bluuebunny/crossref_metadata_2025 huggingface dataset:
THE PROPERTIES OF GOLGI-ASSOCIATED NUCLEOSIDE DIPHOSPHATASE AND THIAMINE PYROPHOSPHATASE: II. ELECTROPHORETIC SEPARATION AND IDENTIFICATION
Authors: ALLEN JOHN M.
Published: April 2011

Nucleoside diphosphatase and thiamine pyrophosphatase have been separated electrophoretically in 12.5% acrylamide gels. Identifications and characterizations of these enzymes was achieved by cytochemical means. Two basic reaction systems were used in these studies. System A contained 33 mM Tris, 15 mM calcium chloride, and 4 mM substrate material, and was adjusted to pH 9.5. System B constained 80 mM Tris-maleate buffer, pH 7.0, 3.6 mM lead nitrate, 5 mM manganous chloride, and 4 mM substrate material. Thiamine pyrophosphatase reacted most strongly with thiamine pyrophosphate, gave weak reactions with cytidine diphosphate, guanosine diphosphate, and inosine diphosphate, and did not react with uridine diphosphate and adenosine diphosphate. Nucleoside diphosphate reacted strongly with uridine diphosphate and inosine diphosphate, gave modest reactions with guanosinse diphosphate, reacted weakly with thiamine pyrophosphate and cytidine diphosphate, and was minimally reactive with adenosine diphosphate. Both enzymes were maximally activated by manganous ion and minimally activated by calcium ion. Thiamine pyrophosphatase was moderately activated by cysteine, glutathione, and potassium cyanide, was unaffected by uranyl nitrate, and was inhibited by sodium fluoride. Nucleoside diphosphatase was inhibited by cysteine, glutathione, and uranyl nitrate and was unaffected by sodium fluoride and potassium cyanide. These properties correspond very closely with those assigned to Golgi-associated nucleoside diphosphatase and thiamine pyrophosphatase in cytochemical studies. Nucleoside diphosphatase is of very general occurrence in mouse cells, whereas thiamine pyrophosphatase is of more limited distribution.
STUDIES OF THE CYTOCHEMISTRY OF NUCLEOPROTEINS III. DEMONSTRATION OF DEOXYRIBONUCLEIC-RIBONUCLEIC ACID COMPLEXES IN MAMMALIAN CELLS
Authors: LOVE ROBERT , RABOTTI GIANCARLO
Published: April 2011

1. On the basis of susceptibility to digestion with dilute nucleases, the nine types of ribonucleoprotein (RNP) stainable by the toluidine blue-molybdate (TBM) method can be divided into four groups. The first group is resistant to the action of dilute ribonuclease (RNase) but is sensitive to deoxyribonuclease (DNase) and consists of both types of chromosomal RNP, perichromosomal RNP and intranuclear amorphous parachromatin. The second group is digested by dilute RNase and by DNase, and consists of the amorphous parachromatin that diffuses into the mitotic spindle and the nucleolini. The third group is digested by dilute RNase, but not by DNase and consists of both types of diffuse cytoplasmic and the granular cytoplasmic RNP. The pars amorpha of the nucleolus constitutes the fourth group which is resistant to digestion by dilute RNase and DNase.

2. Biochemical and other studies indicate that the effects of digestion by the enzymes is not due to contamination of RNase by DNase or vice versa.

3. Staining of chromatin by TBM (method C) is usually due to dye-binding by RNA and DNA and the intensity of staining is reduced by treatment with RNase. When the effective concentration of dye is low only RNA appears to be stained; digestion with RNase then leads to an increase in the intensity of staining. It is suggested that RNA may sterically mask the phosphoryl groups of DNA. A method for selectively staining RNA in the chromatin and chromosomes is described.

4. The observations suggest that the chromosomal, perichromosomal RNP and possibly the nucleolini and intranuclear parachromatin are present as a DNA-RNA hybrid. Current knowledge of the nature and enzyme susceptibility of informational RNA and its DNA hybrid suggest that the major component of the hybrid stained by TBM is informational RNA and that this type of RNA is present in the amorphous parachromatin of the mitotic spindle.
HISTOCHEMICAL DEMONSTRATION OF HYDROLYTIC ENZYMES AND DEHYDROGENASES IN THE ORAL MUCOSA OF CATS
Authors: KAGAWA SHOHEI , OSANAI KIYOSHI
Published: April 2011

A histochemical study was made of several hydrolytic enzymes and dehydrogenases in cat oral mucosae. The activity of alkaline phosphatase, though with some difference in degree, was generally confined to the basal cell layer and superficial cell layer of the oral epithelium. Acid phosphatase was observed in the superficial layer of the oral epithelium. Esterase was demonstrated in epithelial cells, especially in the parakeratinized layer. Aminopeptidase was absent in all the oral mucosa of the eat. β-Glucuronidase activity was observed homogeneously through the epithelium. Succinic, lactic and malic dehydrogenases were generally present in the basal cell layer and adjacent prickle cells, and they were absent in the superficial layer.
QUATERNARY AMMONIUM COMPOUNDS IN CONNECTIVE TISSUE HISTOCHEMISTRY: I. SELECTIVE UNBLOCKING
Authors: KELLY JOHN W. , BLOOM GUNNAR D. , SCOTT JOHN E.
Published: April 2011

Certain features of an accepted biochemical system for fractionating biological polyanions were adapted for more precise identification of acid mucopolysaccharides in histological preparations. Tissue studies were based on two major steps: (1) complete blocking of all polyanions by a quaternary ammonium detergent and (2) selective unblocking by increasing concentrations of an electrolyte. Critical stages were visualized by suitable stains for acid mucopolysaccharides.

