bluuebunny/crossref_metadata_2025 huggingface dataset:
A METHOD FOR THE HISTOCHEMICAL DEMONSTRATION OF DISACCHARIDASE ACTIVITIES: APPLICATION TO INVERTASE AND TREHALASE IN SOME ANIMAL TISSUES
Authors: DAHLQVIST ARNE , BRUN ARNE
Published: April 2011

1) A histochemical method for the demonstration and localization of disaccharidase activity using various disaccharides as the substrate is described. Frozen sections are incubated in a solution containing substrate plus a coupled oxidation-reduction system of glucose oxidase, phenazine methosulfate and a tetrazolium salt. Where disccharidase activity is present, the substrate is hydrolyzed with the liberation of glucose, and a deep blue insoluble formazan is formed.

2) This method has been used for the demonstration of invertase and trehalase in rat tissue sections. The small intestine, colon, stomach and kidney were investigated.

3) All the epithelial cells of the small intestinal mucosa, in the villi, crypts and glands of Brunner, contain invertase and trehalase. These enzymes appear to be localized to small rounded granules which are scattered throughout the cytoplasm of the cells. The activity is especially high in the perinuclear part of the epithelial cells, but no concentration of the granules is to be seen in "brush border" region.

4) The stomach and the kidney also contain weak invertase activity which is present in granules localized to small distinct cell groups. No trehalase activity could be demonstrated in these organs. The colon contains essentially no invertase or trehalase.
NEW OBSERVATIONS ON DISCREPANCIES IN THE HISTOCHEMICAL LOCALIZATION OF LEUCINE AMINOPEPTIDASE
Authors: NACHLAS MARVIN M. , FRIEDMAN MELVIN M. , SELIGMAN ARNOLD M.
Published: April 2011

Differences in the localization of leucine aminopeptidase in the cortex of the rat kidney have been reported with two methods employing the same substrate. Test tube and tissue experiments were carried out in an effort to explain the disparity. Substrate concentration, pH, buffer, the use of an activator, duration of incubation and temperature of incubation were found to have insignificant effects. Four important findings were (1) leucine aminopeptidase activity of the inner cortex of the rat kidney assayed two to three times greater than that of the outer cortex, (2) this difference was confirmed by histochemical demonstration with both methods, (3) fast blue B inhibited the enzyme more than fast garnet GBC and fast Corinth V, and (4) fast blue B coupled faster than the other diazonium salts.
POLYVINYL ALCOHOL AS AN EMBEDDING MEDIUM FOR LIPID AND ENZYME HISTOCHEMISTRY
Authors: FEDER NED
Published: April 2011

Polyvinyl alcohol is a highly polar, water-soluble resin. Fixed, washed specimens are soaked in a mixture of 15% polyvinyl alcohol, 15% glycerol and 70% water, which is then allowed to dry at room temperature to a hard, sectionable block. Sections 1-100 µ thick are floated on water, dried on glass slides and stained. There is good preservation of tissue and cell structure, and of many lipids and enzymes.
ON THE BINDING OF CONGO RED BY AMYLOID
Authors: PUCHTLER HOLDE , SWEAT FAYE , LEVINE M.
Published: April 2011

Congo red in alkaline 80% ethyl alcohol solution saturated with NaCl stains amyloid selectively. The distribution of the dye is very similar to that observed in serial sections stained with Highman's Congo red method.

In sections stained with Congo red in aqueous or alcoholic solutions without differentiation all tissue structures are intensely colored, but only amyloid is dichroic. The rapid removal of dye by alkaline solutions from structures other than amyloid suggests ionic or salt-type bonds between Congo red and these structures. In contrast, the binding of Congo red by amyloid from alkaline alcoholic solutions and the increase in intensity of staining upon addition of NaCl indicate a non-ionic type linkage between amyloid and dye.

The effects of deamination, acetylation and various oxidation procedures on the binding of Congo red indicate interaction of the dye with hydroxyl groups of amyloid.

According to data on the direct dyeing of cellulose in textile dyeing, Congo red is adsorbed by hydrogen bonding between hydroxyl groups of the polysaccharide chains and the amino groups of the dye. It seems probable that a similar mechanism is involved in the binding of Congo red by amyloid.

It is suggested that the selective staining of amyloid with Congo red is due to steric configurations of amyloid or one of its components which favor non-ionic binding of the eye.
STUDIES IN THE HISTOCHEMISTRY OF PLASMALOGENS: I. THE EFFECT OF FORMALIN AND ACROLEIN FIXATION ON THE PLASMALOGENS OF ADRENAL AND BRAIN
Authors: NORTON WILLIAM T. , GELFAND MARTIN , BROTZ MIRIAM
Published: April 2011

Formalin containing fixatives were found to cause a rapid drop in extractable plasmalogens of steer adrenal cortex and medulla and rat brain to 50-60% of the initial value in 4-6 hours. This rapid destructive reaction was followed by a slow decrease so that for several days the plasmalogen levels remained relatively constant. An acrolein containing fixative, although not evaluated with adrenal, showed similar behavior with rat brain. Control experiments show the fixative aldehyde is solely responsible for this plasmalogen loss. This decrease is not paralleled by a similar decrease in extractable total lipid or total phospholipid. It is believed that this decrease represents an actual destruction of the enol-ether bond by chemical reaction with the fixative aldehyde. Although the nature of this reaction is unknown, model experiments show cephalin plasmalogen to be much more labile than choline plasmalogens to formaldehyde, although all the cephalin plasmalogens are not destroyed during the fast reaction in tissue.

