Summary
The reaction rate of glucose-6-phosphate dehydrogenase activity in liver sections from fed and starved rats has been monitored by the continuous measurement at 37 C of the reaction product as it is formed using scanning and integrating microdensitometry. Control media lacked either substrate or both substrate and coenzyme. All reactions were nonlinear; however, subtraction of either of the controls from the test response produced linearity. Differing responses in sections of livers from fed and fasted rats indicate that the appropriate control medium for use in the assay of this dehydrogenase is one lacking both substrate and coenzyme rather than a medium containing coenzyme. The reaction rate was the same with each of the final acceptors. Problems with the diffusion of the formazan of BPST and with the failure to precipitate the formazan of Neotetrazolium make Tetranitro BT and Nitro BT the tetrazolium salts of choice in quantitative dehydrogenase assays.
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References
Altman, F. P. (1976a) The quantification of formazans in tissue sections by microdensitometry. II. The use of BPST, a new tetrazolium salt.Histochem. J. 8, 501–6.
Altman, F. P. (1976b) The quantification of formazans in tissue sections by microdensitometry. III. The effect of objective power and scanning spot size.Histochem. J. 8, 507–11.
Altman, F. P. &Butcher, R. G. (1973) Studies on the reduction of tetrazolium salts. I. The isolation and characterisation of a half-formazan intermediate produced during the reduction of neotetrazolium chloride.Histochemistry 37, 333–50.
Butcher, R. G. (1972) Precise cytochemical measurement of neotetrazolium formazan by scanning and integrating microdensitometry.Histochemistry 32, 171–90.
Butcher, R. G. (1978a) Oxygen and the production of formazan from neotetrazolium chloride.Histochemistry 56, 329–40.
Butcher, R. G. (1978b) The measurement in tissue sections of the two formazans derived from nitroblue tetrazolium in dehydrogenase reactions.Histochem. J. 10, 739–44.
Butcher, R. G. (1979) The oxygen insensitivity phenomenon as a diagnostic aid in carcinoma of the bronchus. InQuantitative Cytochemistry and its Applications (edited byPattison, J. R., Bitensky, L. andChayen, J.), pp. 241–51, London: Academic Press.
Butcher, R. G. (1984) Reaction rate studies of glucose-6-phosphate dehydrogenase in rat tracheal epithelium; the effect of section thickness.Histochemistry 81, 567–72.
Butcher, R. G. &Altman, F. P. (1973) Studies on the reduction of tetrazolium salts. II. The measurement of the half-reduced and fully reduced formazans of neotetrazolium chloride in tissue sections.Histochemistry 37, 351–63.
Butcher, R. G., Dawson, A. L., Knaab, S. A. &Gahan, P. B. (1980) Dehydrogenase activity and loss of formazan from tissue sections.Histochem. J. 12, 591–8.
Delong, A. S. H. (1982) Mechanisms of metal salt methods in enzyme cytochemistry with special reference to acid phosphatase.Histochem. J. 14, 1–33.
Evans, A. W. (1979)Quantitative cytochemistry of glucose-6-phosphate, lactate and succinate dehydrogenases in oral mucosa during hamster cheek pouch cardinogenesis and in human premalignant and malignant lesions. PhD thesis, University of London.
Gahan, P. B. &Dawson, A. L. (1981) Problems encountered with BPST in dehydrogenase histochemistry.Histochem. J. 13, 338.
Goldstein, D. J. (1981) Errors in microdensitometry.Histochem. J. 13, 251–363.
Henderson, B., Loveridge, N. &Robertson, W. R. (1978) A quantitative study of the effects of different grades of polyvinyl alcohol on the activities of certain enzymes in unfixed tissue sections.Histochem. J. 10, 453–63.
Ibrahim, K. S., Husain, O., Bitensky, L. &Chayen, J. (1983) A modified tetrazolium reaction for identifying malignant cells from gastric and colonic cancer.J. clin. Path. 36, 133–6.
Kugler, P. (1981) Kinetic and morphometric measurements of enzyme reactions in tissue sections with a new instrumental setup.Histochemistry 71, 433–49.