Results with tissues and certain polyanion models showed that both acid mucopolysaccharide and nucleic acid structures behaved histochemically, under proper conditions, as predicted from the biochemical reactions of such polyanions as hyaluronic acid, chondroitin sulfate, heparin, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). Chemical fixation involving formaldehyde seriously altered the expected positions of nucleic acid structures in the unblocking sequence; this was corrected by use of cetylpyridinium chloride in the fixing solution.

Considering the biochemical background of these methods and several different histochemical applications already accomplished, numerous possibilities are open for extending the fundamental and practical uses of quaternary ammonium compounds in microscopic studies.
EFFECT OF ALDEHYDE FIXATION ON CELLULAR RHEUMATOID FACTOR AND CERTAIN TISSUE ANTIGENS
Authors: BALL JOHN , EL DIN BAHGAT NOUR , TAYLOR GEOFFREY
Published: April 2011

Rheumatoid facor, nuclei, and γ2 globulins can be identified by immunofluorescence microscopy in alcohol-fixed paraffin-embedded sections, though wax of low melting point is required for preservation of nuclear antigens. Rheumatoid factor also resisted aldehyde fixatives; hence it should be possible to investigate the ultrastructural characteristics of cells containing it.
A QUANTITATIVE STUDY OF THE FEULGEN REACTION WITH THE AID OF HISTOCHEMICAL MODEL SYSTEMS
Authors: HARDONK M. J. , VAN DUIJN P.
Published: April 2011

This paper reports some quantitative results with three histochemical model systems. The model systems are DNA-cellulose, deoxyadenylic acid-cellulose and deoxyguanylic acid-cellulose, and DNP-bovine albumin-polyacrylamide. By measuring the UV-absorption of the nucleotide-cellulose after various hydrolysis times, the rates of splitting-off of the purines by HCl were determined. Adenine is found to be split off twice as fast as guanine.

The phosphorus/pararosaniline ratio is found to differ considerably from the theoretical value calculated on the assumption that every free aldehyde group reacts with pararosaniline.

The differences in the values found for the phosphorus/pararosaniline ratio of the DNA-cellulose and the nucleotide-cellulose can be related to the differences in the absorption spectra of these models, which points to different degrees of substitution at the amino groups of pararosaniline. Addition of protein to DNP in a polyacrylamide film does not alter the phosphorus/pararosaniline ratio.
HISTOCHEMICAL DEMONSTRATION OF MUSCLE LIPASE
Authors: BOKDAWALA F. D. , GEORGE J. C.
Published: April 2011

A method which is an advance over the existing methods for the histochemical demonstration of lipase activity in muscle, using "Tween 85' as substrate and alizarin red S as stain for the calcium soap formed, is described. Lipase has been localized in the mitochondria of the fish trunk and pigeon breast muscles.
HISTOCHEMICAL DEMONSTRATION OF CHANGES IN THE ACTIVITY OF ALKALINE AND ACID PHOSPHATASES IN THE ADRENAL OF A MIGRATORY STARLING
Authors: NAIK D. V. , GEORGE J. C.
Published: April 2011

The activity of alkaline and acid Phosphatases was studied in adrenal cortex and medulla at the beginning and the end of the period of preparation for migration in the migratory starling, Sturnus roseus. Alkaline phosphatase in cortex and acid phosphatase in medulla showed very high increases as migration approached. The increase in alkaline phosphatase activity is correlated with increase in adrenocorticotropic hormone, ribonucleic acid, corticoids and fat in adrenal cortex, and the increase in acid phosphatase in adrenal medulla with increase in adrenalin and nor-adrenalin production. The increase in acid phosphatase in the nuclei of the cortical and medullary cells is correlated with increased nucleic acid metabolism.
THE EFFECT OF MENADIONE AND PHENAZINE METHOSULFATE ON THE TETRAZOLIUM REDUCTION SYSTEM UNDER HISTOCHEMICAL CONDITIONS
Authors: HASHIMOTO T. , KALUZA J. S. , BURSTONE M. S.
Published: April 2011

The influence of phenazine methosulfate and menadione on the activity of DPNH- and TPNH-tetrazolium reductase, succinic, lactic, and triphosphopyridine nucleotide-linked isocitric dehydrogenase were studied with reference to applied histochemistry. Fresh frozen sections of various tissues were employed in conjunction with nitro blue tetrazolium as the hydrogen acceptor.

Phenazine methosulfate was found to produce a variable effect upon staining, enhancing it or suppressing it according to incubation conditions. Suppression was most pronounced with higher concentrations. Menadione had no suppressive influence and was useful in increasing tetrazolium reduction. Its usefulness is, however, limited unless a soluble factor, possibly DT-(DPNH-TPNH)-diaphorase, can be prevented from diffusing into the medium, and "nothing dehydrogenase" is selectively inactivated.
THE LOCALIZATION OF ENZYMES WITHIN SKELETAL MUSCLE FIBERS USING THE TETRAZOLIUM TECHNIQUE
Authors: SMITH BARBARA
Published: April 2011

The tetrazolium technique has been used to study the mitochondrial and glycolytic enzymes in a skeletal muscle of the rabbit. The histological picture given by mitochondrial enzymes in C fibers appeared to show a diformazan distribution pattern similar to that of mitochondria demonstrated by other techniques. The pattern of diformazan distribution given by glycolytic enzymes was similar to the sarcoplasmic reticulum of Veratti. This was considered to be a false localization as it did not coincide with the cellular distribution of these enzymes as shown by differential centrifugation. The tetrazolium technique appears to have limited value in intracellular localization except for insoluble mitochondrial enzymes.
A HISTOCHEMICAL STUDY OF SUBSTRATE SPECIFICITY FOR THE STEROID 3β-ol DEHYDROGENASE AND ISOMERASE SYSTEMS IN HUMAN OVARY AND TESTIS
Authors: GOLDBERG B. , JONES G. E. S. , BORKOWF H. I.
Published: April 2011

Either the enzyme which oxidizes 3β-ol groups in C-19 and C-21 steroids is properly designated a steroid-3β-hydroxy dehydrogenase or there is a multiplicity of such dehydrogenases.

