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Attempts to quantitate immunocytochemistry at the electron microscope level

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Summary

The ability to localize intracellular macromoleculesin situ by high resolution techniques has been made possible by the development of antibody labelling of thin sections obtained either from tissues embedded in an hydrophilic matrix, or by ultracryotomy or from conventional plastic embedded tissue. When particle-tagged immunological reagents are used to visualize intracellular antigens, quantitative information can be obtained by combining particle counts with morphometric estimations of compartment volume. Various detection systems have been used successfully for quantitation, which include ferritin-conjugated antibodies, biotin-avidin-ferritin complexes and, more recently, gold-protein A conjugates. Examples of the use of these techniques the localization of secretory proteins in pancreatic exocrine cells, opsin and a large membrane protein in photoreceptor cells of frog retina, and contractile proteins in skeletal muscle are given. Quantitative data obtained by morphometric analysis, both in bovine and rat pancreatic exocrine cells, are compared with values assessed by biochemical methods.

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References

  • Avrameas, S. &Uriel, J. (1966) Méthode de marquage d'antigènes et d'anticorps avec des enzymes et son application en immunodiffusion.C. r. hebd. Séanc. Acad. Sci., Paris 262, 2543.

    Google Scholar 

  • Batten, T. F. C. & Hopkins, C. R. (1980) Simultaneous localization of LHβ and FSHβ in porcine gonadotrophs by immunoelectromicroscopy using IgG bound through protein A to gold particle markers of specific size.Mol. Cell Endocrinol. (in press).

  • Batten, T. F. C. &Hopkins, C. R. (1978) Use of protein A-coated colloidal gold particles for immunoelectron microscopic localization of ACTH on ultrathin sections.Histochemistry 60, 317.

    Google Scholar 

  • Bendayan, M., Roth, J., Perrelet, A. & Orci, L. (1980) Quantitative immunocytochemical localization of exocrine pancreatic enzymes in subcellular compartments of the rat acinar cell.J. Histochem. Cytochem. (in press).

  • Geuze, J. J., Slot, J. W. &Tokuyasu, K. T. (1979) Immunocytochemical localization of amylase and chymotrypsinogen in the exocrine pancreatic cell with special attention to the Golgi complex.J. Cell Biol. 82, 697.

    Google Scholar 

  • Greene, L. J., Hirs, C. H. &Palade, G. E. (1963) On the protein composition of bovine pancreatic zymogen granules.J. biol. Chem. 238, 2054.

    Google Scholar 

  • Heitzmann, H. &Richards, F. M. (1974) Use of the avidin-biotin complex for specific staining of biological membranes in electron microscopy.Proc. natn. Acad. Sci. U.S.A. 71, 3537.

    Google Scholar 

  • Kawarai, Y. &Nakane, P. K. (1970) Localization of tissue antigens on the ultrathin sections with peroxidase labelled antibody method.J. Histochem. Cytochem. 18, 161.

    Google Scholar 

  • Kraehenbuhl, J. P. &Jamieson, J. D. I. (1972) Solid phase conjugation of ferritin to Fab fragments for use in antigen localization on thin sections.Proc. natn. Acad. Sci. U.S.A. 69, 117.

    Google Scholar 

  • Kraehenbuhl, J. P. &Jamieson, J. D. (1974) Localization of intracellular antigens by immunoelectron microscopy.Int. Rev. Expl. Pathol. 13, 1.

    Google Scholar 

  • Kraehenbuhl, J. P., Racine, L. &Jamieson, J. D. (1977) Immunocytochemical localization of secretory proteins in bovine pancreatic exocrine cells.J. Cell Biol. 72, 406.

    Google Scholar 

  • Maclean, J. D. &Singer, S. J. (1970) A general method for the specific staining of intracellular antigens with ferritin-conjugates.Proc. natn. Acad. Sci. U.S.A. 65, 122.

    Google Scholar 

  • Marchis-Mouren, G. (1965) Etude comparée de l'équipement enzymatique du suc pancréatique de diverses espèces.Bul. Soc. Chim. biol. 47, 2207.

    Google Scholar 

  • Mazurkiewic, J. E. &Nakane, P. K. (1972) Light and electron microscopic localization of antigens in tissues embedded in polyethylene glycol with a peroxidase-labelled antibody method.J. Histochem. Cytochem. 20, 969.

    Google Scholar 

  • Palade, G. E. (1975) Intracellular aspects of the process of protein secretion.Science, N.Y. 189, 347.

    Google Scholar 

  • Nakane, P. K. &Pierce, G. B. (1966) Enzyme-labelled antibodies: preparation and application for the localization of antigens.J. Histochem. Cytochem. 14, 929.

    Google Scholar 

  • Papermaster, D. S., Schneider, B. G., Zorn, M. K. &Kraehenbuhl, J. P. (1978a) Immunocytochemical localization of opsin in outer segments and Golgi zones of frog photoreceptor cells. An electron microscope analysis of cross-linked albumin-embedded retina.J. Cell Biol. 77, 196.

    Google Scholar 

  • Papermaster, D. S., Schneider, B. G., Zorn, M. A. &Kraehenbuhl, J. P. (1978b) Immunocytochemical localization of a large membrane protein to the incisures and margins of frog retinal rod outer segment disks.J. Cell Biol. 78, 415.

    Google Scholar 

  • Roth, J., Bendayan, M. &Orci, L. (1978) Ultrastructural localization of intracellular antigens by the use of protein A-Gold complex.J. Histochem. Cytochem. 26, 1074.

    Google Scholar 

  • Stöström, M. &Squire, J. M. (1977) Cryo-ultramicrotomy and myofibrillar fine structure: a review.J. Microscopy 111, 239.

    Google Scholar 

  • Sternberger, L. A. (1974)Immunochemistry. Engelwood Cliffs, New Jersey: Prentice-Hall.

    Google Scholar 

  • Sternberger, L. A. &Cuculis, J. J. (1969) Methods for enzymatic intensification of the immunocytochemical reaction without use of labeled antibodies.J. Histochem. Cytochem. 17, 190.

    Google Scholar 

  • Tokuyasu, K. (1973) A technique for ultracryotomy of cell suspensions and tissues.J. Cell Biol. 57, 551.

    Google Scholar 

  • Tokuyasu, K. T. (1978a) Immunoferritin localization of intracellular antigens in positively stained frozen sections.Ninth Int. Congr. electron Microsc. 2, 168.

    Google Scholar 

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

    Google Scholar 

  • Tokuyasu, K. T. &Singer, S. S. (1976) Improved procedures for immunoferritin labelling of ultrathin frozen sections.J. Cell Biol. 71, 894.

    Google Scholar 

  • Weibel, E. R. &Bolender, P. (1973) Stereological techniques for electron microscopic morphometry. InPrinciples and Techniques of Electron Microscopy (edited byHayat, M.), Vol. 3. p. 237, New York: Van Nostrand Reinhold.

    Google Scholar 

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Authors and Affiliations

  1. Institut de Biochimie, Université de Lausanne, 1066, Epalinges, Switzerland

    J. P. Kraehenbuhl & L. Racine

  2. European Molecular Biology Laboratory, Postfach 102209, Heidelberg, Germany

    G. W. Griffiths

Authors
  1. J. P. Kraehenbuhl
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  2. L. Racine
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  3. G. W. Griffiths
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Kraehenbuhl, J.P., Racine, L. & Griffiths, G.W. Attempts to quantitate immunocytochemistry at the electron microscope level. Histochem J 12, 317–332 (1980). https://doi.org/10.1007/BF01006953

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  • DOI: https://doi.org/10.1007/BF01006953

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