Journal of Histochemistry & Cytochemistry

CHAUNCEY, HOWARD H.; KRONMAN, JOSEPH H.; LEVITT, MAYER A.

doi: 10.1177/12.9.647pmid: 14221044

Acid phosphatase, alkaline phosphatase, nonspecific esterase, pseudocholinesterase, leucine aminopeptidase, and β-d-galactosidase activities of parotid and submandibular salivary glands were evaluated in six animal species. Assays for total enzyme, lyoenzyme, and demoenzyme were performed on unfixed and chloral hydrate formalin (CHF) fixed frozen cryostat sections.Following fixation lyoenzyme (soluble fraction) activity exhibited extreme reduction for all enzymes except pseudocholinesterase, while the demoenzyme (insoluble fraction) activity of acid phosphatase, nonspecific esterase, pseudocholinesterase, and β-d-galactosidase was maintained at a high level or increased. Since removal of lyoenzyme activity and retention of desmoenzyme activity are requisites in histochemical localization, this fixative may be the preferred method for handling tissues prior to the localization of the above cited enzymes.Several hypotheses are proposed to explain the increased desmoenzyme (nonspecific esterase and β-d-galactosidase) noted subsequent to fixation.

MCCLURKIN, lOLA T.

doi: 10.1177/12.9.654pmid: 14221045

(1) In the histochemical method for adenosine triphosphatase, when magnesium and ATP are in equivalent concentrations, and sodium is present in the medium, staining occurs in the tissues. When sodium is omitted from this medium some structures—previously stained in the presence of sodium—fail to stain.(2) Ouabain inhibits staining when magnesium and ATP are equivalent and sodium is present in the medium.(3) The effects of calcium, potassium and sodium on staining with the proposed medium parallel the effects of these ions on activity of sodium-activated ATPase in vitro.(4) When magnesium ions and ATP are not equivalent in the substrate, sodium and ouabain fail to affect staining.(5) Certain tissue structures—such as the axonic nerve ending—which have been shown to contain sodium-activated ATPase in vitro, stain in the presence of sodium when magnesium and ATP are equivalent, but fail to stain when sodium is omitted from this medium.

HAYASHI, MASANDO

doi: 10.1177/12.9.659pmid: 14221046

The histochemical distribution of β-glucuronidase activity in rat tissues as demonstrated by the naphthol AS-BI glucuronide and hexazonium pararosanilin method has been described.The present method has demonstrated a close correlation between the amount of stain derived from the β-glucuronidase reaction and the reported enzyme activity by homogenate assay in every tissue examined. Tissues known to be enzyme-rich by homogenate assay showed a rapid and intense staining in a great number of cells. These tissues were the preputial gland, spleen, liver, epididymis, ovary, lymph node, thymus, kidney and thyroid. In enzyme-poor tissues, the staining was negative or limited to a small number of specific cells. These tissues were the brain, spinal cord, eye, testis, and muscle. Cells responsible for the intense staining in all tissues examined were those of the macrophage system, parenchymal cells of liver and kidney and several types of mucous membrane cells. In each cell the major staining appeared as discrete and, most often, spherical granules in the cytoplasm, and no staining was observed in the nucleus, Size, number and location of the granules was quite variable depending on the type of cells. Large granules were especially abundant in macrophages. The possible significance of this enzyme distribution is discussed briefly.

BALOGH, KÁROLY

doi: 10.1177/12.9.670pmid: 14221047

20α-Hydroxysteroid dehydrogenase activity was localized histochemically in the corpus luteum of the rat by using Nitro-BT as an indicator. Intensive enzyme activity was obseryed in the corpus luteum cells, especially during involution. The placenta and corpora lutea of pregnancy failed to reveal enzyme activity during the last week of gravidity. Other tissues, including endocrine glands, liver and kidneys were also negative. The Present method offers a possibility to identify the sites of progesterone metabolism in the rat ovary at the microscopic level.