Although DHA is a satisfactory substrate for the human ovary, adrenal, and placenta, it is not generally so for the human testis. Pregnenolone is only at times a moderately satisfactory substrate in the ovary and is not satisfactory for the testis.

In the present histochemical study two Δ4,3β-ol steroids have been reported as satisfactory substrates for human endocrine tissues as well as liver and intestine. These new substrates make it possible to demonstrate activity in human testicular tissues and at probable sites of androgen production.

If these tenets hold, they indicate that it is more satisfactory to use unphysiologic substrates for the histochemical demonstration of enzymes as this will obviate the interplay of homeostatic mechanisms which must constantly be at work to preserve the cellular equilibrium.

Although the data might suggest that the human testis, in contrast to the ovary, is deficient in isomerizing capacity and that this step precedes oxidation of the 3β-ol group, the biochemical data do not support this theory. A more probable explanation is that either DHA, its reaction product 5-androstenedione, or both, inhibit the human testicular enzyme system, while progesterone inhibits that of the corpus luteum; thus a difference between the dehydrogenases of these two human tissues appears to exist.
CHANGES IN HYDROLASE ENZYMORPHOLOGY OF RAT UTERUS AND VAGINA AFTER ESTROGENS; NAPHTHOL AS SUBSTRATES
Authors: WATANABE KEIICHI , FISHMAN WILLIAM H.
Published: April 2011

The early enzymorphologic changes of rat uterus and vagina on castration and following estrogen administration (estradiol-17β, estrone and estriol) were examined with regard to alkaline phosphatase, esterase, β-glucuronidase and acid phosphatase using naphthol AS compounds as substrates.

Castration caused a marked decrease of enzyme activities except for alkaline phosphatase of capillary endothelium and for nonspecific esterase.

All activities, compared to castrated controls, were markedly increased by estrogen administration except for the alkaline phosphatase of capillary endothelia. Estriol and estrone were more potent than estradiol-17β in the early stage of estrogen treatment (4-hour interval).

The response of alkaline phosphatase of striated border of lumenal uterine epithelium to estrogen treatment is unique in that the other hydrolases were absent from this site.

The estrogen sensitivity of the hydrolases of connective tissue cells of rat uterine stroma is clearly evident for the first time.

Of interest also is a comparison of β-glucuronidase and acid phosphatase in which naphthol AS-BI is the product of enzyme hydrolysis in each case. Although the intensity of the respective staining reactions correlated well with each other in relation to the events of castration and estrogen administration, there were clear differences evident.

Finally, the new findings demonstrate the value of the improved techniques as well as resolve some ambiguities of earlier findings obtained with diverse methods.
ANALYTICAL CYTOLOGY OF MUCIN PRODUCTION. V. A NEW MUCINOUS TISSUE CULTURE LINE
Authors: ROSAN ROBERT C. , KERRIGAN JUDITH A.
Published: April 2011

1. Morphologic and cytologic detail is presented on a hitherto unreported tissue culture line.

2. The cultures, which are derived from a biologically stabilized transplantable homologous mucinous tumor of hamster, display many of the features of the donor tumor. They elaborate mucinous materials, some of which accept the periodic acid Schiff stain and others of which take the Alcian blue stain.

3. The fluorescent periodic acid Schiff reaction identifies structures not generally considered to be Schiff positive following classical oxidation and/or blockade procedures.

4. In these cells, the nucleolus appears to have a definite envelope. It appears to complex with an acridine dye at pH 4 and gives reactions for aldehyde and ethylene groups.
A HISTOCHEMICAL ANALYSIS OF DEHYDROGENASE VARIETY IN THE DIFFERENT TYPES OF MUSCULAR TISSUES
Authors: DICULESCO I. , ONICESCO D. , MISCHIU L.
Published: April 2011

Histochemically, we examined several types of dehydrogenases in the myoblasts from "in vitro" cultures of the chick embryos, in the gastroenemius of frog, in the pigeon pectoralis, in the heart of several animal species and in the myometrium of the rabbit in different functional states (non-gestatory, gestatory and postpartum

In the myoblasts were active: succinic, isocitric, malic dehydrogenases, di- and triphosphopyridine nucleotide diaphorases, glutamic, α-glycerophosphate and lactic dehydrogenases and lactic acid, the two dehydrogenases of the pentose cycle and β-hydroxybutyric dehydrogenase.

The cardiac muscular tissues and red striated muscle show high and very high reactions for the enzymes of the Krebs cycle and the diaphorases, and negative reactions for the rest of the studied enzymes.

The white striated muscular tissue presents all the reactions, except those of the enzymes of the hexose shunt. This type of tissue is marked by a high enzyme activity of the anaerobic glycolysis.

The gestatory and non-gestatory myometrium shows all the enzymatic reactions except lactic dehydrogenase. In the myometrium of the postpartum period one notes an enzyme decrease of the Krebs cycle, of the pentose cycle and of the lipidic metabolism and an intensification of the anaerobic glycolysis and the glucose dehydrogenase.

