scholarly journals Improvement in the Histochemical Localization of Leucine Aminopeptidase with a New Substrate, L-Leucyl-4-Methoxy-2-Naphthylamide

1960 ◽  
Vol 7 (2) ◽  
pp. 261-264 ◽  
Author(s):  
Marvin M. Nachlas ◽  
Benito Monis ◽  
David Rosenblatt ◽  
Arnold M. Seligman
Keyword(s):  
Leucine Aminopeptidase ◽  
Hydrolysis Product ◽  
Histochemical Demonstration ◽  
New Method ◽  
Lipid Solubility ◽  
Rapid Rate ◽  
Frozen Sections ◽  
Enzyme Localization ◽  
Copper Chelate ◽  
Fresh Frozen

A new method for the histochemical demonstration of leucine aminopeptidase in fresh frozen sections was developed with the substrate L-leucyl-4-methoxy-2-naphthylamide. The superior enzyme localization is due to the more rapid rate of coupling of the hydrolysis product, 4-methoxy-2-naphthylamine as compared to 2-naphthylamine itself, and to the low lipid solubility and high substantivity for protein of the copper chelate of the dye formed on coupling with tetrazotized diorthoanisidine. A comparison of the old and the new method is illustrated, and a description is given of the localization of leucine aminopeptidase in the tissues of the rat and man.

scholarly journals Freeze substituted tissue in 5'-nucleotidase histochemistry. Comparative histochemical and biochemical investigations.

1979 ◽  
Vol 27 (12) ◽  
pp. 1582-1587 ◽  
Author(s):  
K Klaushofer ◽  
H von Mayersbach
Keyword(s):  
Enzyme Activity ◽  
Sensitivity And Specificity ◽  
Rat Liver ◽  
Freeze Substitution ◽  
High Sensitivity ◽  
Histochemical Demonstration ◽  
Frozen Sections ◽  
Enzyme Localization ◽  
Fresh Frozen ◽  
Preparation Techniques

The suitability of freeze-substitution in n-butanol and paraffin embedding of tissues for the histochemical demonstration of 5'-nucleotidase was investigated and compared with commonly used preparation techniques, such as fresh frozen sections and cryostate sections of cold formalin and glutaraldehyde-fixed rat liver. The influences of each step of the preparation techniques on the enzyme activity were controlled by a quantitative radiochemical assay. Freeze substitution was revealed to be superior to the commonly used preparation techniques with respect to: 1) high sensitivity and specificity of the histochemical 5'-nucleotidase reaction (this is based on the fact that incubation media with very low lead concentrations (0,6 mM/1) can be used); 2) excellent morphological appearance of the tissues showing cytological details of enzyme localization; 3) unlimited storage of the tissue materials and ease of sectioning.

scholarly journals A NEW HISTOCHEMICAL METHOD FOR ACID PHOSPHATASE BY THE USE OF 5-IODOINDOXYL PHOSPHATE

1966 ◽  
Vol 14 (2) ◽  
pp. 171-176 ◽  
Author(s):  
G. W. EVANS ◽  
CECILIA L. WHINNEY ◽  
K. C. TSOU
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Azo Dye ◽  
Redox System ◽  
Histochemical Demonstration ◽  
Crystal Formation ◽  
Frozen Sections ◽  
Fresh Frozen ◽  
Ph Range

5-Iodoindoxyl phosphate has been found to be a useful indigogenic substrate in the histochemical demonstration of acid phosphatase activity. Its superiority to other indoxyl phosphates is apparently due to a rapid oxidation of 5-iodoindoxyl to 5,5'-diiodoindigo in the acid pH range. A redox system of ferri-ferrocyanide enhances the oxidation and improves the localization. This method can be applied to calcium-formol-fixed tissues or to fresh frozen sections, although fixed tissues yield better results. The method is not recommended for the demonstration of enzyme activity in lipid-rich tissues because of the complexing property of lipids with 5,5'-diiodoindigo that results in crystal formation. The distribution of acid phosphatase activity with this method is generally similar to that obtained using azo dye methods.

