Summary
The sensitivity of mouse liver glucose-6-phosphatase activity towards glutaraldehyde fixation has been analysed by biochemical and cytochemical means. The degree of enzymatic inhibition and various enzymatic properties have been studied. Several differences have been observed in the Km determination, the sensitivity to pH 5 and the activity related to pH between fixed and unfixed enzymes. The role of Pb++ ions in the cytochemical media has also been estimated.
It is concluded that several enzymatic differences appear between fixed and unfixed enzymes and that the inhibition by Pb ions is dependent on the buffer and on the amount of substrate used.
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Allen, J. M.: The histochemistry of glucose-6-phosphatase in the epididymis of the mouse. J. Histochem. Cytochem. 9, 681–689 (1961)
Aloyo, V. J., Geller, A. M., Marcus, C. J., Byrne, W. L.: Properties of glutaraldehyde modified bovine hepatic fructose-1, 6-diphosphatase. Biochim. biophys. Acta (Amst.) 289, 242–246 (1972)
Anderson, P. J.: Purification and quantitation of glutaraldehyde and its effect on several enzyme activities in skeletal muscle. J. Histochem. Cytochem. 15, 652–661 (1967)
Arborgh, B., Ericsson, J. L. E., Helminen, H.: Inhibition of renal acid phosphatase and aryl sulfatase activity by glutaraldehyde fixation. J. Histochem. Cytochem. 19, 449–451 (1971)
Beaufay, H., De Duve, C.: Le système hexose-phosphatasique. IV. Spécificité de la glucose-6-phosphatase. Bull. Soc. Chim. biol. (Paris) 36, 1525–1568 (1954)
Beaufay, H., Hers, H. G., Berthet, J., De Duve, C.: Le système hexose-phosphatasique. V. Influence de divers agents sur l'activité et la stabilité de la glucose-6-phosphatase. Bull. Soc. Chim. biol. (Paris) 36, 1539–1550 (1954)
Bowes, J. H., Cater, C. W.: The reaction of glutaraldehyde with proteins and other biological materials. J. roy. Micr. Soc. 85, 193–200 (1965)
Butcher, R. G.: The estimation of the nucleic acids of tissue sections and its use as a unit of comparison for quantitative histochemistry. Histochemie 13, 262–275 (1968)
Casanova, S., Marchetti, M., Bovina, C., Laschi, R.: A study of the effects of fixation on liver glucose-6-phosphatase activity for electron microscope cytochemistry. J. Submicro. Cytol. 4, 261–270 (1972)
Chabarek, S., Martell, A. E.: Organic sequestering agents. New York: John Wiley & Sons 1959
Chiquoine, A. D.: The distribution of glucose-6-phosphatase in the liver and kidney of the mouse. J. Histochem. Cytochem. 1, 429–435 (1953)
Chiquoine, A. D.: Further studies on the histochemistry of glucose-6-phosphatase. J. Histochem. Cytochem. 3, 471–478 (1955)
Christie, K., Stoward, P.: A quantitative study of the fixation of acid phosphatase by formaldehyde and its relevance to histochemistry. Proc. roy. Soc. Lond. 186, 137–164 (1974)
Cornelisse, C. J.: The localization problem in enzyme cytochemistry. Ph. D. thesis, Rijksuniversitet, Leiden, 80 pages (1974)
Dayan, J., Wilson, I.: The phosphorylation of Tris by alkaline phosphatase. Biochim. biophys. Acta (Amst.) 81, 620–623 (1964)
Dryer, R. L., Tammes, A. R., Routh, J. I.: The determination of phosphorus and phosphatase with N-phenyl-p-phenylene diamine. J. biol. Chem. 225, 171–183 (1957)
Engel, A. G., Tice, L. W.: Cytochemistry of phosphatases of the sarcoplasmic reticulum. I. Biochemical studies. J. Cell Biol. 31, 473–487 (1966)
Ericsson, J. L. E.: On the fine structural demonstration of glucose-6-phosphatase. J. Histochem. Cytochem. 14, 361–362 (1966)
Freiman, D. G.: Use of an organic chelating agent in the histochemical study of alkaline phosphatase activation. Proc. Soc. exp. Biol. (N.Y.) 84, 338–341 (1953)
Ganote, C. E., Rosenthal, A. S., Moses, H. L., Tice, L. W.: Lead and phosphate as sources of artifact in nucleoside phosphatase histochemistry. J. Histochem. Cytochem. 17, 641–650 (1969)
Gillis, J. M., Page, S. G.: Localization of ATPase activity in striated muscle and probable sources of artifact. J. Cell Sci. 2, 113–118 (1967)
Goldfischer, S., Essner, E., Novikoff, A. B.: The localization of phosphatase activities at the level of ultrastructure. J. Histochem. Cytochem. 12, 72–95 (1964)