These experiments also show that steer adrenal cortex and medulla have the same plasmalogen content and that the plasmalogen in either part of the gland responds the same to fixation, even though plasmal staining would indicate that the medulla is richer in plasmalogen. After a period of fixation sufficient to render the plasmal reaction negative in both adrenal cortex and medulla and in rat brain there is still at least 35-50% of the initial plasmalogen present. The intensity of the plasmal stain appears to parallel the concentration of the fixative labile plasmalogen. In these particular tissues there was no suggestion of an increase in free aldehyde over plasmalogen until the tenth day of fixation.
A HISTOCHEMICAL ANALYSIS OF COENZYME-LINKED DEHYDROGENASES IN SCORBUTIC SEMINAL VESICLES
Authors: CAVAZOS L. F. , MANNING J. P. , JEFFREY J. E. , FEAGANS W. M.
Published: April 2011

The effects of ascorbic acid depletion and inanition on the seminal vesicle of the guinea pig were studied histochemically. Through use of the appropriate substrate, several enzymes of the Embden-Meyerhof scheme, the citric acid cycle and the hexose monophosphate shunt were examined. Results indicate that in avitaminosis C, and to a lesser extent in inanition, there is alteration of the functional activity of the cytoplasm of the seminal vesicle epithelium. This change is indicated by decrease of the hexose monophosphate shunt enzymes and, to a limited degree, changes in the citric acid cycle and glutamate dehydrogenase side chain.
THE CYTOCHEMICAL DEMONSTRATION OF SUCCINIC DEHYDROGENASE IN MOUSE LEUKOCYTES
Authors: MORRISON JOHN H. , KRONHEIM SUZANNE
Published: April 2011

The specificity of Nitro BT as an electron acceptor for the intracellular localizations of succinic dehydrogenase in mammalian blood cells was investigated. Imprints of mouse spleen were fixed either by immersion in acetone or formol calcium under varying conditions of time and temperature or by freezing in cold isopentane. The incubation mixture used for the detection of enzyme activity contained succinate, Nitro BT, amytal, and phosphate buffer (pH 7.4).

By comparison of the intensity and site of formazan production in imprints prepared under varying conditions of fixation, it was found that specific intracellular localization of succinic dehydrogenase occurred only in cells fixed in formol calcium prior to, or after incubation with Nitro BT. Using this fixation method, succinic dehydrogenase was detected in the hemocytoblast, lymphocyte, myelocyte, and megakaryocyte. In all these cell types the enzyme was confined exclusively to the mitochondria. Only slight activity was observed in occasional metamyelocytes and none in any of the mature segmented forms.

Imprint cells fixed in formol calcium failed to exhibit endogenous enzyme activity when succinate was omitted from the incubation medium (control series). It was also shown that reduction of Nitro BT was inhibited completely in the presence of 6 mM malonic acid (inhibition series) and that this inhibition was reversible in the presence of excess succinate (reversal series).

The results of this study are discussed in terms of their possible significance in the differentiation process leading to formation of the mature granulocyte. On the basis of our findings and other information in the literature concerning modifications in fine structure of the mitochondria during blood cell maturation, it was suggested that cytogenesis of the granulocyte is associated with a progressive loss in oxidative enzyme activity and that oxidation reactions in the mature granulocyte may be mediated, for the most part, by glycolysis.
AN IMPROVED APPARATUS FOR QUANTITATIVE CYTOCHEMICAL MODEL STUDIES AND ITS USE IN AN EXPERIMENTAL TEST OF THE TWO-WAVELENGTH METHOD
Authors: VAN DUIJN P. , DEN TONKELAAR E. M. , HARDONK M. J.
Published: April 2011

The construction of an improved apparatus for quantitative studies on cytochemically-stained membranes is described. The membrane-holder of this apparatus was constructed in such a way that the area of the film whose extinction is measured coincides exactly with the area punched out for weight determination and chemical analyses. In addition, provisions were made to assure an uniform response of the photomultiplier to light rays traversing different parts of the field. Using this system, an experimental test of the so called two-wavelength method was carried out.

The spectra of Feulgen-stained deoxyribonucleic acid cellulose films or periodic acid Schiff-stained cellulose films were measured. The stained membranes were punched out, after which the resulting discs were cut, folded, and brought back to their original position in the holder between two unstained cellulose membranes. In this way the number of chromophores in the measured area was kept constant although their macroscopic distribution had changed considerably.

From the spectra of the folded stained discs the extinction of the same material in unfolded form could be calculated by applying the two-wavelength method of Patau and Ornstein. The two-wavelength method was found to correct well, but some residual error remained even in this macrosystem, due to the relatively strong effect of small measuring errors on the end results.
ENZYMORPHOLOGY OF RAT BRAIN; β-GLUCURONIDASE, ALKALINE PHOSPHATASE, ESTERASE
Authors: FISHMAN JOEL S. , HAYASHI MASANDO
Published: April 2011

The enzymorphology technique with chloral hydrate fixative as applied to rat brain gives good preservation of the cell morphology and enzyme activity. The findings are in harmony with those obtained by others using separate techniques. Thus, β-glucuronidase shows an intense activity in the Purkinje cells of the cerebellum; α-naphthyl esterase and indoxyl esterase show strong activity in the neurons, astrocytes and their processes and alkaline phosphatase enriched the walls of the capillaries.
THE REACTIVITY OF PROTEINS OF SOME CONNECTIVE TISSUES AND EPITHELIAL STRUCTURES WITH 2,4-DINITROFLUOROBENZENE
Authors: ZERLOTTI EUGENIO , ENGEL MILTON B.
Published: April 2011

