Summary
The quantitation of electron dense labelling is very tedious when it is done “by hand”. Accordingly we developed software allowing, at electron microscopic level, a semi-automatic counting of dense markers in biological specimens. It includes the digitization of images and extraction of dense particles from the grey level of the background. The definition of the areas of interest was carried out by the observer but all quantitative calculations were done automatically. This method was applied to different biological materials (phospholipid and lysozyme labelling in secretory granules of human submucosal bronchial gland cells). The results obtained by this semi-automatic procedure were in good agreement with those obtained by manual counting of colloïdal gold labelling (r=0.97).
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References
Bendayan M, Roth J, Perrelet A, Orci L (1980) Quantitative immunocytochemical localization of pancreatic secretory proteins in subcellular compartments of the rat acinar cell. J Histochem Cytochem 28:149–160
Bendayan M, Nanci A, Herbener G, Gregoire S, Duhr M (1986) A review of the study of protein secretion applying the protein gold immunocytochemical approach. Am J Anat 175:379–400
Cau P, Negre-Aminou P, Massacrier A (1989) Immunocytochimie et quantification. In: Calas A, Gounon P, Hernandez-Verdun D, Nicolas G, Reynes M, Tougard C (eds) Immunocytochimie. Méthodes pratiques. Société Française de Microscopie Electronique, Paris, pp 149–160
Coster M, Chermant JL (1985) Precis d'analyse d'images. CNRS, Paris
De Harven E, Soligo D, Christensen H (1987) Should we be counting immunogold marker particles on cell surfaces with the SEM? J Microsc 146:183–189
Desjardins M, Bendayan M (1989) Heterogeneous distribution of type IV collagen, entactin, heparan sulfate proteoglycan, and laminin among renal basement membranes as demonstrated by quantitative immunocytochemistry. J Histochem Cytochem 37:885–897
Girod S, Fuchey C, Galabert C, Lebonvallet S, Bonnet N, Ploton D, Puchelle E (1991) Identification of phospholipids in secretory granules of human submucosal gland respiratory cells. J Histochem Cytochem 32:193–198
Griffiths G, Hoppeler H (1986) Quantitation in immunocytochemistry: correlation of immunogold labeling to absolute number of membrane antigens. J Histochem Cytochem 34:1389–1398
Hinnrasky J, Chevillard M, Puchelle E (1990) Immunocytochemical demonstration of quantitative differences in the distribution of lysozyme in human airway secretory granule phenotypes. Biol Cell 68:239–243
Howell K, Reuter-Carlson U, Devaney E, Luzio JP, Fuller S (1987) One antigen, one gold? A quantitative analysis of immunogold labeling of plasma membrane 5'-nucleotidase in frozen thin sections. Eur J Cell Biol 44:318–327
Ichikawa M, Sasaki K, Ichikawa A (1989) Immunocytochemical localization of amylase in gerbil salivary gland acinar cells processed by rapid freezing and freeze-substitution fixation. J Histochem Cytochem 37:185–194
Kempf J, Kempf C (1982) Autoradiographic grain counting by fully automated image analysis. Microsc Acta 86:215–223
Meyer F (1979) Iterative image transformations for an automatic screening of cervical smears. J Histochem Cytochem 27:128
Nanci A, Slavkin H, Smith C (1987) Immunocytochemical and radioautographic evidence for secretion and intracellular degradation of enamel proteins by amenosblasts during the maturation stage of amelogenesis is rat incisors. Anat Rec 217:107–123
Pratt W (1978) Digital image processing. Wiley, New York Chichester Brisbane
Serra J (1982) Image analysis and mathematical morphology. Academic Press, London Orlando San Diego
Sklarew R (1982) Simultaneous Feulgen densitometry and autoradiographic grain counting with the Quantimet 720D image-analysis system II. Automatic grain counting. J Histochem Cytochem 30:49–57
Weibel ER (1979) Stereological methods, vol I. Practical methods for biological morphometry. Academic Press, London
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Lebonvallet, S., Mennesson, T., Bonnet, N. et al. Semi-automatic quantitation of dense markers in cytochemistry. Histochemistry 96, 245–250 (1991). https://doi.org/10.1007/BF00271542
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DOI: https://doi.org/10.1007/BF00271542