Alcohol dehydrogenase was negative in all tissues studied with the exception of one weak reaction in the myometrium during the first postpartum hours.
HISTOCHEMICAL IDENTIFICATION OF ACID MUCOPOLYSACCHARIDES WITH ACRIDINE ORANGE
Authors: SAUNDERS ALEX M.
Published: April 2011

It was demonstrated that acridine orange, analogous to cetyl pyridinium chloride, precipitates acid mucopolysaccharides, and that the precipitates can be redissolved in solutions of graded sodium chloride concentration. The established fractional solution technique for acid mucopolysaccharide complexes with cetyl pyridinium chloride has been applied to histochemical staining by replacing the cetyl pyridinium chloride with acridine orange. The staining method described differentiates between the presence of hyaluronic acid, chondroitin sulfuric acid and heparin.
HISTOCHEMICAL STUDY OF OXIDATIVE ENZYMES IN VERTEBRATE MUSCLES
Authors: OGATA TAKURO , MORI MASAHIKO
Published: April 2011

A histochemical study was carried out on oxidative enzymes, succinic dehydrogenase, diphosphopyridine nucleotide (DPN)-dependent dehydrogenases (lactic, malic, α-glycerophosphate, glutamic, and β-hydroxybutyric dehydrogenases), DPNH-diaphorase, triphosphopyridine nucleotide (TPN)-dependent dehydrogenases (isocitrate, and glucose-6-phosphate dehydrogenase) and TPNH-diaphorase on various kinds of vertebrate muscles. The results obtained were as follows:

1. Most of the mammalian striated muscle fibers fell into three types following a histochemical demonstration of oxidative enzyme reaction: Small red muscle fibers showed higher activities of oxidative enzymes, while large white fibers lower activities and the third type of fibers, "medium fiber" which were intermediate in size showed activities between red and white muscle fibers.

2. The intrafusal fibers of mammalian muscle spindles are classified into three types by oxidative enzyme reactions, i.e., red intrafusal fibers, medium intrafusal fibers and white intrafusal fibers.

3. Regarding birds, chicken muscle was composed of three types of fibers, while pigeon and lovebird muscles displayed two types of fibers.

4. Reptile and amphibian muscle fibers were classed as small red fibers showing higher activities, medium fibers moderate activities and white fibers lower activities.

5. Fish red muscle was composed only of red fibers, and the white muscle of white fibers. The transitional part of red and white muscles, revealed three types of fibers.

6. Generally, succinic dehydrogenase, lactic dehydrogenase, malic dehydrogenase, α-glycerophosphate dehydrogenase showed a higher activity, DPNH-diaphorase, TPNH-diaphorase a moderate activity, glutamic dehydrogenase, isocitrate dehydrogenase a lower activity and β-hydroxybutyric dehydrogenase and glucose-6-phosphate dehydrogenase a trace or nonactivity in most vertebrate striated muscles.
HISTOCHEMICAL STUDY OF OXIDATIVE ENZYMES IN INVERTEBRATE MUSCLES
Authors: OGATA TAKURO , MORI MASAHIKO
Published: April 2011

A histochemical study of oxidative enzymes was carried out on various kinds of invertebrate muscles and the results obtained were as follows:

1. Insect flight muscles showed the presence of the most intense activity of succinic dehydrogenase and high activity of α-glycerophosphate and glutamic dehydrogenases.

2. Regarding crustacean striated muscles, prawn muscle was distinguished into types of fibers by oxidative enzyme reactions.

3. In molluscan muscles, the squid and octopus muscle fibers were distinguished into two types by their oxidative enzyme activities.

4. The earthworm body wall muscle was made of only one type of fibers, whose oxidative enzyme activities were low.
DIRECT FLUOROCHROMING OF RICKETTSIAE
Authors: ANDERSON DORIS , GREIFF DONALD
Published: April 2011

Several fluorochromes were used to stains directly R. mooseri in smears and tissues. The effects of fixatives, hydrogen ion concentration, time in stain and concentration of stain were studied also. Interference by background fluoresence was minimized by a preliminary staining of fixed preparations with an alcian blue, periodic acid-Schiff sequence. Maximum fluorescence of rickettsiae was obtained when tissues were fixed in Carnoy's or smears in methanol or absolute ethanol-chloroform (1:1) and stained with acridine orange.
HISTOCHEMICAL DEMONSTRATION OF NOREPINEPHRINE AT A FINE STRUCTURAL LEVEL
Authors: WOOD JOE G. , BARRNETT RUSSELL J.
Published: April 2011

The present work describes a method for the histochemical demonstration of, and the differentiation between, norepinephrine and epinephrine at a fine structural level in cells of the adrenal medulla. The two catecholamines are definitely contained in granules which occur in two different cell types; one for norepinephrine, the other for epinephrine. No evidence could be adduced from the present material that norepinephrine material served as a precursor for epinephrine granules. Preliminary observations are included on the occurence of catecholamine-containing granules in neuronal processes of the hypothalamus.
DEHYDROGENASES IN DEVELOPING BONE IN THE RAT
Authors: FULLMER HAROLD M.
Published: April 2011

Sections of mandibles from seventy-five 5-day old rats were studied histochemically for malic, isocitric, α-glycerophosphate, glutamic, lactic, 6-phosphogluconic, glucose-6-phosphate, d(—)- β-hydroxybutyric and succinic dehydrogenases and TPN- and DPN-diaphorases. All the enzymes studied (except TPN-glutamic dehydrogenase) were found in osteocytes, osteoblasts and osteoclasts. The results suggest that all bone cells possess fully functional citric acid cycles, pentose cycles, and the capacity to metabolize fatty acids and carbohydrates.