2,4-Dinitrofluorobenzene was used without any coupling procedure as a histochemical reagent to localize some reactive groups of proteins in connective tissues and epithelial structures. Tissues were fixed by freezing-drying to avoid uncontrolled alterations of proteins and the reactions were modified in some sections by postfixation or pre-treatment with a variety of reagents, including alcohol, formalin, formaldehyde vapors, acetic anhydride, nitrous acid and N-ethylmaleimide. The staining and resolution were greatly enhanced by viewing the sections with monochromatic light at 410 mµ. When used in this way, 2,4-dinitrofluorobenzene is specific for: (1) α-amino groups of terminal amino acids; (2)ε-amino groups of lysine and hydroxylysine; (3) sulfhydryl groups of cysteine. However, the reaction seems to be dominated by the ε-amino groups of lysine in most of the tissues. In the native state not all functional groups of tissue colloids react presumably because the groups are masked by the interractions involved in forming tertiary structures or covalent bonds. This is specially notable in calcified tissues. Pyknotic nuclei, pre-dentin, carious dentin and newly formed bone are more reactive suggesting a less highly organized state of their structural proteins or some other modification in the lysine and hydroxylysine containing proteins. 2,4-Dinitrofluorobenzene is valuable as a specific cytochemical reagent and, in some instances, as an indicator of the state of organization of cellular and extracellular colloids.
DISTRIBUTION OF PROTEIN-BOUND SULFHYDRYL GROUPS IN YEAST CELLS
Authors: POMERANZ Y.
Published: April 2011

Young and old cultures of 12 yeasts have been stained by the Barrnett-Seligman histochemical procedure for localization of sulfhydryl groups. Thiol groups have been found distributed in the cytoplasm, but not in the cell wall. The intense staining of sulfhydryl groups by the specific stain was clearly showns in the slender thread of protoplasm between bud and mother cell.

The intensity of staining in the proliferating cell was higher than in the resting cell; variations in thiol content between various yeast species were recorded. No decrease in thiol content of yeast spores, as compared with vegetative cells, has been found in Schizosaccharomyces octosporus, whereas the staining of yeast spores in Saccharomyces and Hansenula was found to be negligible. Implications of these findings are discussed.
HISTOCHEMICAL STUDY ON GLYCOGEN SYNTHESIS FROM GLUCOSE-1-PHOSPHATE IN BLOOD CELLS UNDER NORMAL AND PATHOLOGICAL CONDITIONS
Authors: TAKEUCHI TADAO , TADOKORO NOZOMU , IDE HIDEMARO
Published: April 2011

Polysaccharides of glycogen class were histochemically synthesized from glucose-1-phosphate at pH 5.7 in various blood and bone marrow cells except the crythrocytic series. The intense activity of phosphorylase was indicated blue with iodine which stained a long straight chain of 1,4-linkages, while the branching enzyme (amylo-1,4 → 1,6-transglucosidase) activity was indicated red violet by staining shorter chains of a branched polysaccharide. These reactions appeared most frequently in neutrophilic, pseudoeosinophilic or amphophil series, particularly in the mature cells of the series. The histochemical synthesis of glycogen from glucose-1-phosphate in neutrophil leucocytes in the peripheral blood varies under pathologic conditions.
THIAMINE DIPHOSPHATASE: A HISTOCHEMICAL STUDY
Authors: NAIDOO D.
Published: April 2011

The sites of action in the central nervous system of the rat of thiamine diphosphatase, an enzyme liberating phosphate from Vitamin B1, is described and illustrated. The optic tectum of the domestic fowl has also been examined. Some histological and cytochemical properties of the enzyme in these two species is compared. The activity of this enzyme may be involved in the pathological disturbances and tissue damage occurring in thiamine deficiency.
AN AUTORADIOGRAPHIC STUDY OF THE LOCALIZATION AND DISTRIBUTION OF TRITIATED HISTIDINE IN BONE
Authors: TONNA E. A. , CRONKITE E. P. , PAVELEC M.
Published: April 2011

The localization, distribution and cellular turnover of tritium labeled-histidine was followed autoradiographically in the femora of young mice. Initially the labeled amino acid was taken up by the cellular phase of the skeletal system within the first hour after isotope administration. From 1 to 4 hours, intracellular histidine appeared within newly deposited cartilage and bone matrix. By 14 days skeletal cells had lost their label, however, red blood cells were still labeled at 14 days.
TWO-EMULSION RADIOAUTOGRAPHY
Authors: BASERGA RENATO , NEMEROFF KEN
Published: April 2011

Two-emulsion radioautography can distinguish between beta particles emitted from C14 atoms and those originating in H3 atoms. It uses two layers of sensitized emulsion separated by an inert layer of celloidin: the C14 beta particles, because of their longer range, are recorded in the second emulsion, but the beta particles from tritium are arrested by the first emulsion. Experiments have shown that the combination of two NTB emulsions separated by a celloidin layer is the most satisfactory. The total thickness of the radioautograph is 19 µ. The first emulsion plus the celloidin layer measure together 12 µ in thickness and decrease by 38% the flux of C14 beta particles reaching the second emulsion. Staining of the specimen with Mayer's hematoxylin and eosin and celloidin coating follow the exposure and processing of the first emulsion. The second emulsion is applied after the celloidin coating and processed after a suitable second exposure.