scholarly journals p-HYDROQUINONE AND p-BENZOQUINONE AS HISTOCHEMICAL REAGENTS I. A NEW TETRAZOLIUM METHOD FOR AMINO GROUPS, AND THE MECHANISM OF FORMAZAN STAINING OF PARAFFIN AND FRESH FROZEN SECTIONS

1967 ◽  
Vol 15 (7) ◽  
pp. 404-408 ◽  
Author(s):  
G. G. CARMICHAEL ◽  
STEPHANIE T. K. MANDER
Keyword(s):  
Electron Acceptor ◽  
Thermal Shrinkage ◽  
Frozen Sections ◽  
Amino Groups ◽  
Paraffin Sections ◽  
Tetrazolium Bromide ◽  
Acid Ph ◽  
Reduction System ◽  
Oxidation Reduction ◽  
Fresh Frozen

The staining of amino groups by formazan when dehydrated paraffin sections are incubated in a mixture of hydroquinone and 3-(4,5-dimethyl thiazolyl-2)-2 ,5-diphenyl-2H-tetrazolium bromide at an acid pH is reported. The mechanism of this reaction and of the cytoplasmic deposition of formazan in fresh frozen sections incubated under similar conditions is investigated. It is shown that the oxidation of hydroquinone to semiquinone is responsible for the reaction, the tetrazole acting as electron acceptor. The tissue amino groups, exposed by dehydration and thermal shrinkage, and the nitrogen groupings of phosphobipid behave as "catalysts." The relevant properties of the hydroquinone-benzoquinone oxidation-reduction system are described, and the reactions between benzoquinone and tissue constituents are reviewed.

Effect of bacterial endotoxemia on succinic dehydrogenase of liver and kidney of rabbit

1961 ◽  
Vol 201 (1) ◽  
pp. 16-18 ◽  
Author(s):  
J. Cascarano ◽  
A. D. Rubin ◽  
A. K. Neumann ◽  
B. W. Zweifach
Keyword(s):  
Succinic Dehydrogenase ◽  
Significant Inhibition ◽  
Frozen Sections ◽  
Experimental Animals ◽  
E Coli ◽  
Fresh Frozen ◽  
Tissue Homogenates ◽  
Liver And Kidney ◽  
Lethal Doses

The in vivo inhibition of liver and kidney succinic dehydrogenase by administration of lethal doses of bacterial endotoxin ( Escherichia coli and Salmonella typhosa) was investigated. Quantitative determinations conducted on tissue homogenates revealed significant inhibition of activity only in liver of rabbits injected with E. coli lipopolysaccharide. The histochemical distribution of succinic dehydrogenase in fresh frozen sections of kidney was the same in both control and experimental animals. However, the centrolobular areas of liver appeared considerably depressed in activity in both E. coli and S. typhosa endotoxin-treated animals. These data, along with those presented by other studies in the literature, suggest that the action of endotoxin appears to be restricted to certain cells.

Lipoproteins and Fibrinogen in Atherogenesis

1979 ◽  
Author(s):  
K.W. Walton
Keyword(s):  
Low Density Lipoproteins ◽  
Atherosclerotic Plaques ◽  
Low Density ◽  
Frozen Sections ◽  
Fibrin Monomer ◽  
Fresh Frozen ◽  
Arterial Intima

Previous work from this and other laboratories has shown that material antigenically related to the low-density lipoproteins (LDL) and to fibrinogen is demonstrable in atherosclerotic plaques. When arterial intima is extracted electrophoretically, ‘bound’ and ‘labile’ fractions of these antigens are in each case demonstrable. In the case of the “bound’ fraction of fibrinogen-related antigen (FRA) it has not been clear as to whether the material is in the form of native fibrinogen, as fibrin monomer, or as fibrin. To examine this problem fresh-frozen sections of arteries containing FRA have been examined by immunohistological techniques using antisera specific for fibrinogen, fibrinopeptides, plasmin and fibronectin. The results obtained with these antisera will be compared with one another and with results obtained with antisera to the antigens of LDL.

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