Osteoclasts generally exhibited greater enzymatic activity than either osteoblasts or osteocytes indicating a strong broad metabolic capacity. The demonstration of isocitric dehydrogenase in osteoclasts tends to discredit Walker's theory of bone resorption (18).
APPLICATION OF THE STEREOSPECIFIC INHIBITOR L-PHENYLALANINE TO THE ENZYMORPHOLOGY OF INTESTINAL ALKALINE PHOSPHATASE
Authors: WATANABE KEIICHI , FISHMAN WILLIAM H.
Published: April 2011

The intestine-specific inhibitor of alkaline phosphatase, l-phenylalanine, has been employed in the study of alkaline phosphatase in rat intestine. Five substrates were used, β-glycerophosphate, o-carboxyphenylphosphate, α-naphthyl acid phosphate, naphthol AS-BI phosphate and naphthol AS-TR phosphate. The hydrolytic rate and specific l-phenylalanine inhibition was most marked in the case of the first three substrates. When formol-Ca fixed, gum-sucrose treated sections of rat intestine were incubated in these substrates in the presence separately of 0.05 Md- and l-phenylalanine and Gomori's calcium-cobalt method was employed, a striking inhibition by the l-isomer was regularly found. Azo dye methods were applied under different conditions of time, temperature and cation concentration in some instances. The contrast was marked but not dramatic in the case of α-naphthyl acid phosphate, and it was evident but not marked in the case of naphthol AS phosphate substrates. These findings have been discussed.

The alkaline phosphatase of intestinal epithelial cells (human) grown in tissue culture exhibited great sensitivity to l-phenylalanine. Rat kidney and leukocyte alkaline phosphatase were relatively insensitive under the same conditions.

Morphologically, the l-phenylalanine-sensitive alkaline phosphatase in rat intestine is largely confined to the striated border of the epithelial cells. In animals maintained on a high fat diet, azo dye methods demonstrated the presence of fine granules in the region of the Golgi apparatus and terminal web in the presence of d-phenylalanine. These reactions were absent in companion sections incubated in l-phenylalanine-containing substrate solutions.

The possibility of comparing the morphology of a tissue in which both test and control solutions are identical except for the configuration of one component may contribute to a more precise interpretation of the results obtained in the test section.
A HISTOCHEMICAL METHOD FOR DEMONSTRATING PROTEIN-BOUND SULFHYDRYL AND DISULFIDE GROUPS WITH NITRO BLUE TETRAZOLIUM
Authors: DEGUCHI YASUO
Published: April 2011

Nitro-BT was applied to an alkaline tetrazolium method at low alkaline pH (pH 8.4) for the demonstration of sulfhydryl and disulfide groups. Evidence of specificity was given in test tube experiments and histochemical techniques. Histochemical findings with the method employed were almost similar to those of Barrnett and Seligman's method, except for a little better contrast on sections, and a fine formazan of nitro-BT gave a strict localization of sulfhydryl groups.

A comparative distribution of sulfhydryl and disulfide groups was observed in the skin of foot-sole. The detachment and destruction of sections were prevented by the present method which was advantageous in shortening the reaction time to ⅕-1/10 and in keeping a low pH in the solution.
AMINO ACID CONSTITUTION OF CHROMOSOMES
Authors: SHARMA ARUN KUMAR , CHATTERJI ASOKE KUMAR
Published: April 2011

The present experiment was designed to demonstrate the presence of two protein-bound amino acids, viz. tryptophan and tyrosine, in the different, cellular constituents with particular reference to chromosomes. The materials chosen were the root-tips of Allium cepa and Vicia faba. Studies were made from the smeared tissue preparation.

For tryptophan, indole reactions following two methods, viz., that of Gurr (1958) and of Glenner and Lillie (1957) were followed and the latter yielded satisfactory results; while for tyrosine, Bensley and Gersh's modified Millon's reaction (1953) was adopted.

The results of the reactions show that tryptophan is present in much less amount than tyrosine in the protein component of the chromosomes. The same is true with cytoplasm as well. The strongly positive tyrosine reaction of nuclear membrane, with distinct strands towards the cell membrane, has been attributed to its role in nucleocytoplasmic transfer and synthesis of cytoplasmic protein. The less positive reaction of nuclear membrane with tryptophan suggests its comparatively little importance in transfer.

Only in case of tyrosine, the differential reaction of the euchromatic and heterochromatic parts could he made out, the latter giving negative response. It has been suggested that it might appear only during the time of transfer through the nucleolus. Tryptophan on the other hand shows uniform distribution throughout the chromosome length.
THE ACTIVITY OF URIDINE DIPHOSPHATE GLUCOSE-GLYCOGEN SYNTHETASE IN SOME EMBRYONIC TISSUES
Authors: GRILLO T. ADESANYA IGE , OKUNO GIICHI , PRICE SWETLANA , FOÁ PIERO P.
Published: April 2011

Uridine diphosphate glucose (UDPG)-glycogen synthetase has been studied using chemical and histochemical methods. Activity was demonstrated in whole egg and in homogenates of whole 3-day old chick embryos, in the heart and skeletal muscle from the 5th day and in the liver from the 7th day of incubation onwards. The activity increases with age until the 16th day at which time a sharp fall was noted in all tissues studied. Enzyme activity was demonstrated also in heart, liver, skeletal muscle and brain of rat embryos from the 14th day onwards. In these tissues the activity increases with age until birth.