By using a tritiated precursor of DNA and a C14-labeled precursor of RNA or proteins, or viceversa, two-emulsion radioautography can be applied to investigate two distinct metabolic processes occurring at the same time in the same cell.
HISTOCHEMICALLY SELECTIVE ACIDOPHILIA OF BASIC NUCLEOPROTEINS IN CHROMATIN AND NUCLEOLI AT ALKALINE pH
Authors: SPICER S. S.
Published: April 2011

Chromatin and nucleoli of tissues fixed with formalin-free, mercurial or alcoholic solutions, have the unique property of staining metachromatically with the acid dye Biebrich scarlet in the restricted pH range 9.0-10.0. The nuclear structures lack affinity for fast green FCF under these conditions so that a mixture of Biebrich scarlet and fast green stains chromatin and nucleoli a characteristic orange-red at pH 9.5. A Feulgen-Biebrich scarlet sequence distinguishes magenta DNA in chromatin and orange-red basic protein in nucleoli. Although nuclei in tissues fixed with the buffered mercurial show strong affinity for the basic dye alcian blue, nuclei in formalin-fixed tissue do not.

In formalin-fixed, trichloroacetic acid (TCA)-extracted tissues, chromatin stains with Biebrich scarlet or fast green but nucleoli do not. The acidophilia induced by TCA may in part depend on the fact that deoxyribonucleic acid (DNA) is removed by this treatment. However such tissues lose the acidophilia of the chromatin when treated again with formaldehyde but regain the acidophilia after re-extraction with TCA, indicating removal of formaldehyde condensates from amines by TCA.

The affinity of chromatin and nucleoli for Biebrich scarlet at pH 9.5 is highly susceptible to destruction by relatively brief nitrosation, acetylation or exposure to formaldehyde. Extraction with a sulfuric acid-mercurial reagent also eliminates this staining; but other solvents used for isolation of histone do not, except that 1 N HCl has such an effect when employed on sections of tissue fixed with HgCl2.

After mild treatment with TCA, exposure to pH 9.5 buffer extracts DNA whereas buffer at pH 6.0 has no such effect.
THE ACTION OF FORMALDEHYDE SOLUTIONS ON HUMAN BRAIN LIPIDS
Authors: HESLINGA F. J. M. , DEIERKAUF F. A.
Published: April 2011

The composition of the lipids extracted from fresh normal human brain and from brains preserved up to 24 years in unbuffered formaldehyde solutions was analysed quantitatively using column and paper chromatography, followed by various determinations.

The amounts of cholesterol, cerebrosides, sulphatides, phosphoinositides and sphingomyelin remain unaffected.

Lecithin, phosphatidylethanolamine and phosphatidylserine are broken down to the corresponding lyso compounds, fatty acids, phosphatidic acid and probably lysophosphatidic acid. The lyso compounds are further broken down by the liberation of the second fatty acid. By this process the fatty acid content is markedly increased. The total lipid content decreases slightly by the formation of water soluble phosphoryl compounds. Consequently the phosphorus content of the total lipid extract suffers marked diminution.

Histochemical reactions on lipids in formaldehyde stored tissues should preferably be performed in combination with an analysis of the lipid extract.
THE MECHANISM OF TISSUE STAINING BY FERRIC HYDROXYQUINOLINE METHODS FOR β-GLUCURONIDASE
Authors: JANIGAN D. T. , PEARSE A. G. E.
Published: April 2011

1. The investigations reported in this paper show that the staining product of the Fishman-Baker and other ferric hydroxyquinoline methods for the demonstration of β-glucuronidase is entirely uninfluenced by the presence or absence of the enzyme.

2. Staining is due to the uptake of ferric iron from the complex equilibrium mixture of the incubating medium.

3. The uptake of iron is influenced by a number of factors but is essentially dependent on the nature of the tissue.

4. Changes in the localization of iron staining which occur in tissues in physiological or pathological states are likely to be due to significant though ill-understood differences in affinity for iron shown by intracellular constituents.
THE FINE STRUCTURAL LOCALIZATION OF GLUCOSE-6-PHOSPHATASE IN RAT LIVER
Authors: TICE LOIS W. , BARRNETT RUSSELL J.
Published: April 2011

Glucose-6-phosphatase activity was demonstrated histochemically in rat liver using either the Wachstein-Meisel medium or a modified Chiquoine medium, and the characteristic properties of the enzyme activity were confirmed. The distribution of activity in both unfixed and hydroxyadipaldehyde-fixed material was demonstrated with the electron microscope. Activity was found in both smooth- and rough-surfaced elements of the endoplasmic reticulum of hepatic cells, including the nuclear envelope, but was absent from the plasma membrane. These findings further implicate the endoplasmic reticulum as an organelle of transport, and in addition suggest that the nuclear envelope has functional as well as morphological continuity with the endoplasmic reticulum.
Dialogical Action: Moving From Community-Based to Community-Driven Participatory Research
Authors: Montoya Michael J. , Kent Erin E.
Published: April 2011

Proponents of community-based research advocate for the active involvement and engagement of community members, citing improved construct validity, intervention efficacy, and accountability. However, to create the conditions in which expertise is mutually constructed and in which no one is the object of research, a reconsideration of the fundamental ethos of community involvement and engagement is required. In this article, we seek to accomplish two goals: (a) to briefly assess the definitions of community health, focus groups, and dissemination that are often used in community-based research; and (b) to introduce an application of dialogical action that goes beyond traditional focus group methodology to promote the creation of an evolving and dynamic dialogue among campus and community stakeholders. An urban case study is presented.
Discourses of Depression of Australian General Practitioners Working With Gay Men
Authors: Körner Henrike , Newman Christy E. , Mao Limin , Kidd Michael R. , Saltman Deborah C. , Kippax Susan
Published: April 2011

