Quantitative cytochemistry of alkaline phosphatase activity

Howat, D. D. C.; Chayen, J.

doi:10.1002/cbf.290010206

Cited by 2 publications

(1 citation statement)

References 19 publications

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“…The reaction medium contains sufficient sodium azide to largely inhibit any GTPase activity in this tissue. Despite the fact that the reaction was done at pH 7.4, which is far from the optimal pH for alkaline phosphatase activity (pH 9.4 in these sections) (Howat and Chayen, 1983) and for acid phosphatase activity (pH 4.5 to 5.5). (Morton, 1961) we considered it advisable to measure whether the specific inhibitors for these enzyme activities had any effect on the amount of reaction product generated in the presence of GTP.…”

Section: Discussionmentioning

confidence: 98%

Histochemistry of guanylate cyclase activity.

Mehdizadeh

O'farrell

Bitensky

et al. 1995

J Histochem Cytochem.

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Guanylate cyclase liberates pyrophosphate from guanosine triphosphate (GTP). In studies published previously, this phosphate is trapped by lead ions even though it is known that free lead ions inactivate a considerable proportion of this enzymatic activity. To overcome the damaging effects of fixation, this study used fresh cryostat sections stabilized with a sufficient concentration of a collagen-derived polypeptide to ensure no measurable loss of guanylate cyclase activity. To avoid the damaging influence of free lead ions, we used a hidden metal capture reagent, i.e., a complex of lead ammonium citrate/acetate that does not react with GTP but which rapidly forms a precipitate with the pyrophosphate liberated by the enzyme. The lead precipitate is then converted into the colored sulfide which is measured in individual cells by microdensitometry. This system was used to measure guanylate cyclase activity in individual cells in unfixed sections of rat liver.

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Section: Discussionmentioning

confidence: 98%

Histochemistry of guanylate cyclase activity.

Mehdizadeh

O'farrell

Bitensky

et al. 1995

J Histochem Cytochem.

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Enzyme histochemical reactions in unfixed and undecalcified cryostat sections of mouse knee joints with special reference to arthritic lesions

Noorden

Vogels

1986

Histochemistry

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The use of unfixed and undecalcified cryostat sections of mouse knee joints is described for the study of enzyme histochemical reactions. Non-inflamed knee joints and knee joints of mice with antigen induced arthritis have been used. Joints were embedded in gelatin and subsequently cut at low speed with a motor-driven cryostat fitted with a tungsten carbide knife at an obtuse angle (10 degrees). The sections were attached to transparent tape to keep the integrity of the tissue intact. The following histochemical reactions were carried out successfully: the tetrazolium salt reaction for dehydrogenase and reductase activity, the post-azo-coupling method for acid phosphatase and cathepsin B activity and the simultaneous azo-coupling method for esterase activity. In all cases the morphology and integrity of the sections were well kept and serial sections were obtained without any difficulty. Nonspecific staining of the tape did not occur. The localization of the final reaction product was meeting criteria for specific and precise histochemical methods with the exception of the metal salt method because of nonspecific staining of undecalcified bone. Cytophotometry of the final reaction product appeared to be reproducible and valid as demonstrated by reaction for glucose-6-phosphate dehydrogenase activity in synoviocytes from knee joints with induced arthritis. End point measurements as well as kinetic measurements of the formazan production were performed and linear relationships were found between the specific formazan formation and section thickness or incubation time, respectively. It is concluded that cryostat sections attached to transparent tape are an excellent tool for the study of the metabolism in tissues adjacent to bone matrix.(ABSTRACT TRUNCATED AT 250 WORDS)

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Quantitative cytochemistry of alkaline phosphatase activity

Abstract: Verheyen, A., Borgers, M., Vanparijs, 0. and Thienpont, D.(1 976) Biochemistry of parasites and host-parasite relationships (Van den Boscche, H., Ed.), pp. 605-618. Elsevier/ North-Holland Biomedical Press, Amsterdam

Cited by 2 publications

References 19 publications

Histochemistry of guanylate cyclase activity.

Histochemistry of guanylate cyclase activity.

Enzyme histochemical reactions in unfixed and undecalcified cryostat sections of mouse knee joints with special reference to arthritic lesions

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