Summary
Microwave irradiation has been applied to reduce the immunogold staining time of ultrathin sections of Lowicryl embedded specimens. Labelling has been stimulated by microwave irradiation during incubation with 10nm colloidal gold particls coated with either goat anti-mouse antibodies (GaM-gold) or goat anti-rabbit antibodies (GaR-gold) and has been compared with control incubations. Quantification has been performed on cytoplasmic membranes or lysosomes labelled with a primary antibody. Counting the gold particles over specific and non-specific sites in electron micrographs and electron microscopic images by IBAS 2000 revealed that irradiation of 25 μl droplets both at 80W and 150 W resulted in an accelerated immunogold labelling, while the non-specific background levels were not increased. A plateau level in immunogold labelling intensity was reached after 25 min incubation under microwave irradiation at 150W as compared to 120 min incubation without microwaves. No improvement in localization sharpness of immunogold labelling on membranes was achieved by microwave irradiation. The microwave-mediated acceleration of immunogold staining may be considered as an example of a staining method with a restricted thermal action on microvolumes as indicated by direct temperature measurements using a fibre-optic thermometer.
Similar content being viewed by others
References
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.
Boon, M. E. &Kok, L. P. (editors) (1987) In:Microwave Cookbook of Pathology: The art of microscopic visualization. Ch 5 & 6. Leiden: Coulomb Press, Leyden.
Chiu, K. Y. (1987) Use of microwaves for rapid immunoperoxidase staining of paraffin sections.Med Lab. Sci. 44, 3–5.
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.
Geuze, H. J., Slot, J. W., Van der Ley, P. A., Scheffer, R. C. T. &Griffith, J. M. (1981) Use of colloidal gold particles in double labelling immunoelectron microscopy of ultrathin frozen tissue sections.J. cell. Biol. 89, 653–65.
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.
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, Y. (1986) Rapid immunoperoxidase staining of lymphocyte antigens using microwave irradiation.J. Pathol. 148, 183–7.
Login, G. R. &Dvorak, A. M. (1985) Microwave energy fixation for electron microscopy.Am. J. Pathol. 120, 230–4.
Login, G. R., Stavinoha, W. B. &Dvorak, A. M. (1986) Ultrafast microwave energy fixation for electron microscopy.J. Histochem. Cytochem. 34, 381–7.
Van der Kwast, Th. H., Versnel, M. A., Delahaye, M., De Jong, A., Zondervan, P. E. &Hoog-Steden, H. (1988) Expression of epithelial membrance antigen on malignant mesothelioma cells: an immunocytochemical and immunoelectron microscopic study.Acta Cytologica 32, 169–74.
Zondervan, P. E., van der Kwast, Th. H., De Jong, A. Visser, W. J. &De Bruijn, W. C. (1986) Lysosomal localization of secretory prostatic acid phosphatase in human hyperplastic prostate epithelium.Urol. Res. 14, 331–5.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Zondervan, P.E., De Jong, A., Sorber, C.W.J. et al. Microwave-stimulated incubation in immunoelectron microscopy: A quantitative study. Histochem J 20, 359–364 (1988). https://doi.org/10.1007/BF01002730
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF01002730