Skip to main content
Log in

A fluorescent redox dye. Influence of several substrates and electron carriers on the tetrazolium salt—formazan reaction of Ehrlich ascites tumour cells

The Histochemical Journal Aims and scope Submit manuscript

Summary

This study was performed to elaborate the best conditions for measuring the redox activity of Ehrlich ascites tumour cells by using a new tetrazolium salt, cyantolyl tetrazolium chloride (CTC). This tetrazolium salt forms a fluorescent water-insoluble formazan on reduction on the surface of intact vital cells. The influences of fixation and of various substrates and electron carriers on the cellular reduction of CTC were investigated quantitatively using an elution technique. The amount of formazan obtained after incubating vital cells with Meldola Blue as electron carrier was greater than that obtained with Methylene Blue, menadione, 2,6-dichloroindophenol, 1-methoxyphenazine methosulphate or phenazine methosulphate. Using flow cytometry, the formazan production per cell and, after staining the nuclear DNA, the distribution of the redox activity in the cell population can be visualized with satisfactory resolution. We conclude from our findings that dehydrogenases are only partially involved in the reduction of tetrazolium salts by intact cells and that a redox activity, probably related to a cell membrane-bound NAD(P)H—oxidase system, is mainly measured.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

References

  • ALTMAN, F. P. (1972) Quantitative dehydrogenase histochemistry with special reference to the pentose shunt dehydrogenases.Prog. Histochem. Cytochem. 4, 225–73.

    Google Scholar 

  • ALTMAN, F. P. (1976) Tetrazolium salts and formazans.Prog. Histochem. Cytochem. 9, 1–56.

    Google Scholar 

  • ALTMAN, F. P. (1981) Some aspects of the use of tetrazolium salts in quantitative histochemistry.Acta histochemia Suppl. XXIV, 221–30.

    Google Scholar 

  • ALTMAN, F. P. & BUTCHER, R. G. (1973) Studies on the reduction of tetrazolium salts. I. The isolation and characterization of a half-formazan intermediate product during the reduction of neotetrazolium chloride.Histochemie 37, 333–50.

    Google Scholar 

  • BARKA, T. & DALLNER, G. (1957) The reduction of tetrazolium salts by ascites tumour cells in the presence of glucose.J. Histochem. Cytochem. 6, 174–80.

    Google Scholar 

  • BENGTSSON, G. & ANDERSSON, G. (1981) The effect of Ehrlich ascites tumour growth on lactate dehydrogenase activity in tissues and physiological fluids of tumour bearing mice.Int. J. Biochem. 13, 53–61.

    Google Scholar 

  • BRIGGS, R. T., DRATH, D. B., KARNOVSKY, M. L. & KARNOVSKY, M. J. (1975) Localization of NADH oxidase on the surface of human polymorphonuclear leukocytes by a new cytochemical method.J. Cell Biol. 67, 566–86.

    Google Scholar 

  • 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.Histochemie 37, 351–63.

    Google Scholar 

  • ECHTERMEYER, V. (1973) Zur Quantifizierung topochemischer Tetrazoliumreaktionen.Histochemie 36, 35–7.

    Google Scholar 

  • KUGLER, P. (1982) Quantitative dehydrogenase histochemistry with exogenous electron carriers, (PMS, MPMS, MB).Histochemistry 75, 95–112.

    Google Scholar 

  • LIPPOLD, H. J. (1982) Quantitative succinate dehydrogenase histochemistry. A comparison of different tetrazolium salts.Histochemistry 76, 381–407.

    Google Scholar 

  • SCHÜNZEL, G. & KUHNE, H. (1982) Quantitative Untersuchungen zur Empfindlichkeitssteigerung der Tetrazolium-Methoden für den histochemischen Dehydrogenasen-Nachweis.Acta histochem. Suppl. XXVI, 167–71.

    Google Scholar 

  • SEIDLER, E. (1982) The importance of the superoxide dismutases in histochemistry of dehydrogenases.Acta histochem. 66, 106–14.

    Google Scholar 

  • SEIDLER, E. & WENK, H. (1982) Zur Eignung von Monotetrazoliumsalzen für die Quantifizierung histochemischer Dehydrogenasereaktionen.Acta histochem. Suppl. XXVI, 157–60.

    Google Scholar 

  • SEVERIN, E. & STELLMACH, J. (1984) Flow cytometry of redox activity of single cells using a newly synthesized fluorescent formazan.Acta histochem. 75, 101–5.

    Google Scholar 

  • SEVERIN, E. & STELLMACH, J. (1985) Fluorimetric assay of redox activity in cells.Anal. Chim. Acta 170, 341–6.

    Google Scholar 

  • STELLMACH, J. (1984) Fluorescent redox dyes. I. Production of fluorescent formazan by unstimulated and phorbol ester or digitonin-stimulated Ehrlich ascites tumor cells.Histochemistry 80, 137–43.

    Google Scholar 

  • STELLMACH, J. & SEVERIN, E. (1984) Fluoreszierende Redoxindikatoren und ihre Anwendung in der Cytodiagnostik. German patent no. DE OS 34 18 852.

  • VAN BERKEL, T. C. J. & KRUIJT, J. K. (1977) Distribution and some properties of NADPH and NADH oxidase in parenchymal and nonparenchymal liver cells.Arch. Biochem. Biophys. 179, 8–14.

    Google Scholar 

  • VAN NOORDEN, C. J. F & TAS, J. (1982) The role of exogenous electron carriers in NAD(P)-dependent dehydrogenase cytochemistry studiedin vitro and with a model system of polyacrylamide films.J. Histochem. Cytochem. 30, 12–20.

    Google Scholar 

  • VAN NOORDEN, C. J. F., KOOIJ, A., VOGELS, I. M. C. & FREDERIKS, W. M. (1985) On the nature of the ‘nothing dehydrogenase’ reaction.Histochem, J. 17, 1111–8.

    Google Scholar 

  • WOHLRAB, F., SEIDLER, E. & KUNZE, K. D. (1979) ‘Histound Zytochemie dehydrierender Enzyme’, p. 216. Leipzig: J. A. Barth.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Stellmach, J., Severin, E. A fluorescent redox dye. Influence of several substrates and electron carriers on the tetrazolium salt—formazan reaction of Ehrlich ascites tumour cells. Histochem J 19, 21–26 (1987). https://doi.org/10.1007/BF01675289

Download citation

  • Received:

  • Revised:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF01675289

Keywords

Navigation