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Immunochemical Protocols pp 297–311Cite as

Immunogold Probes in Electron Microscopy

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Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 80))

Abstract

Electron microscopy permits the detailed study of cell relationships within tissues and organelles within cells. Two ultrastructural techniques in which gold probes have proven invaluable are immunocytochemistry and in situ hybridization, and these will be described here. Until 1980, peroxidase was the marker of choice, but now colloidal gold is almost universally used. The advantages of colloidal gold are that:

  1. 1.

    It is particulate and very dense, therefore easily identified on biological sections;

  2. 2.

    It is small and will not obscure fine structural details,

  3. 3.

    It can be prepared in several different sizes for use in multiple labeling experiments and quantitation, and

  4. 4.

    It is easily conjugated to immunoglobulins or proteins A or G.

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References

  1. Beesley, J. (1989) Colloidal gold: a new perspective for cytochemical marking. Royal Microscopical Society Handbook 17. Oxford Science Publications, Oxford, UK.

    Google Scholar 

  2. Gross, U., Bormuth, H., Gaissmaier, C, Dittrich, C, Krenn, V., Bohne, W, and Ferguson, D. J. P. (1995) Monoclonal rat antibodies directed against Toxoplasma gondii suitable for studying tackyzoite-bradyzoite interconversion in vivo Clin Diag Lab Immunol 2, 542–548

    CAS  Google Scholar 

  3. Levy, E R. and Hemngton, C. S (eds). (1995) Nonisotopic Methods in Molecular Biology—A Practical Approach Oxford University Press, Oxford

    Google Scholar 

  4. Morel, G. (ed). (1993) Hybridization Techniques for Electron Microscopy CRC, Boca Raton, FL

    Google Scholar 

  5. Morey, A L, Ferguson, D J P, Leslie, K O., Taatses, D. J, and Fleming, K. A (1993) Detection of parvoviral and human DNA sequences at the ultrastructural level by in situ hybridization with digoxigenin-labeled probes. Histochem J 25, 421–429

    Article  PubMed  CAS  Google Scholar 

  6. Morey, A L. (1995) Nonisotopic in situ hybridization at the ultrastructural level J Pathol 176, 113–121

    Article  PubMed  CAS  Google Scholar 

  7. Morey, A L., Ferguson, D. J P., and Fleming, K. A. (1995) Combined immunocytochemistry and nonisotopic in situ hybridization for ultrastructural investigation of human parvovirus B19 infection Histochem J 27, 46–53.

    Article  PubMed  CAS  Google Scholar 

  8. Slot, J W. and Geuze, H J (1984) Gold markers for single and double immunolabeling of ultrathin cryosections, in Immunolabehng for Electron Microscopy (Polak, J M. and Varndell, I. M, eds), Elsevier, Amsterdam, pp 129–142

    Google Scholar 

  9. Tokuyasu, K T. (1978) A study of positive staining of ultrathin frozen sections. J Ultrastruct Res 63, 287–307

    Article  PubMed  CAS  Google Scholar 

  10. Reynolds, E. S. (1963) The use of lead citrate at high pH as an electron-opaque stain in electron microscopy J Cell Biol 17, 208–212

    Article  PubMed  CAS  Google Scholar 

  11. Carlemalm, E, Garavito, R M., and Villiger, W (1982) Resin development for electron microscopy and an analysis of embedding at low temperature J Microsc 126, 123–143.

    CAS  Google Scholar 

  12. Griffiths, G, McDowall, A., Back, R, and Dubochet, J (1984) On the preparation of cryosections for immunocytochemistry J Ultrastruct Res 89, 68–78

    Article  Google Scholar 

  13. Tokuyasu, K. T. (1986) Immunocryoultramicrotomy J Microsc 143, 139–149

    PubMed  CAS  Google Scholar 

  14. Mannweiler, K., Hohenberg, H., Bohn, W., and Rutter, G. (1982) Protein A-gold particles as markers in replica immunocytochemistry. high resolution electron microscope investigations of plasma membrane surfaces J Microsc 126, 145–149

    PubMed  CAS  Google Scholar 

  15. Hodges, G M., Southgate, J, and Toulson, E. C (1987) Colloidal gold-a powerful tool in S E. M. immunocytochemistry an overview of bioapplications. Scanning Electron Microscopy 1, 301–318.