The appearance of UDPG-glycogen synthetase in the tissues of the chick embryo coincides with the time at which, according to previously published data, glycogen begins to appear. In the liver of the rat embryo, the enzyme appears before glycogen formation starts. This is contrary to the behavior of phosphorylase which appears later, at least in skeletal muscle. These findings are in agreement with the view that, in vivo, glycogen synthesis occurs via the UDPG, rather than the phosphorylase, pathway.
MONOAMINE OXIDASE DISTRIBUTION IN THE RABBIT EYE
Authors: SHANTHAVEERAPPA T. R. , BOURNE G. H.
Published: April 2011

Monoamine oxidase distribution in the different components of the eye has been described in detail. The degree of enzyme activity varied from one component of the eye to another, the corneal stroma and Descemet's membrane being completely negative. The corneal epithelium, choroid and nonpigmented cells of the iris stroma showed mild positive activity. Strong positive activity was present in the myelin sheaths of optic and oculomotor nerves and also in the ciliary epithelium. The irido-corneal angle (meshwork) showed a moderately strong positive activity against the surrounding tissues. The most interesting observations were made in the retina. The pigment epithelial cell layer and the outer plexiform layer showed moderate positive activity while the activity in the inner plexiform layer, the outer and inner nuclear layers and in the ganglion cell layer was negligible. The layer of nerve fibers and internal limiting membranes were completely negative. In the layer of neuroreceptors, the outer half of this layer gave a very strong positive reaction and the inner half a very mild positive reaction.

The significance of this varied distribution in the neuroreceptor layer is discussed. A comparison of distribution of some important enzymes in the myelin sheaths of peripheral nerves, pigment epithelial cells of the retina, and the inner and outer segments of the neuroreceptor layer is made to show that each tissue is differently equipped with various enzymes, due probably to a fundamental difference in their functions.
NAPHTHOL AS-BI β-d-GLUCOSIDURONIC ACID; ITS SYNTHESIS AND SUITABILITY AS A SUBSTRATE FOR β-GLUCURONIDASE
Authors: FISHMAN WILLIAM H. , NAKAJIMA YASUO , ANSTISS C. , GREEN SIDNEY
Published: April 2011

1. Naphthol AS-BI β-d-glucosiduronic acid has been prepared by first reacting potassiumnaphtholate AS-BI with acetobromomethyl glucuronate in absolute alcohol and then by removing the protecting acetyl and methyl groups with barium hydroxide and hydrochloric acid. A stable crystalline product gave the elemental and group analytical values expected for C24H22O9NBr·0.6 H2O and yielded the expected equimolar amounts of naphthol AS-BI and glucuronic acid on complete hydrolysis with purified rat liver β-glucuronidase. Contaminants were found absent after examining the product by thin layer chromatography. The glucuronide is stable at room temperature (20°C) as a solid or in aqueous solution.

2. Purified rat liver β-glucuronidase hydrolyzes the glucosiduronic acid at a rate somewhat more rapidly than it does phenolphthalein-, p-nitrophenyl- and 8-hydroxyquinoline β-d-glucosiduronic acids. The Michaelis constant is 0.000021 M, the optimum pH is 4.5. There is a linear relationship to enzyme concentration. Saccharolactone and the presence of a second substrate effectively inhibit the reaction.

3. Incidentally to the main objectives of the study, a simple colorimetric method was devised for measuring enzymically-liberated naphthol AS-BI in the kinetic studies.

4. It is concluded that naphthol AS-BI β-d-glucosiduronic acid is a suitable substrate for β-glucuronidase and that it lacks none of the desirable attributes of the classical biochemical and histochemical substrates. Moreover, it is now possible to undertake definitive studies on the design of enzymorphologic techniques for β-glucuronidase utilizing this new substrate.
THE REACTIONS OF SULFATED POLYSACCHARIDES TO SEVERAL HISTOCHEMICAL TESTS
Authors: YAMADA KAZUYORI
Published: April 2011

1. The reactions of several sulfated polysaccharides and glycogen to a series of histochemical tests are described.

2. The accepted significance of a positive periodic acid Schiff reaction appears to hold true in these substances.

3. Although the alcian blue staining of the sulfated polysaccharides is attributed primarily to their acidic groups, their appropriate hydroxyl groups can influence the stainability.

4. The azure A metachromasia and methylene blue staining of the sulfated polysaccharides confirm largely the validity of the hitherto established reaction mechanisms. A finding worthy of recording is that chromic acid oxidation abolishes metachromasia of some sulfated polysaccharides.

5. Throughout the present study discussions were made on the possible mechanisms involved in the reactions observed.
ISOLATION OF NUCLEI FROM MAMMALIAN TISSUES THROUGH THE USE OF TRITON X-100
Authors: HYMER W. C. , KUFF E. L.
Published: April 2011

It has been found that nuclei from a variety of mammlian cell types maintain their structural integrity (at the light microscope level) when exposed to the non-ionic detergent Triton X-100, in the presence of Mg or Ca ions. This observation has led to the development of a simple, rapid, and reproducible method of nuclear isolation. Nuclei isolated by this method contain more than 90% of the deoxyribonucleic acid (DNA) from filtered homogenates and exhibit diphosphopyridine nucleotide (DPN) pyrophosphorylase activity. The isolated nuclei also retain rapidly labeled ribonucleic acid (RNA) after in vivo incorporation of labeled RNA precursors. Free mitotic metaphase figures have been seen in nuclear fractions prepared from tumor cells in the Triton X-100, an observation which suggests that metaphase aggregates could be isolated in quantity with the use of this detergent and appropriate choice of tissue.
THE HISTOCHEMICAL DEMONSTRATION OF PANCREATIC-LIKE LIPASE AND COMPARISON WITH THE DISTRIBUTION OF ESTERASE
Authors: ABE MUNEAKI , KRAMER STANLEY P. , SELIGMAN ARNOLD M.
Published: April 2011

A new histochemical method for lipase was developed using naphthol-AS nonanoate as a substrate and sodium taurocholate as an activator. In a comparison with esterase, pancreatic-like lipase activity was studied in various tissues. As a result, it was disclosed that the pancreatic acinar cells show the highest activation with sodium taurocholate, and lipase activity is located in the region immediately surrounding the zymogen granules and not within the granules themselves.

