The histochemical demonstration of choline acetyltransferase by semipermeable membrane technique

1976 ◽  
Vol 49 (4) ◽  
pp. 303-307 ◽  
Author(s):  
Gerald H�ther ◽  
Hans Luppa
Keyword(s):  
Choline Acetyltransferase ◽  
Histochemical Demonstration ◽  
Semipermeable Membrane ◽  
Membrane Technique

scholarly journals Enhanced resolution of histochemical distribution of glucose-6-phosphate dehydrogenase activity in rat neural tissue by use of a semipermeable membrane.

1991 ◽  
Vol 39 (7) ◽  
pp. 937-943 ◽  
Author(s):  
M A Philbert ◽  
C M Beiswanger ◽  
T L Roscoe ◽  
D K Waters ◽  
H E Lowndes
Keyword(s):  
Vestibular Nucleus ◽  
Neural Tissue ◽  
Semipermeable Membrane ◽  
G6pd Activity ◽  
Ependymal Cells ◽  
Regional Localization ◽  
Enhanced Resolution ◽  
Membrane Technique ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
The Brain

We examined the histochemical distribution of glucose-6-phosphate dehydrogenase (G6PD) activity in neural tissue using different diffusion barriers. Although polyvinyl alcohol and agar overlays permitted regional localization of G6PD, a semipermeable membrane revealed cellular differences in G6PD activity within populations of neurons. Distribution of G6PD activity in selected regions of the nervous system was examined using the membrane technique. White matter usually exhibited strong G6PD activity. The neuronal somata of the dorsal root ganglia (L4-L6) and anterior horns of the spinal lumbar enlargement demonstrated a variation in activity which was independent of somal size. Satellite cells showed intense activity when the membrane technique was used. Hippocampal pyramidal and granular cells of the dentate gyrus exhibited moderate, uniform G6PD activity, but only weak activity was seen in hippocampal and dentate molecular layers. High levels of activity were observed in the vascular endothelial cells of the brain, spinal cord, and choroid plexus, and in the ependymal cells of the spinal central canal and ventricles of the brain. The superior vestibular nucleus appeared to have little G6PD activity in either the neuron cell bodies or the surrounding parenchyma. The use of a semipermeable membrane for localization of G6PD activity in neural tissues permits enhanced resolution of neuron elements and may provide a more accurate assessment of G6PD activity in histological preparations.

scholarly journals Quantitative histochemical study of acid phosphatase activity in rat liver using a semipermeable membrane technique.

1987 ◽  
Vol 35 (2) ◽  
pp. 175-180 ◽  
Author(s):  
W M Frederiks ◽  
F Marx ◽  
G N Jonges ◽  
C J Van Noorden
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Rat Liver ◽  
Acid Phosphatase Activity ◽  
Enzyme Inhibitors ◽  
Semipermeable Membrane ◽  
Coupling Technique ◽  
Lysosomal Acid ◽  
Membrane Technique ◽  
Post Coupling

Acid phosphatase activity has been demonstrated in rat liver with the semipermeable membrane technique using naphthol AS-BI phosphate as substrate and hexazotized pararosaniline (HPRA) as simultaneous coupling agent. With this method the final reaction product (FRP) appeared in rat liver as intensely colored red granules in liver parenchymal cells and in Küpffer cells. The absorbance spectrum of the FRP peaks between 510 and 550 nm. A nonspecific reaction product, as has been found in skeletal muscle, did not occur in rat liver. A substrate concentration of 5 mM and a HPRA concentration of 10 mM result in optimum localization and activity. We concluded from the results with different enzyme inhibitors that lysosomal acid phosphatase was demonstrated. The mean absorbance of the FRP increased linearly with incubation time (15-60 min). Furthermore, we found a linear increase of the FRP with increasing section thickness (4-10 micron). When the simultaneous coupling method was replaced by a post-coupling technique, the colored reaction product was diffusely located throughout the cytoplasm. In conclusion, the simultaneous coupling technique in combination with the semipermeable membrane method is a valuable tool for detecting and quantifying lysosomal acid phosphatase activity in rat liver. We demonstrated that acid phosphatase activity is 1.2 times higher periportally than pericentrally in rat liver, and that 24 hr fasting before the experiments did not change the acid phosphatase activity.

scholarly journals A histochemical procedure for light microscopic demonstration of xanthine oxidase activity in unfixed cryostat sections using cerium ions and a semipermeable membrane technique.

1993 ◽  
Vol 41 (5) ◽  
pp. 667-670 ◽  
Author(s):  
W M Frederiks ◽  
F Marx
Keyword(s):  
Xanthine Oxidase ◽  
Oxidase Activity ◽  
Semipermeable Membrane ◽  
Myeloperoxidase Activity ◽  
Final Reaction Product ◽  
Final Reaction ◽  
Xanthine Oxidase Activity ◽  
Membrane Technique ◽  
Cryostat Sections ◽  
Unfixed Cryostat

Xanthine oxidoreductase exists in two functionally distinct forms. Under normal conditions, the larger part of the enzyme occurs as an NAD(+)-dependent dehydrogenase form which produces NADH and urate. The dehydrogenase can be transformed under various (patho)physiological conditions to an oxygen-dependent oxidase form which produces oxygen radicals and/or hydrogen peroxide and urate. Tetrazolium salts are used to demonstrate the total activity of both the dehydrogenase and the oxidase form of the enzyme. We have developed a procedure to detect the oxidase form only in unfixed cryostat sections with the use of cerium on the basis of the semipermeable membrane technique. The incubation medium contained hypoxanthine as substrate, cerium ions, and sodium azide to inhibit catalase and peroxidase activity. In a second-step reaction, diaminobenzidine was polymerized in the presence of cobalt ions by decomposition of cerium perhydroxide. Large amounts of final reaction product were found in milk droplets in the acini of lactating bovine mammary gland, whereas milk-secreting epithelial cells contained hardly any final reaction product. In rat duodenum, enzyme activity was found in the cytoplasm of enterocytes and goblet cells but not in the mucus. Control reactions performed in the absence of substrate or in the presence of substrate and allopurinol, a specific inhibitor of xanthine oxidase, were completely negative in both tissues, with the exception of polymorphonuclear leukocytes in the lamina propria of duodenum. The positive nonspecific reaction in these cells was caused by myeloperoxidase activity. We conclude that the present method is specific for the detection of xanthine oxidase activity. Moreover, conversion of the dehydrogenase form into the oxidase form can be prevented by omission of chemical fixation of the tissue in the present procedure.

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