Summary
It was demonstrated that microwave energy used simultaneously in combination with low concentrations of glutaraldehyde (0.05%) and formaldehyde (2.0%) rapidly preserved light microscopic histology and excellent fine structural details, as well as a variety of cytoplasmic and membrane-bound antigens. Specimen blocks up to 1 cm3 can be fixed in as brief a time as 26 ms using a specially designed microwave device (ultrafast microwave fixation method). The fast microwave fixation method, using a commercially available device, was successfully used to preserve granule-bound rat mast cell chymase which was subsequently detected by a postembedding immunogold procedure. Control of the following parameters is important to the microwave fixation method: (1) specimens with one dimension less than 1 cm; (2) irradiation temperatures lower than 50°C; (3) irradiation times less than 50 s; (4) immediate replacement of the postirradiation solution with cold storage buffer; (5) fixing the specimen within 15 min after it is removed from its blood supply.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Battifora, H. &Kopinski, M. (1986) The influence of protease digestion and duration of fixation on the immunostaining of keratins. A comparison of formalin and ethanol fixation.J. Histochem. Cytochem. 34, 1095–100.
Bendayan, M. &Zollinger, M. (1983) Ultrastructural localization of antigenic sites on osmium-fixed tissues applying the protein A-gold technique.J. Histochem. Cytochem. 31, 101–9.
Benditt, E. P. &Arase, M. (1959) An enzyme in mast cells with properties like chymotrypsin.J. Exp. Med. 110, 451–60.
Bernard, G. R. (1974) Microwave irradiation as a generator of heat for histological fixation.Stain Technol. 49, 215–24.
Boon, M. E., Kok, L. P. &Ouwerkerk-Noordam, E. (1986) Microwave-stimulated diffusion for fast processing of tissue: reduced dehydrating, clearing, and impregnating times.Histopathology 10, 303–9.
Brinn, N. T. (1983) Rapid metallic histological staining using the microwave oven.J. Histochnol. 6, 125–9.
Chew, E. C., Riches, D. J., Lam, T. K. &Hou Chan, H. J. (1983) A fine structural study of microwave fixation of tissues.Cell Biol. Int. Rep. 7, 135–9.
Chiu, K. Y. &Chan, K. W. (1987) Rapid immunofluorescence staining of humam ranal biopsy specimens using microwave irradiation.J. Clin. Pathol. 40, 689–92.
Clements, J. R., Beitz, A. J. (1985) The effects of different pretreatment conditions and fixation regimes on serotonin immunoreactivity: a quantitative light microscopic study.J. Histochem. Cytochem. 33, 778–84.
Dvorak, A. M., Klebanoff, S. J., Henderson, W. R., Monahan, R. A., Pyne, K. &Galli, S. J. (1985a) Vesicular uptake of eosinophil peroxidase by guinea pig basophils and by cloned mouse mast cells and granule-containing lymphoid cells.Am. J. Pathol. 118, 425–38.
Dvorak, A. M., Schulman, E. S., Peters, S. P., Macglashan, D. W., Jr, Newball, H. H., Schleimer, R. P. &Lichtenstein, L. M. (1985b) Immunoglobulin E-mediated degranulation of isolated human lung mast cells.Lab. Invest. 53, 45–56.
Ellar, D. J., Munoz, E. &Salton, M. R. J. (1971) The effect of low concentrations of glutaraldehyde on Micrococcus lysodeikticus membranes: changes in the release of membrane-associated enzymes and membrane structure.Biochim. Biophys. Acta 225, 140–50.
Enerbäck, L. &Svensson, I. (1980) Isolation of rat peritoneal mast cells by centrifugation on density gradients of Percoll.J. Immunol. Methods 39, 135–45.
Estrada, J. C., Brinn, N. T. &Bossen, E. H. (1985) A rapid method of staining ultrathin sections for surgical pathology TEM with the use of the microwave oven.Am J. Clin. Pathol. 83, 639–41.
Fox, C. H., Johnson, F. B., Whiting, J. &Roller, P. P. (1985) Formaldehyde fixation.J. Histochem. Cytochem. 33, 845–853.