The data for this article are from a primary health care project on HIV and depression, in which the prevalence, nature, clinical management, and self-management of depression among homosexually active men attending high-HIV-caseload general practice clinics were investigated. One of the qualitative arms consisted of in-depth interviews with general practitioners (GPs) with high caseloads of gay men. The approach to discourse analysis was informed by Halliday’s systemic functional linguistics. GPs constructed three discourses of depression: engaging with psychiatric discourse, engaging with the patient’s world, and engaging with social structures. When GPs drew on the discourse of psychiatry, this discourse was positioned as only one possible construction of depression. This discourse was also contextualized in the social lives of gay men, and it was explicitly challenged and rejected. Engaging with their patients’ social world was considered vital for recognizing depression in gay men. Finally, the GPs’ construction of depression was inextricably linked to social disadvantage and marginalization. Depression is highly heterogeneous and constructed in terms of social relationships rather than as an independent entity that resides in the individual. There is a synergy between GPs’ constructions of depression and men’s experiences of depression, which differs from conventional medical views, and which enables GPs to be highly effective in dealing with the mental health issues of their gay patients.
A Burger Model for the Effective Behavior of a Microcracked Viscoelastic Solid
Authors: Nguyen Sy Tuan , Dormieux Luc , Pape Yann Le , Sanahuja Julien
Published: April 2011

This article aims at the determination of the effective behavior of a microcracked linear viscoelastic solid. Due to the nonlinearity of the strain concentration in the cracks, the latter cannot be derived directly from a combination of the correspondence theorem with the Eshelby-based homogenization schemes. The proposed alternative approach is based on the linear relationship between the macroscopic strain and the local displacement discontinuity across the crack. An approximation of the effective behavior in the framework of a Burger model is derived analytically.
Desmoplastic Small Cell Tumor With Bizarre Giant Nuclei
Authors: Bono Francesca , Testi Maria Adele , Rosai Juan
Published: April 2011

A case is reported of an intra-abdominal desmoplastic small cell tumor featuring giant bizarre nuclei, an event not previously recorded in this entity to the best of the authors’ knowledge. The diagnosis was confirmed by immunohistochemistry and molecular genetics. A list is provided of the many other conditions in which this morphologically spectacular but clinically inconsequential nuclear change has been recorded.
Mitochondrial Mutations
Authors: Lax Nichola Z. , Turnbull Doug M. , Reeve Amy K.
Published: April 2011

Mutations in mitochondrial DNA cause a number of neurological diseases with defined neuropathology; however, mutations in this genome have also been found to be important in a number of more common neurodegenerative diseases. In this review, the authors discuss the importance of mitochondrial DNA mutations in a number of different diseases and speculate how such mutations could lead to cell loss. Increasing our understanding of how mitochondrial DNA mutations affect mitochondrial metabolism and subsequently result in neurodegenerative disease will prove vital to the development of targeted therapies and treatments.
Effect of a thin soft core on the bending behavior of a sandwich with thick CFRP facings
Authors: Iaccarino P. , Leone C. , Durante M. , Caprino G. , Lamboglia A.
Published: April 2011

Three-point bending tests in the elastic domain were carried out on sandwich specimens with the facings made of unidirectional T800H/3900-2 laminates, and the core of a soft and thin viscoelastic layer, varying the slenderness ratio. A closed-form formula available in the literature, developed for sandwich beams with thick facings, was used to predict the rigidities in flexure, adopting three different methods for the calculation of the shear rigidity. The best results were obtained by the classical sandwich theory, which provided reasonably good correlation with the experimental results, slightly overestimating the actual values. A finite element model consisting of three layers, made of standard CQUAD4 and CHEXA8 elements for facings and core, respectively, was developed, accurately predicting beam bending behavior. From the numerical results, the inability of classical sandwich theory to correctly estimate the bending rigidity is correlated with the through-the-thickness shear deformation of facings, not accounted for in the analysis.
MICRODETERMINATION OF CYTOCHROME OXIDASE IN RAT TISSUES BY THE OXIDATION OF N-PHENYL-p-PHENYLENEDIAMINE OR ASCORBIC ACID
Authors: PEARL W. , CASCARANO J. , ZWEIFACH B. W.
Published: April 2011

The cytochrome oxidase content of rat tissues was investigated, utilizing the accumulation of the free radical formed by the emzymatic, univalent oxidation of a stable, non-toxic substrate, N-phenyl- p-phenylenediamine ( p-aminodiphenylamine). This procedure eliminated the non-enzymatic coupling of the radical with α-naphthol, found in the classic Nadi reaction. Using homogenates, the following decreasing order of enzyme activity was found: heart, kidney, diaphragm, liver, rectus abdominis. A more sensitive assay was also developed, involving the reduction of cytochrome c by ascorbic acid, and using 2-( p-iodophenyl)-3-( p-nitrophenyl)-5-phenyl tetrazolium chloride to analyze unoxidized ascorbic acid. The same order of tissue activity was demonstrated. The cytochrome oxidase content of a tissue may be a reflection of its physiological role.
ARE ACTIVE AND INACTIVE PHOSPHORYLASES HISTOCHEMICALLY DISTINGUISHABLE?
Authors: GODLEWSKI HENRY G.
Published: April 2011