Maximum activity of esterase and lipase differed among various tissues of the rat. Esterase activity is most intense in liver and kidney followed by pancreas. Pancreatic-like lipase activity is most striking in pancreas, less so in the liver and in the kidney the activity is rather weak. Because of the exceptional esterase activity of macrophages, mast cells and pericytes of the cerebral capillaries, pancreatic-like lipase activity should be ascribed to them with caution and only in terms of the difference between reaction without and with sodium taurocholate.
DIHYDROLIPOIC DEHYDROGENASE ACTIVITY: A STEP IN FORMATION OF ACYL-COENZYME A, DEMONSTRATED HISTOCHEMICALLY
Authors: BALOGH KÁROLY
Published: April 2011

A method for the histochemical localization of dihydrolipoic dehydrogenase has been developed, using diphospyridine nucleotide (DPN) as a coenzyme and Nitro-blue tetrazolium (Nitro-BT) as the final electron acceptor. With this technique the enzyme, which plays an important role in the synthesis of coenzyme-A linked acyls, was found to be widespread in the tissues of the rat, and its distribution in various organs is presented. The problems involved in the histochemical demonstration of dihydrolipoic dehydrogenase are discussed.
CYTOCHEMISTRY OF PARACOCCIDIOIDES BRASILIENSIS. I. CYTOCHEMISTRY OF CYTOPLASMIC POLYSACCHARIDES IN YEAST FORM CULTURES WITH LIGHT MICROSCOPE
Authors: CARBONELL LUIS M. , CASTEJON HAYDEE , POLLAK LADISLAO
Published: April 2011

1. The histochemical distribution and localization of cytoplasmic polysaccharides are studied in the yeast form of P. brasiliensis. Glycogen and an acid mucopolysaccharide probably containing sulfate groups are demonstrated. These polysaccharides disappear after the sixth day of culture. The glycogen and the acid mucopolysaccharide are bound to protein.

2. Stains and enzyme treatments applied in sequence help demonstrate the site of different polysaccharides.
THE USE OF THE GEVAERT NUC-307 NUCLEAR EMULSION FOR RADIOAUTOGRAPHY AT THE ELECTRON MICROSCOPE LEVEL
Authors: YOUNG B. A. , KOPRIWA BEATRIX M.
Published: April 2011

The fine-grain Gevaert NUC-307 emulsion was investigated for use in radioautography. First, an attempt was made to develop grains visible in the light microscope in order to compare the sensitivity and the background fog of this emulsion with that of other emulsions and thus obtain the data required to devise an adequate set of conditions for work with the electron microscope. A good method for electron microscopic radioautography was to dip EM grids attached to glass slides in undiluted NUC-307 emulsion warmed to 40°C. The exposed preparations were developed for 5 minutes in D-170 at 18°C. The silver grains were found to be smaller and, therefore, to allow a better resolution than those of Ilford L4 emulsion. The high resolution as well as the high sensitivity of the NUC-307 emulsion make it well suited for electron microscopic radioautographs.
ACID DEOXYRIBONUCLEASE IN RAT LIVER CELL NUCLEI ISOLATED IN THE PRESENCE OF CALCIUM IONS
Authors: SWINGLE KARL F. , COLE LEONARD J.
Published: April 2011

Nuclear fractions centrifugally separated from rat liver homogenates in isotonic sucrose or in water, containing in each case 1.8 mM CaCl2, and washed repeatedly in the same media, retained a considerable amount of acid deoxyribonuclease activity. Similarly prepared nuclear fractions using calcium-free media lost nearly all of their deoxyribonuclease activity. This retention of activity in the presence of calcium was shown to be neither cosedimentation of unbroken cells or of lysosomes with the nuclear fraction nor precipitation of soluble deoxyribonuclease in the presence of calcium. The nuclear-bound enzyme was active under conditions shown not to remove it from the nuclei.
FACTORS AFFECTING THE CYTOCHEMICAL REACTION OF PEROXIDASE WITH BENZIDINE AND THE STABILITY OF THE BLUE REACTION PRODUCT
Authors: STRAUS WERNER
Published: April 2011

The influence of pH, temperature, time of incubation, concentration of the substrate and acceptor, addition of alcohol and salts on the cytochemical reaction of peroxidase with benzidine and H2O2 was investigated. The stability of the blue and brown reaction products towards oxidizing and reducing agents was tested. It was observed that a slightly acid pH and low temperature are main factors for the formation of a blue rather than of brown or yellow reaction products of peroxidase with benzidine. A procedure was described for the stabilization of the blue reaction product consisting in the treatment of the tissue with 70% alcohol solutions of sodium nitroprusside, buffered at pH 5.0, subsequent to the staining with benzidine. Crystallization artifacts of the blue pigment could be prevented in the absence of added salts (buffers) by regulating the amount of "benzidine blue" produced by peroxidase. The main variables used for this regulation were the time of incubation with benzidine and H2O2 at 0-4°C, and secondary variables were the concentrations of benzidine, H2O2, and added alcohol. Procedures for the staining of peroxidase in alcohol-containing and in aqueous solutions of benzidine were described and compared. The specificity of the cytochemical reaction of peroxidase with benzidine was discussed.
QUANTITATIVE STUDIES ON ISOLATED PANCREATIC ISLETS OF MAMMALS: ADENOSINE TRIPHOSPHATASE ACTIVITY IN NORMAL AND OBESE-HYPERGLYCEMIC MICE
Authors: TÄLJEDAL INGE-BERT , HELLMAN BO , HELLERSTRÖM CLAES
Published: April 2011