Goodlad, G. A. J., Mills, G. T.: The acid phosphatases of rat liver. Biochem. J. 66, 346–354 (1957)
Habeeb, A. F. S. A., Hiramoto, R.: Reaction of proteins with glutaraldehyde. Arch. Biochem. Biophys. 126, 16–26 (1968)
Holt, S. J.: Factors governing the validity of staining methods for enzymes and their bearing upon the Gomori acid phosphatase technique. Exp. Cell Res., Suppl. 7, 1–32 (1959)
Hopwood, D.: Some aspects of fixation with glutaraldehyde. A biochemical and histochemical comparison of the effects of formaldehyde and glutaraldehyde and glutaraldehyde fixation on various enzymes and glycogen, with a note on penetration of glutaraldehyde into liver. J. Anat. (Lond.) 101, 83–92 (1967)
Hopwood, D.: Fixatives and fixation: A review. Histochemical J. 1, 323–360 (1969)
Hopwood, D., Allen, C. R., McCabe, M.: The reactions between glutaraldehyde and various proteins. An investigation of their kinetics. Histochemical J. 2, 137–150 (1970)
Howell, D. S.: Current concepts of calcification. J. Bone Jt Surg. A 53, 250–258 (1971)
Hugon, J. S., Maestracci, D., Ménard, D.: Glucose-6-phosphatase activity in the intestinal epithelium of the mouse. J. Histochem. Cytochem. 19, 515–525 (1971)
Hugon, J. S., Maestracci, D., Ménard, D.: Stimulation of glucose-6-phosphatase in the mucosal cells of the mouse intestine. J. Histochem. Cytochem. 21, 426–440 (1973)
Janigan, D. T.: Tissue enzyme fixation studies. I. The effects of aldehyde fixation on β-glucuronidase, β-galactosidase, N-acetyl-β-glucosamidase and β-glucosidase in tissue blocks. Lab. Invest. 13, 1038–1050 (1964)
Janigan, D. T.: The effects of aldehyde fixation on acid phosphatase activity in tissue blocks. J. Histochem. Cytochem. 13, 473–483 (1965)
Jones, G. R. N.: Quantitative histochemical studies on the succinate-neotetrazolium reductase system. Losses of nitrogenous material occuring from unfixed frozen sections in rat liver during incubation. Exp. Cell Res. 43, 268–280 (1966)
Kanamura, S.: Difference in resistance to glutaraldehyde or formaldehyde fixation between mouse and rat glucose-6-phosphatase. Acta histochem. cytochem. 3, 160–162 (1970)
Kanamura, S.: Demonstration of glucose-6-phosphatase activity in hepatocytes following transparenchymal perfusion fixation with glutaraldehyde. J. Histochem. Cytochem. 19, 386–387 (1971)
Kanamura, S.: Optimal postfixation washing time for ultrastructural demonstration of glucose-6-phosphatase activity. J. Histochem. Cytochem. 21, 1086–1089 (1973)
Leskes, A., Siekevitz, P., Palade, G. E.: Differentiation of endoplasmic reticulum in hepatocytes. I. Glucose-6-phosphatase distribution in situ. J. Cell Biol. 49, 264–287 (1971)
Lewis, J. A., Tata, J. R.: Heterogeneous distribution of glucose-6-phosphatase in rat liver microsomal fractions as shown by adaptation of a cytochemical technique. Biochem. J. 134, 69–78 (1973)
Lineweaver, H., Burk, D.: The determination of enzyme dissociation constants. J. Amer. chem. Soc. 56, 658–669 (1934)
Lowry, O. H., Rosebrough, J. J., Farr, A. L., Randall, R. J.: Protein measurement with the Folin phenol reagent. J. biol. Chem. 193, 265–275 (1951)
Marchesi, V. T., Palade, G. E.: The localization of Mg−Na−K activated adenosine triphosphatase on red cell ghost membranes. J. Cell Biol. 35, 385–404 (1967)
Matt, Ch. A. von, Fuenfschilling, H., Moppert, J. M., Gander, E. S.: Comparative determination of liver acid phosphatase activity in decapited and perfused rats. Histochemie 25, 72–76 (1971)
Moses, H. L., Rosenthal, A. S.: Pitfalls in the use of lead ion for histochemical localization of nucleoside phosphatases. J. Histochem. Cytochem. 16, 530–539 (1968)
Moses, H. L., Rosenthal, A. S., Beaver, D. L., Schuffman, S. S.: Lead ion and phosphatase histochemistry. II. Effect of adenosine triphosphate hydrolysis by lead ion on the histochemical localization of adenosine triphosphatase activity. J. Histochem. Cytochem. 14, 702–710 (1966)
Nielsen, A. E.: Kinetics of precipitation. Oxford: Pergamon Press 1964