Some features of the phosphorylase activity derived from the biochemical datas are listed. It is considered worthwhile to assess the results of the histochemical reaction. Possibilities to demonstrate the phosphorylase a, phosphorylase b, and the total phosphorylase activity by the modifications of the original Takeuchi and Kuriaki method are successively discussed. In order to distinguish between the activities of two forms of enzyme it is necessary to block the mutual and spontaneous conversions of the active and inactive phosphorylase which may occur in the course of incubation. The intensity of the reactions performed in those conditions should then be compared with the intensity of the Guha and Wegmann' or Grillo's reactions for the total phosphorylase activity. Applying NaF, and EDTA as the blocking agents, and ATP, MgSO4, AMP as the activators, makes it possible to assess the content of two forms of phosphorylase in the tissue section examined.
ELECTRON MICROSCOPIC STUDY ON SPECIFIC PROTECTION OF ISOLATED BORDETELLA BRONCHISEPTICA ANTIBODY DURING EXHAUSTIVE LABELLING WITH URANIUM
Authors: STERNBERGER LUDWIG A. , DONATI EDWARD J. , WILSON CATHERINE E.
Published: April 2011

Exposure of antibody to relatively high concentrations of uranium yielded an electron-dense specific localizing reagent. Destruction of antibody activity during labelling was prevented by protection of the antibody-antigen combining sites: Anti- Bordetella bronchiseptica antibody was removed from serum by absorption with B. bronchiseptica. The washed agglutinate was exposed to uranium. The antibody recovered from this agglutinate by brief alkali treatment in the cold was 80 to 100% pure and contained 28 to 312 atoms of uranium per unit of 156,000 molecular weight. The material stained the walls of living or formalin killed, and the cytoplasm of living B. bronchiseptica. E. coli and mouse spleen cells were not stained. Contrast was diminished when B. bronchiseptica was exposed to unlabelled anti- B. bronchiseptica antiserum prior to uranium-antibody conjugate. Absorption with B. bronchiseptica, but not with E. coli, abolished the staining capacity of purified uraniumn-antibody conjugate. Whole antiserum labelled with uranium without specific protection, even though retaining some antibody activity, was not suitable as a stain since it deposited a large amount of debris and apparently stained both E. coli and B. bronchiseptica.
OBSERVATIONS ON THE SUBCELLULAR LOCALIZATION OF OXIDATIVE ENZYMES WITH NITRO BLUE TETRAZOLIUM
Authors: HITZEMAN JEAN WALTER
Published: April 2011

The effectiveness of nitro blue tetrazolium in localizing enzymes at the subcellular level has been investigated. The primary source of interference with enzyme localization was lipid material which progressively accumulated formazan deposits during incubation in a tetrazole medium containing pyridine nucleotide. Acetone fixation at 2°C extracted this lipid and permitted the visualization of both mitochondrial and microsomal enzymes. There was no evidence for an unspecific adsorption of Nitro-BT on lipoprotein membranes which led to an antifactual staining pattern. The demonstration of oxidative enzyme activity in the alkaline pH range was not impeded by "nothing dehydrogenase" if fats had been removed. Otherwise, lipid-tetrazole interactions contributed heavily to nonenzymatic formazan deposition which was difficult to distinguish visually from mitochondrial activity. MTT-Co++ was also adsorbed by lipids. After cold acetone fixation, patterns of this formazan were not equivalent to those found with Nitro-BT. The relation of these findings to the current evaluations of enzyme activity at the subcellular level is discussed. At the level of the light microscope, nitro blue tetrazolium is an adequate indicator of the intracellular sites of oxidative enzyme activity if lipid interference is counteracted.
THE DISTRIBUTION OF CHOLINESTERASES IN THE SENSORY GANGLIA OF MAN AND OF SOME MAMMALS
Authors: CAUNA N. , NAIK N. T.
Published: April 2011

The distribution of cholinesterases in the cranial and spinal sensory ganglia was investigated in the cat, guinea-pig, rat, mole and man using a modified Koelle's histochemical technique.

It was found that the cytoplasm of the sensory nerve cells contained acetylcholinesterase in all species, but the concentration of the enzyme varied from cell to cell. Nerve fibers gave a negative acetylcholinesterase reaction in the cat and guinea-pig, and a weak to moderate positive reaction in the rat, mole and man.

Non-specific cholinesterase was only found in the sensory ganglia of the cat and guinea-pig. In human sensory ganglia, it was found during the fetal life but not after birth.

The sensory ganglia of the rat were found to be nearer to those of man than the ganglia of the cat or guinea-pig in respect of cholinesterase distribution.

It is suggested that the variations in acetyl-cholinesterase activity among different nerve cells may signify the existence of several cell types in the sensory ganglia.

The role of intracytoplasmic cholinesterase in the sensory nerve cells is discussed.
INTRACELLULAR MITOCHONDRIAL VARIATION IN ENZYME ACTIVITY AS SHOWN BY HISTOCHEMICAL STUDIES USING LIGHT AND ELECTRON MICROSCOPY
Authors: AVERS CHARLOTTE J. , TKAL MARGARET M.
Published: April 2011