Comori, G. (1953) Chloracetyl esters as histochemical substrates.J. Histochem. Cytochem. 1, 469–70.
Gordon, H. W. &Daniel, E. J. (1974) Preliminary report: microwave fixation of human tissues.Am. J. Med. Technol. 40, 441–2.
Hafiz, S., Spencer, R. C., Lee, M., Gooch, H. &Duerden, B. I. (1985) Use of microwaves for acid- and alcohol-fast staining.J. Clin. Pathol. 38, 1073–84.
Hand, J. W. (1977) Microwave heating patterns in simple tissue models.Phys. Med. Biol. 22, 981–7.
Hautzer, N. W., Wittkuhn, J. F. &Mccaughey, W. T. E. (1980) Trypsin digestion in immunoperoxidase staining.J. Histochem. Cytochem. 28, 52–3.
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.
Johnson, C. C. &Guy, A. W. (1972) Non-ionizing electromagnetic wave effects in biological materials and systems.Proc. IEEE 60, 692–718.
Jones, D. J. &Stavinoha, W. B. (1979) Microwave inactivation as a tool for studying the neuropharmacology of cyclic nucleotides. InNeuropharmacology of Cyclic Nucleotides (edited byPalmer, G. C.), pp. 253–81 Baltimore: Urban & Schwarzenberg.
Karnovsky, M. J. (1965) A formaldehyde-glutaraldehyde fixative of high osmolality for use in electron microscopy.J. Cell Biol. 27, 137A-8A.
Kok, L. P. (1986) Fundamentals of microwave-stimulated staining. InStandardization and Quantitation of Diagnostic Staining in Cytology (edited byBoon, M. E. &Kok, L. P.), pp. 82–97 Leyden: Coulomb Press.
Kok, L. P., Boon, M. E., Ouwerkerk-Noordam, E. &Gerrits, P. O. (1986) The application of a microwave technique for the preparation of cell blocks from sputum.J. Microsc. 144, 193–9.
Lagunoff, D. &Benditt, E. P. (1963) Proteolytic enzymes of mast cells.Ann. N.Y. Acad. Sci. 103, 185–98.
Leong, A. S. Y., Daymon, M. E. &Milios, J. (1985) Microwave irradiation as a form of fixation for light and electron microscopy.J. Pathol. 146, 313–21.
Leong, A. S. Y.,&Milios, J. (1986) Rapid immunoperoxidase staining of lymphocyte antigens using microwave irradiation.J. Pathol. 148, 183–7.
Login, G. R. (1978) Microwave fixation versus formalin fixation of surgical and autopsy tissue.Am. J. Med. Technol. 44, 435–7.
Login, G. R. &Dvorak, A. M. (1985) Microwave energy fixation for electron microscopy.Am. J. Pathol. 120, 230–43.
Login, G. R., Galli, S. J., Morgan, E., Arizono, N., Schwartz, L. B. &Dvorak, A. M. (1987a) Rapid microwave fixation of rat mast cells. I. Localization of granule chymase with an ultrastructural postembedding immunogold technique.Lab. Invest. 57, 592–9.
Login, G. R., Schnitt, S. J. &Dvorak, A. M. (1987b) Rapid microwave fixation of human tissues for light microscopic immunoperoxidase identification of diagnostically useful antigens.Lab. Invest. 57, 585–91.
Login, G. R., Stavinoha, W. B. &Dvorak, A. M. (1986) Ultrafast microwave energy fixation for electron microscopy.J. Histochem. Cytochem. 34, 381–7.
Login, G. R., Stavinoha, W. B. &Dvorak, A. M. (1988) Fast and ultrafast microwave energy fixation techniques for immunofluorescence, and light and electron microscopy. InProceedings of the 4th International Congress of Toxicology—Satellite Symposium on the Application of Microwave Irradiation, 26 July 1986 (edited byBlank C. L. &Maruyama, Y.) Tokyo: Soft Science Publishers, in press.
Mayers, C. P. (1970) Histological fixation by microwave heating.J. Clin. Pathol. 23, 273–5.