Nolte, J. &Pette, D. (1971) Quantitative microscope-photometric determination of enzyme activities in cryostat sections. InRecent Advances in Quantitative Histo- and Cytochemistry (edited byDubach, U. C. andSchmidt, U.), pp. 54–60. Bern, Stuttgart, Vienna: Hans Huber.
Nolte, J. &Pette, D. (1972) Microphotometric determination of enzyme activity in single cells in cryostat sections. I. Application of the gel film technique to microphotometry and studies on the intralobular distribution of succinate dehydrogenase and lactate dehydrogenase activities in rat liver.J. Histochem. Cytochem. 20, 567–76.
Pette, D. (1981) Microphotometric measurements of initial maximum reaction rates in quantitative enzyme histochemistryin situ.Histochem. J. 13, 319–27.
Pette, D., Wasmund, H. &Wimmer, M. (1979) Principle and method of kinetic microphotometric enzyme activity determinationin situ.Histochemistry 64, 1–10.
Pette, D. &Wimmer, M. (1979) Kinetic microphotometric activity determination in enzyme containing gels and model studies with tissue sections.Histochemistry 64, 11–22.
Pette, D. &Wimmer, M. (1980) Microphotometric determination of enzyme activities in cryostat sections by the gel film technique. InTrends in Enzyme Histochemistry and Cytochemistry. Ciba Foundation Symposium, Vol. 73 (new series), pp. 121–34. Amsterdam, Oxford, New York: Excerpta Medica.
Potter, V. R., Gebert, R. A. &Pitot, H. C. (1966) Enzyme levels in rats adapted to 36-hour fasting.Adv. Enzyme Reg. 4, 247–65.
Racker, E. (1955) Mechanism of action and properties of pyridine nucleotide-linked enzymes.Physiol. Rev. 35, 1–56.
Rieder, H. (1981) NADP-dependent dehydrogenases in rat liver parenchyma. III. The description of a lipogenic area on the basis of histochemically demonstrated enzyme activities and the neutral fat content during fasting and refeeding.Histochemistry 72, 579–615.
Rieder, H., Teutsch, H. F. &Sasse, D. (1978) NADP-dependent dehydrogenases in rat liver parenchyma. I. Methodological studies on the qualitative histochemistry of G6PDH, 6PGDH, malic enzyme and ICDH.Histochemistry 56, 283–98.
Robertson, W. R., Davison, B., Gandy, M., Burdge, R. &Lambert, A. (1984) Computerized continuous monitoring of cytochemical enzyme reaction product formation by the Vickers M85A microdensitometer.Histochem. J. 16, 651–6.
Sinowatz, F., Scheubeck, M., Wrobel, K. H. &Zwack, M. (1983) Histochemical localization and quantification of glucose-6-phosphate dehydrogenase in bovine Leydig cells.Histochem. J. 15, 831–44.
Van Noorden, C. J. F. (1984) Histochemistry and cytochemistry of glucose-6-phosphate dehydrogenase.Prog. Histochem. Cytochem. 15/4, 1–85.
Van Noorden, C. J. F. &Butcher, R. G. (1984) Histochemical localization of NAD(P)-dependent dehydrogenase activity with four different tetrazolium salts.J. Histochem. Cytochem. 32, 998–1004.
Van Noorden, C. J. F., Vogels, I. M. C., James, J. &Tas, J. (1982) A sensitive cytochemical staining method for glucose-6-phosphate dehydrogenase activity in individual erythrocytes.Histochemistry 75, 493–506.
Van Noorden, C. J. F., Tas, J., Vogels, I. M. C. (1983) Cytophotometry of glucose-6-phosphate dehydrogenase activity in individual cells.Histochem. J. 15, 583–99.
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Butcher, R.G., Van Noorden, C.J.F. Reaction rate studies of glucose-6-phosphate dehydrogenase activity in sections of rat liver using four tetrazolium salts. Histochem J 17, 993–1008 (1985). https://doi.org/10.1007/BF01417948
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DOI: https://doi.org/10.1007/BF01417948