    CAS  Google Scholar 

  16. Murphy, J A. (1980) Noncoating techniques to render biological specimens conductive/1980 update Scanning Electron Microscopy 1, 209–220

    Google Scholar 

  17. mnPinto da Silva, P., Barbosa, M. L. F, and Aguas, A P. (1986) A guide to fracture label, cytochemical labeling of freeze-fractured cells, in Advanced Techniques in Biological Electron Microscopy (Koehler, J K, ed), Springer-Verlag, Berlin, pp 201–227

    Google Scholar 

  18. mnPinto da Silva, P. and Kan, F. W. (1984) Label-fracture a method for high resolution labeling of cell surfaces J Cell Biol 99, 1156–1161

    Google Scholar 

  19. Robards, A. W. and Sleytr, U. B (1985) Low temperature methods in biological electron microscopy, in Practical Methods in Electron Microscopy, vol 10 (Glauert, A M, ed), Elsevier, Amsterdam

    Google Scholar 

  20. Robertson, D, Monaghan, P., Clark, C, and Atherton, A. J (1992) An appraisal of low temperature embedding by progressive lowering of temperature into Lowicryl HM20 for immunocytochemical studies J Microsc. 168, 85–100

    PubMed  CAS  Google Scholar 

  21. Dudek, R W, Varndell, I. M., and Polak, J. M (1984) Combined quick-freeze and freeze-drying techniques for improved electron immunocytochemistry, in Immunolabeling for Electron Microscopy (Polak, J M and Varndell, I M, eds.), Elsevier, Amsterdam, pp 235–248

    Google Scholar 

  22. Oprins, A D., Geuze, H J, and Slot, J W (1994) Cryosubstitution dehydration of aldehyde-fixed tissue, a favourable approach to quantitative immunocytochemistry J Histochem Cytochem 42, 497–503

    PubMed  CAS  Google Scholar 

  23. Monaghan, P and Robertson, D (1990) Freeze-substitution without aldehyde or osmium fixatives ultrastructural and implication for immunocytochemistry. J Microsc 158, 355–363

    PubMed  CAS  Google Scholar 

  24. Shimizu, H. and Nishikawa, T. (1993) Application of an image analyser to gold labeling in immunoelectron microscopy to achieve better demonstration and quantitative analysis J Histochem Cytochem 41, 123–128

    PubMed  CAS  Google Scholar 

  25. Morris, R. E, Ciraolo, G M, and Saelinger, C. B (1991) Gold enhancement of gold-labeled probes gold-intensified staining technique (GIST). J Histochem Cytochem 39, 1585–1591.

    PubMed  CAS  Google Scholar 

  26. Lackie, P M, Hennessy, R J, Hacker, G W, and Polak, J M (1985) Investigation of immunogold-silver staining using electron microscopy Histochemistry 83, 545–550

    Article  PubMed  CAS  Google Scholar 

  27. Manara, G C, Ferrari, C, Torresam, C, Sansoni, P, and De Panfilis, G. (1990) The immunogold-silver staining approach in the study of lymphocyte subpopulations in transmission electron microscopy. J Immunol Methods 128, 59–63

    Article  PubMed  CAS  Google Scholar 

  28. Otsuki, Y, Maxwell, L. E, Magan, S., and Kubo, H (1990) Immunogold-silver staining method for light and electron microscopic detection of lymphocyte cell surface antigens with monoclonal antibodies J Histochem Cytochem 38, 1215–1221.

    PubMed  CAS  Google Scholar 

  29. Burry, R. W., Vandrè, D D., and Hayes, D M. (1992) Silver enhancement of gold antibody probes in preembedding electron microscopic immunocytochemistry J Histochem Cytochem 40, 1849–1856

    PubMed  CAS  Google Scholar 

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Author information

Authors and Affiliations

  1. Department of Molecular Pathology, Glaxo-Wellcome Medicines Research Centre, Stevenage, UK

    David J. P. Ferguson & Julian E. Beesley

  2. Nuffield Department of Surgery, John Radcliffe Hospital, Oxford, UK

    David A. Hughes

Authors
  1. David J. P. Ferguson
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  2. David A. Hughes
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  3. Julian E. Beesley
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Editor information

Editors and Affiliations

  1. The University of Birmingham, UK

    John D. Pound

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© 1998 Humana Press Inc., Totowa, NJ

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Ferguson, D.J.P., Hughes, D.A., Beesley, J.E. (1998). Immunogold Probes in Electron Microscopy. In: Pound, J.D. (eds) Immunochemical Protocols. Methods in Molecular Biology™, vol 80. Humana Press. https://doi.org/10.1007/978-1-59259-257-9_31

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  • DOI: https://doi.org/10.1007/978-1-59259-257-9_31

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-493-8

  • Online ISBN: 978-1-59259-257-9

  • eBook Packages: Springer Protocols

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