Mcclean, V. E. R., Sheppard, R. J. &Grant, E. H. (1981) A generalized model for the interaction of microwave radiation with bound water in biological material.J. Microwave Power 16, 1–7.
Mclees, B. D. &Finch, E. D. (1973) Analysis of reported physiologic effects of microwave radiation. InAdvances in Biological and Medical Physics, Vol. 14 (edited byLawrence, J. H. &Gofman, J. W.), pp. 163–223 New York: Academic Press.
Mepham, B. L. (1982) Influence of fixatives on the immunoreactivity of paraffin sections.Histochem. J. 14, 731–7.
Merritt, J. H. &Frazer, J. W. (1977) Microwave fixation of brain tissue as a neurochemical technique—a review.J. Microwave Power 12, 133–9.
Patterson, M. K., Jr &Bulard, R. (1980) Microwave fixation of cells in tissue culture.Stain Technol. 55, 71–5.
Pinkus, G. S., O'Connor, E. M., Etheridge, C. L. &Corson, J. M. (1985) Optimal immunoreactivity of keratin proteins in formalin-fixed, paraffin-embedded tissue requires preliminary trypsinization. An immunoperoxidase study of various tumours using polyclonal and monoclonal antibodies.J. Histochem. Cytochem. 33, 465–73.
Ruoff, H. J. (1977) Cyclic AMP and cyclic GMP in the rat gastric mucosa: the significance of post-mortem changes and tissue fixation on the estimation of thein vivo level.Naunyn Schmiedebergs Arch. Pharmacol. 298, 167–73.
Schick, B., Austen, K. F. &Schwartz, L. B. (1984) Activation of rat serosal mast cells by chymase, an endogenous secretory granule protease.J. Immunol. 132, 2571–7.
Schmidt, M. J., Schmidt, D. E. &Robison, G. A. (1971) Cyclic adenosine monophosphate in brain areas: microwave irradiation as a means of tissue fixation.Science 173, 1142–3.
Schnitt, S. J. &Vogel, H. (1986) Meningionmas. Diagnostic value of immunoperoxidase staining for epithelial membrane antigen.Am. J. Surg. Pathol. 10, 640–9.
Seppa, H., Vaananen, K. &Korhonen, K. (1979) Effect of mast cell chymase of rat skin on intercellular matrix: a histochemical study.Acta Histochem. 64, 64–70.
Stavinoha, W. B. (1983) Study of brain neurochemistry utilizing rapid inactivation of brain enzyme activity by heating with microwave irradiation. InMicrowave Fixation of Labile Metabolites, Vol. 45,Advances in Bioscience (edited byBlank, C. L., Stavinoha, W. B. &Maruyama, Y.), pp. 1–12. Oxford: Pergamon Press.
Tovar, C. A., Armati, P. J. &Pollard, J. D. (1987) Microwave fixation of whole peripheral nerve for rapid and efficient enzyme-linked immunosorbent assay (EL-ISA) screening of monoclonal antibodies.J. Immunol. Methods 94, 127–30.
Wakabayashi, T., Asano, M., Kurono, C. &Kimura, H. (1975) Effects of extremely low concentrations of glutaraldehyde together with shortening of fixation times on the maintenance of various enzyme activities.J. Histochem. Cytochem. 23, 632–4.
Wintroub, B. U., Schechter, N. B., Lazarus, G. S., Kaempfer, C. E. &Schwartz, L. B. (1984) Angiotensin I conversion by human rat chymotryptic proteinases.J. Invest. Dermatol. 83, 336–9.
Yurt, R. &Austen, K. F. (1977) Preparative purification of the rat mast cell chymase.J. Exp. Med. 146, 1405–19.
Zimmerman, G. R. &Raney, J. A. (1972) Fast fixation for surgical pathology specimens.Lab. Med. 3, (12), 29–30.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Login, G.R., Dvorak, A.M. Microwave fixation provides excellent preservation of tissue, cells and antigens for light and electron microscopy. Histochem J 20, 373–387 (1988). https://doi.org/10.1007/BF01002732
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF01002732