Summary
Acid phosphatase activity has been measured in cultured human gingival fibroblasts using a validated histochemical simultaneous coupling semi-permeable membrane technique. The histochemical reaction was linear over a three hour incubation period and had a pH optimum of 5.0. The activity was not increased by prior exposure to hypotonic acetate buffer and was inhibited by fluoride and molybdate but not by formaldehyde. These results indicate that the semi-permeable membrane technique described may be used for observing and measuring acid phosphatase activity in cultured fibroblasts. From results obtained using inhibitors, it appears that in these cells most of the acid phosphatase observed is lysosomal. The absence of any activation of activity following pre-incubation with hypotonic buffer indicates that the method is not suitable for monitoring lysosomal membrane function.
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Altman, F. P. (1980) tissue stabilizer methods in histochemistry. InTrends in Enzyme Histochemistry and Cytochemistry, Ciba Foundation Symposium, pp. 81–102. Amsterdam: Excerpta Medica.
Arvidson, K., Cottler-Fox, M. &Friberg, U. (1980) The effects of dental root posts on human gingival fibroblastsin vitro.J. dent. Res. 59, 651–6.
Bitensky, L. (1980). Microdensitometry. InTrends in Enzyme Histochemistry and Cytochemistry, Ciba Foundation Symposium, pp. 181–202. Amsterdam: Excerpta Medica.
Goldstein, D. J. (1971) Aspects of scanning microdensitometry. II. Spot size, focus and resolution.J. Microscopy 93, 15–42.
Half, T. W. &Wenzel, D. G. (1978) A quantitative method for measurement of lysosomal acid phosphatase latency in cultured rat heart cells with210Pb.Histochem. J. 10, 505–15.
Heath, J. K., Gowen, M., Meikle, M. C. &Reynolds, J. J. (1982) Human gingival tissues in culture synthesize three metaloproteinases and a metaloproteinase inhibitor.J. Perio. Res. 17, 183–90.
Lake, B. D. &Ellis, R. B. (1976) What do you think you are quantifying? An appraisal of histochemical methods in the measurement of the activities of lysosomal enzymes.Histochem. J. 8, 357–66.
Maggi, V. &Carbonell, A. W. (1969) Lysosomes and acid phosphatases during growth and differential in mice: a light and electron microscope study.Histochem. J. 1, 383–403.
Maggi, V. &Chng, W. L. (1966) A study of acid phosphatase in mouse kidney using naphthol AS/BI phosphate as a substrate.Histochemie 7, 267–72.
Mandys, V. &Elleder, M. (1980) Demonstration of enzymes in cells cultured on semipermeable membrane in a double chamber.Histochemistry 65, 325–7.
Meijer, A. E. F. H. (1972) Semipermeable membranes for improving the histochemical demonstration of enzyme activities in tissue sections. 1. Acid phosphatase.Histochemie 30, 31–9.
Meijer, A. E. F. H. (1980) Semipermeable membrane techniques in quantitative enzyme histochemistry. InTrends in Enzyme Histochemistry and Cytochemistry, Ciba Foundation Symposium, pp. 103–20. Amsterdam: Excerpta Medica.
Neil, M. W. &Horner, M. W. (1964) Studies on acid hydrolases in adult and foetal tissues.Biochem. J. 92, 217–24.
Stoward, P. J. &Al-Sarraj, B. (1981a) Quantitative histochemical investigations of semi-permeable membrane techniques for the assay of acid phosphatase in skeletal muscle. I. Meijer's Method.Histochemistry 71, 405–31.
Stoward, P. J. &Al-Sarraj, B. (1981b) Quantitative histochemical investigations of semi-permeable membrane techniques for the assay of acid phosphatase in skeletal muscle. III. A modified simultaneous coupling technique.Histochemistry 71, 599–607.
Tyas, M. J. (1978) Microdensitrometric quantification of the lead capture method for the histochemical demonstration of acid phosphatase in cultured cells.Histochem. J. 10, 239–45.
Yoffe, J. R. (1980) A comparison of two cytochemical methods used to determine lysosomal function in cultured endothelial cells.Histochem. J. 12, 537–42.
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Duguid, R., Rumgay, L.M.A. Acid phosphatase activity of human gingival fibroblasts measured using a simultaneous coupling semi-permeable membrane technique. Histochem J 15, 815–824 (1983). https://doi.org/10.1007/BF01003344
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DOI: https://doi.org/10.1007/BF01003344