Nordlie, R. C.: Glucose-6-phosphatase, hydrolytic and synthetic activities. The enzymes, vol. IV, 12, p. 543–610. Edit.: P. Boyer. New York: Academic Press 1971
Nordlie, R. C., Arion, W. J.: Liver microsomal glucose-6-phosphatase, inorganic pyrophosphatase and pyrophosphate—glucose phosphatransferase. III. Associated nucleoside triphosphate and nucleoside diphosphate—glucose phosphotransferase activities. J. biol. Chem. 240, 2155–2164 (1965)
Nordstrom, C., Dahlquist, A., Josefsson, L.: Quantitative determination of enzymes in different parts of the villi and crypts of rat small intestine. Comparison of alkaline phosphatase, disaccharidases and dipeptidases. J. Histochem. Cytochem. 15, 713–721 (1967)
Novikoff, A. B., Hausmann, D. H., Podber, E.: The localization of adenosine triphosphatase in liver: in situ staining and cell fractionation studies. J. Histochem. Cytochem. 6, 61–79 (1958)
Ockerman, P. A.: Glucose-6-phosphatase in human jejunal mucosa; lack of activity in glycogenois of Cori's type I. Clin. chim. Acta 9, 151–156 (1964)
Ockerman, P. A., Lundborg, H.: Conversion of fructose to glucose by human jejunum: absence of galactose-to-glucose conversion. Biochim. biophys. Acta (Amst.) 195, 34–37 (1956)
Poelman, R. E., Daems, W. T.: Problems associated with the demonstration by lead methods of adenosine triphosphatase activity in resident peritoneal macrophages and exudate monocates of the guinea-pig. J. Histochem. Cytochem. 21, 488–498 (1973)
Pratt, O. E.: Some factors affecting rat brain phosphatase activity in fresh tissue suspensions and in histochemical methods. Biochim. biophys. Acta (Amst.) 14, 380–389 (1954)
Rosenthal, A. S., Moses, H. L., Beaver, D. L., Mchuffman, S. S.: Lead ion and phosphatase histochemistry. I. Nonenzymatic hydrolysis of nucleoside phosphates by lead ion. J. Histochem. Cytochem. 14, 698–701 (1966)
Rosenthal, A. S., Moses, H. L., Tice, L. W., Ganote, C. E.: Lead ion and phosphatase histochemistry. III. The effects of lead and adenosine triphosphate concentration on the incorporation of phosphate into fixed tissue. J. Histochem. Cytochem. 17, 608–612 (1969)
Sabatini, D. D., Bensch, K., Barrnett, R. J.: Cytochemistry and electron microscopy. The preservation of cellular ultrastructure and enzymatic activity by aldehyde fixation. J. Cell Biol. 17, 19–58 (1963)
Täljedal, I. B.: Kinetics of glucose-6-phosphatase in pancreatic islets as revealed by staining histochemistry. Histochemie 19, 355–362 (1969)
Tice, L. W.: Lead-adenosine triphosphate complexes in adenosine triphosphatase histochemistry. J. Histochem. Cytochem. 17, 85–94 (1969)
Tice, L. W., Barrnett, R. J.: The fine structural localization of glucose-6-phosphatase in rat liver. J. Histochem. Cytochem. 10, 754–762 (1962)
Tormey, J.: Significance of the histochemical demonstration of ATPase in epithelia noted for active transport. Nature (Lond.) 210, 820–822 (1966)
Tunik, B. D.: Cytochemical localization of myofibrillar adenosine triphosphatase activity in sarcomeres of glycerinated muscle by the calcium precipitation method. J. Histochem. Cytochem. 19, 75–89 (1971)
Wachstein, M., Meisel, E.: On the histochemical demonstration of glucose-6-phosphatase. J. Histochem. Cytochem. 4, 592 (1956)
Walton, A. C.: The formation and properties of precipitation. New York: Interscience Publishers and John Wiley & Sons 1967
Wang, J. H., Tu, J.: Modification of glycogen phosphorylase b by glutaraldehyde. Preparation and isolation of enzyme derivatives with enhanced stability. Biochemistry (Wash.) 8, 4403–4410 (1969)
Wang, J. H., Tu, J.: Allosteric properties of glutaraldehyde—modified glycogen phosphorylase b. J. biol. Chem. 245, 176–182 (1970)
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This work was supported by a grant from the Medical Research Council of Canada (J.H.).
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Berteloot, A., Hugon, J.S. Effect of glutaraldehyde and lead on the activity of hepatic glucose-6-phosphatase. Histochemistry 43, 197–214 (1975). https://doi.org/10.1007/BF00499701
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DOI: https://doi.org/10.1007/BF00499701