Activities of reduced diphosphopyridine nucleotide (DPNH)-tetrazolium reductase, succinic dehydrogenase, and reduced triphosphopyridine nucleotide-tetrazolium reductase were studied using the substrates glutamate, iso-citrate, and DPNH for the first enzyme; succinate; and 6-phosphogluconate, glucose-1-phosphate, and TPNH for the last enzyme system. These activities were quantitated in light microscopical studies by counting the formazan particles in non-vacuolated epidermal meristematic cells of timothy grass seedling roots. Using Nitro-blue tetrazolium as the H+-acceptor, the average counts per cell were: 57, DPNH-tetrazolium reductase; 65.5, succinic dehydrogenase; 78.2, TPNH-tetrazolium reductase. These significantly different counts were higher than those found in an earlier study using neotetrazolium, but the three relative levels of activity were unchanged. One discrepancy was that 6-phosphogluconate and succinate gave the same activity levels although 6-phosphogluconate is believed to be oxidized via TPN-linked systems. INT also was tested, but the results suggested non-specific enzyme activity and those data were discarded. Electron microscopical studies were reported in which DPNH-tetrazolium reductase activity was shown to be restricted to the mitochondrial membranes in about half the mitochondrial population of individual cells. Since preliminary counts of mitochondrial profiles from electron micrographs indicated about 1000 mitochondria per cell, the absolute numbers of formazan particles using light microscopy probably is an unreliable measure. Also, clumping of mitochondria after histochemical incubation was demonstrated in some of the electron microgaphs. This suggested that lower brightfield counts might result since one or more mitochondria may have been associated as a single formazan deposit. Generally, the data verified the earlier conclusions that (a) mitochondria were the specific site of this oxidative enzyme activity, and (b) that only a fraction of the mitochondrial population of a cell was enzymatically active for this enzyme.
THE APPLICABILITY OF A NEW SCHIFF BASE, GLYOXAL BIS (2-HYDROXYANIL),FOR THE CYTOCHEMICAL LOCALIZATION OF IONIC CALCIUM
Authors: KASHIWA HERBERT K. , ATKINSON WILLIAM B.
Published: April 2011

A simple pocedure for localizing calcium ions in tissues and in cells with glyoxal bis (2-hydroxy-anil) (GBHA) has been described. The GBHA molecule, in alcoholic solution made alkaline with NaOH, chelated with Ca, Ba, Sr, Cd, Cu, Co, and Ni ions, forming colored precipitates. All the colored GBHA complexes, with the exception of the red Ca-GBHA granules, were decolorized when immersed in a solution containing carbonate and cyanide. Of the methods available for demonstrating calcium ions, the sensitivity of the GBHA method was comparable only to the Scott-Packer electron microscope method; the latter, however, localized calcium plus magnesium while the GBHA method was specific for calcium ions.

In tissues that were fixed in 10% neutral formalin, alcohol-formalin or absolute alcohol, false localization due to binding of extraneous or diffused calcium ions to polyanionic sites was evident. The false localization of calcium ions was prevented by processing the tissues by the freeze-dry or the freeze-substitution method, dehydrating the tissue below the cutectic point of calcium chloride. The GBHA method was not able to demonstrate calcium ions present as insoluble salts, such as calcium carbonate or as apatite crystals, giving false negative results.
CYTOCHEMICAL STUDIES OF RABBIT AORTIC CELLS IN TISSUE CULTURE
Authors: KOIDE REIZO , POLLAK O. J.
Published: April 2011

The enzymatic activities of acid and alkaline phosphatase, adenosine triphosphatase, carboxyl esterase and succinic dehydrogenase as well as the lipid, cholesterol and acid mucopolysaccharide content in three types of cells cultured from normal and cholesterol-fed rabbit aortae were studied.

Although enzymatic activities varied from cell to cell in the same culture, there was good evidence that "endothelial" type III cells had more activity than the other two cell types. With normal aging enzymatic activity seemed to diminish in all three cell types. Giant cells were usually more active. Degenerated cells also had somewhat increased enzymatic activity. However, type I and II cells always had considerably less activity than type III cells, regardless of their origin, age or shape.

Type III cells exhibited intense activity in acid phosphatase; moderate activity in esterase and in succinic dehydrogenase; and weak or almost negligible activity in alkaline phosphatase and adenosine triphosphatase. In "fibroblastic," type I or II cells all these enzymatic activities were weak or negligible. No differences of enzymatic activity were seen between cells from normal and atherosclerotic aorta but for the esterase activity which was highest in type III cells from plaques.

With the exception of carboxyl esterase activity which seemed to be slightly higher in the type III cells from atherosclerotic aorta, cells of different origin did not differ in enzymatic activity. Some type I or II cells changed their shape and became round (9), but these round cells did not exhibit stronger enzymatic activities than the spindle forms. On the other hand, large size or differently shaped type III cells retained their greater activity. Thus, greater activity of type III cells was not due to shrinking of the cytoplasm and subsequent condensation of the cell. Studies of other enzyme systems are under way. Very preliminary results indicate that "endothelial" cells yield considerably higher (G. Nadi) cytochrome-c oxidase than other cells.

"Endothelial" or type III cells contained more lipid and acid mucopolysaccharide than "intermediate" or type I cells and "fibroblastic" or type II cells. "Endothelial" our type III cells from atherosclerotic plaques contained cholesterol in their cytoplasm while none of the cells from normal aorta did.

Type III cells are metabolically the most active cells cultured form the aortic wall. Their activity may play a major role in the development of the atherosclerotic plaques. Morphologic and functional characteristics of these cells led to the suggestion that they may be endothelial cells or, possibly, macrophages. The fact that only type III cells from atherosclerotic aorta contain much cholesterol and that they appear in larger numbers than cells from normal aorta is held supportive evidence that they are endothelial cells.

