A quantitative study of N-acetyl-beta-glucosaminidase activity in unfixed tissue sections of the guinea-pig thyroid gland.
A post-coupling procedure for the quantitative measurement of N-acetyl-beta-glucosaminidase activity in unfixed tissue sections of guinea-pig thyroid is described. The method depends on the cleaving of a naphthol AS-BI substrate and the insoluble reaction product is post-coupled with Fast Garnet GBC salt (in acetate buffer, pH 6.2) at 4 degrees C. Even though this enzyme is localized in the lysosomes, an inert colloid stabiliser, polyvinyl alcohol (G18/140) is included in the reaction medium to allow the use of the optimal substrate concentration (0.5 mg/ml) whilst employing a low concentration (5%) of ethylene glycol monomethyl ether. The high molecular weight (90 000) grade of polyvinyl alcohol used did not stabilize the lysosomal membrane, although a lower molecular weight (15 000) grade of polyvinyl alcohol (G04/140) may do. The enzyme activity was not affected by the metal ions Ca2+ and Zn2+ and was totally abolished by the specific inhibitor 2-acetamido-2-deoxy-D-gluconolactone.
D000118:Acetylglucosaminidase / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D005260:Female; D005404:Fixatives; D006168:Guinea Pigs; D006596:Hexosaminidases / Q000378:metabolism*; D006651:Histocytochemistry; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology; D011142:Polyvinyl Alcohol / Q000494:pharmacology; D013961:Thyroid Gland / Q000201:enzymology*; D015032:Zinc / Q000494:pharmacology
D005404:Fixatives; D011142:Polyvinyl Alcohol; D006596:Hexosaminidases; D000118:Acetylglucosaminidase; D015032:Zinc; D002118:Calcium