromosome replication by BrdU antibody technique and electron microscopy. Hum Genet 84:237–240Google Scholar

Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • A. Migheli
    • 1
  • C. Mocellini
    • 1
  • M. T. Giordana
    • 1
  1. 1.Clinica Neurologica IIUniversità di TorinoTorinoItaly

Personalised recommendations