Results with cells in tissue cultures were compared with reports on histochemical findings in aortae.
HISTOCHEMICAL DEMONSTRATION OF LIPIDS AND LIPASE ACTIVITY IN RAT TESTIS
Authors: GEORGE J. C. , AMBADKAR P. M.
Published: April 2011

Lipase activity has been demonstrated by improved techniques in the interstitial tissue as well as in the seminiferous elements of the rat testis. The distribution of lipids in the testis has been studied histochemically. It is seen that both the distribution of the enzyme as well as the substrate follow a definite parallel pattern in four different phases. The physiological significance of this enzyme (which is present in quite large concentrations in the interstitium as well as germinal elements) in the metabolism of the testis and the sperms, is thus revealed.
RELIABILITY OF THE RADIOAUTOGRAPHIC TECHNIQUE FOR THE DETECTION OF NEWLY SYNTHESIZED PROTEIN
Authors: DROZ B. , WARSHAWSKY H.
Published: April 2011

It is widely believed that the radioactivity detected by radioautography after administration of a labelled amino acid is due to newly-synthesized protein. The validity of this postulate was examined in work done on the organs of mice injected with leucine-C14 and sacrificed one-half or 24 hours later. Pancreas, liver, kidney and cerebellum were removed and divided into 3 aliquots to determinse (a) the total radioactivity, (b) the biochemical distribution of the radioactivity in the fresh organs, (c) the losses of radioactivity occurring after Bouin fixation and in the course of histological processing as well as the biochemical distribution of the radioactivity retained in histological sections. In addition, an attempt was made to extract radioactivity from mounted sections by various reagents.

It was found that 10-50% of the total radioactivity is lost in Bouin's fluid (presumably as free leucine) and in ethanol and dioxane (presumably as lipid-linked metabolites).

As for the radioactivity persisting in the processed organs, only insignificant amounts were found in the nucleic acid fraction, while most of it was associated with protein.

The possibility that the radioactive amino acids are bound to protein by ionic linkage was examined by extracting sections with strong acids and bases, but only insignificant amounts were thus lost.

It is concluded that at least 91-97% of the radioactivity retained in histological sections after labelled amino acid injection is firmly bound to protein (presumably by peptide bonds). Thus, radioautography under these conditions is a reliable method to detect and trace newly-synthesized protein.
OSMIUM TETROXIDE FIXATION OF LIPIDS: NATURE OF THE REACTION PRODUCTS
Authors: RIEMERSMA JELLE C.
Published: April 2011

To study the possible reaction mechanisms involved in osmium tetroxide fixation as used in electron microscopy, the products were analyzed which result from the interaction of osmium tetroxide and a number of pure substances. By means of paper chromatography, differences could be observed between the reaction products of 1-decene and oleic acid on the one side and of lecithin on the other. In the case of 1-decene and oleic acid, an intermediate osmic acid ester, as postulated by Criegee's reaction scheme, appears to be the main reaction product. In the case of lecithin, the reaction product is mainly a corresponding dihydroxy compound; here the intermediate osmic ester is readily split with formation of osmium dioxide and dihydroxy lecithin.

The presence of a phosphoryl choline group as in lecithin thus appears to influence the course of the reaction with osmium tetroxide. If a polar group as well as double bonds play a part in the reactions, this might explains electron microscopic findings suggesting that during osmium fixation the deposition of osmium dioxide probably occurs at the polar ends of the lipid molecules. Our chromatographic experiments show that osmium dioxide, as a colloidally dispersed hydrate, is negatively charged and capable of Coulomb interaction with the quaternary ammonium group present in lecithin. Combined with the finding that osmium dioxide is readily split off from the osmium ester derivative of lecithin which is formed initially, this evidence strongly favors the hypothesis that osmium dioxide is deposited in the polar strata of lipid micelles.
THE OXIDATION OF LACTATE AND α-GLYCEROPHOSPHATE BY RED AND WHITE SKELETAL MUSCLE: II. HISTOCHEMICAL STUDIES
Authors: VAN WIJHE M. , BLANCHAER M. C. , JACYK W. R.
Published: April 2011

The use of phenazine methosulfate (PMS) in the histochemical demonstration of lactate and α-glycerophosphate dehydrogenases in skeletal muscle has been examined. When PMS is added to the conventional dehydrogenase reagents, these enzymes appear to be at least as active in the white fibers as in the red. In the absence of PMS, the red fibers appear to be much richer than the white in lactate and α-glycerophosphate dehydrogenase. The conventional methods without PMS apparently give a false impression of the distribution of diphosphopyridine nucleotide (DPN)-linked dehydrogenases in skeletal muscle because of their dependence on the indigenous tissue level and intracellular localization of reduced DPN (DPNH)-diaphorase.

A leakage of both enzymes from the sections into the histochemical dehydrogenase reagents occurred both in the presence and absence of PMS. When allowances are made for such losses, it appears that the PMS results are compatible with other evidence that the glycolytic dehydrogenases are more active in white than in red skeletal muscle fibers and that these enzymes are largely confined to the cytoplasm.
HISTOCHEMICAL STUDIES ON THE DISTRIBUTION OF ADENOSINE TRIPHOSPHATASE IN THE TRIGEMINAL GANGLION CELLS OF THE RAT
Authors: TEWARI H. B. , BOURNE G. H.
Published: April 2011

A detailed histochemical study of the distribution of adenosine triphosphatase among trigeminal ganglion cells, associated nerves and epi-, peri-, and endoneurium has been carried out. The latter as well as the pericellular zones show the highest degree of the enzymatic activity. The intracellular activity, although feeble in most of the cases as compared to the pericellular zones, is revealed as morphological entities—as filamentous or vesicular bodies or solid clumps. These are sometimes arranged in the perinuclear zone, at other times as concentric rings in the middle part of the cytoplasm. They may be concentrated at the peripheral part or distributed as isolated bodies throughout the cytoplasm. It has been suggested that these bodies, distributed in various topographical fashions, represent the enzymatically active site of the endoplasmic reticulum. Further, a coordination of various stages seen among different cells, suggests that there is a shifting of the enzymatic activity from the perinuclear zone to the peripheral part of the cytoplasm. At the latter site and in the adjacent pericellular areas there is a strong possibility of an enzymatic role in the membrane permeability. The enzyme may also be playing such a role in epi-, peri- and endoneurium.