HOPSU, V. K.; GLENNER, G. G.

doi: 10.1177/12.9.674pmid: 14223934

BRODY, IRWIN A.; ENGEL, W. KING

doi: 10.1177/12.9.687pmid: N/A

The addition of chemical agents to the standard lactate dehydrogenase (LDH) incubating medium permits the histochemical display of selective LDH isozymes. Excess lactate decreases the intensity of the fast-moving isozymes, and urea decreases the intensity of the slow-moving isozymes. Application of these modified incubating media to histologic sections allows the display of selective LDH isozymes in unhomogenized tissues. Thus, predominance of the slow-moving LDH isozymes in guinea pig gastroenemius and of the fast-moving isozymes in guinea pig soleus were found on the electrophoretic patterns of the muscle extracts and confirmed on the intact tissues.The modified incubating media were used to localize specific LDH isozymes in normal human skeletal muscle. Ry extraction of the sections with saline or acetone the intracellular sites of isozyme activity were identified with recognized cell constituents.It was concluded that the slow-moving LDH isozymes predominate in the aqueous sarcoplasm of Type I fibers while the fast-moving isozymes predominate in the aqueous sarcoplasm of Type II fibers and in the mitochondria and lipid-mitochondrial complexes. The Pattern of dark and light fibers, seen after incubation in the standard LDH medium, was found to be due to the greater activity of the slow-moving isozymes in the Type I than the Type II fibers.Thus technique of isozyme histochemistry thus allows study of the intracellular localizations and the biochemical roles of specific LDH isozymes in skeletal muscles.

DE LORES ARNAIZ, GEORGINA RODRIGUEZ

doi: 10.1177/12.9.696pmid: 14221049

HOFSTEE, B. H. J.

doi: 10.1177/12.9.700pmid: 14221050

A comparison has been made of the meaning of the experimental kinetic constants Vm, KM and Vm/KM with respect to group transfer by a hydrolase proceeding either with or without obligatory intermediary transfer of the group to the enzyme.The results have been applied to the interpretation of kinetic data in a number of actual cases involving esterases whereby either several substrates have one of these experimental constants in common or the addition of a modifier has no effect on one of these constants with respect to a particular substrate.

ROBBINS, ELLIOTT; GONATAS, NICHOLAS K.

doi: 10.1177/12.9.704pmid: 14221051

Mammalian cells of the HeLa (S3) strain, when exposed to spindle inhibitors, have been found to undergo several morphological transformations during interphase as well as during mitosis. Some of these have been studied both histochemically and ultrastructurally. The lysosomes, represented by the multivesicular bodies in HeLa cells, form clusters and become circumferentially disposed instead of occupying the polarized juxtanuclear position characteristic of these organelles. In the electron microscope it is seen that they have acquired a dense osmiophilic core that is separated from the bounding unit membrane by an electron lucent halo. The Golgi apparatus fragments under the influence of spindle inhibitors and also takes up a circumferential distribution in a pattern similar to that of the lysosomes. On the ultrastructural level, no significant modifications in this organelle are seen. Also noted in the interphase cell are numerous 60-80 Å fibrils coursing through the cytoplasm as well as a paucity of spindle our microtubules.A striking similarity has been pointed out between the behavior of the lysosomes in the drugtreated interphase cell and the untreated, normal mitotic cell.A possible explanation of some of the changes noted has been given in terms of an interruption of protoplasmic flow resulting from the disappearance of microtubules.

doi: 10.1177/12.9.711pmid: N/A

In the article by Drs. Carbonell, Castejon, and Pollak entitled "Cytochemistry of Parcoccidioides brasiliensis. I. Cytochemistry of Cytoplasmic Polysaccharides in Yeast Form Cultures with Light Microscope" appearing in the June issue of the Journal (12:413, 1964), the correct numbering of the figures in the color plate should have been in vertical rows, rather than horizontal. In the left hand column, from the top, are Figs. 1, 2, and 3. In the right hand column are Figs. 4, 5, and 6. The Journal regrets this error.