Microchemical and histochemical methods were used for the characterization, localization and assay of adenosine triphosphate (ATP) splitting enzymes in homogenates and sections of the endocrine pancreas from obese-hyperglycemic mice and their lean litter mates. The following observations were made:

1. Dephosphorylation of ATP was maximal at pH 9.1. It was strongly stimulated by magnesium ions at an optimal concentration of 1 mM. ATP cleavage was inhibited by adenosine diphosphate, sodium azide and p-chloromercuribenzoic acid. The addition of l-cysteine, sodium cyanide or sodium fluoride to the substrate medium had no effect on the enzyme activity. Substitution of ATP by equimolar amounts of other phosphate esters in the medium considerably reduced the substrate cleavage. These results are taken as evidence for the presence of sulfhydryl-dependent adenosine triphosphatase (ATPase) in the islet tissue.

2. Histochemical staining revealed a strong ATP splitting enzyme activity in the capillaries and walls of larger blood vessels throughout the pancreas; a rather weak and diffuse cytoplasmic reaction being found in the islet cells.

3. Microchemical assays revealed a lower cleavage of ATP in the islets as compared with the exocrine pancreas and the liver. The cleavage of ATP was more intense in the islets of the obese-hyperglycemic mice than in those of the lean litter mates.

4. Starvation for 7 days induced no significant changes in the enzyme activity of the endocrine pancreas.
CARBOHYDRATE HISTOCHEMISTRY OF THE SALIVARY GLANDS AND PANCREAS IN CYSTIC FIBROSIS
Authors: SHACKLEFORD JOHN M. , BENTLEY HERSCHEL P.
Published: April 2011

Various histochemical methods for demonstrating complex tissue carbohydrates were applied to salivary gland and pancreatic tissue from four known cases of cystic fibrosis. Aside from the usual methods, e.g., periodic acid Schiff and alcian blue, the p-hydrazinobenzenesulfonic acid reaction was found to be particularly valuable as a survey method for estimating concentrations of vicinal hydroxyl groups. In the pancreas, submandibular and parotid glands striking changes in carbohydrate histochemistry occur in the small intralobular (intercalated) ducts which normally secrete relatively little or no mucopolysaccharide. The lumina of salivary gland striated ducts and large excretory ducts frequently contain inspissated mucus. In the sublingual glands the inspissated mucus is mixed with considerable amounts of deoxyribonucleic acid. Vibrio cholera neuraminidase significantly reduces acid group staining in most of the epithelial mucus although some resistance to digestion by this enzyme is evident. Some acid group substances appear in the interstices of sublingual glands which exhibit the histochemical characteristics of epithelial mucin. This reaction is consistent with a "leakage" of some of the sialomucins from the acini into the interstitial areas.
EXTRACTION RESISTANCE OF SUDAN STAINED MAST CELLS AFTER PREVIOUS ACID TREATMENT
Authors: FEAGLER JOHN R. , MCMANUS J. F. A.
Published: April 2011

1. Sudan black B alone stains the mast cells very faintly. The granules do not appear very distinct or numerous.

2. The periodic acid Schiff (PAS) technique stains the mast cells; however, the results are variable. When stained, the mast cell granules appear slightly darker than the cytoplasm.

3. When PAS and Sudan black are used together, the mast cell granules gain an avidity for the dye and are stained black.

4. Brief acid treatment prior to immersion in Sudan black results in granules gaining an avidity for the Sudan dye that is extraction resistant. The reaction appears dependent upon modifying the mast cell granules by acid treatment. It appears as though the hydrogen ion changes some physical or chemical property of the granules so that they gain an avidity for the dye.

5. Acetylated Sudan black shows an affinity for the mast cell granules greater than Sudan black alone. This is not increased by previous acid treatment. Acetylated Sudan black also stains the mast cell nucleus.
SYNTHESIS AND PROPERTIES OF MODEL SYSTEMS, WITH THEIR USE IN STUDYING THE SCHIFF REACTION IN HISTOCHEMISTRY
Authors: HARDONK M. J. , VAN DUIJN P.
Published: April 2011

The synthesis of cellulose films containing several reactive groups is described. These films were developed for use as histochemical model systems to study several Schiff-type reactions.

The synthesized model systems include cellulose membranes to which amino groups are covalently bound, cellulose membranes containing sulfhydryl groups, and cellulose membranes to which deoxyribonucleotides have been bound by means of their phosphate group.

Evidence is given for the structure of these cellulose derivatives.
NEW SUBSTRATES OF TRYPSIN AND SOME TRYPSIN-LIKE ENZYMES
Authors: GLENNER G. G. , HOPSU V. K. , COHEN L. A.
Published: April 2011

The results of the investigation of two new naphthyl substrates of trypsin and the trypsinlike enzyme(s) in human mast cells, carbobenzyloxy-l-arginine and benzoyl-l-arginine β-naphthylamide, have led directly to the synthesis and study of a further substrate, carbobenzyloxy-l-arginine methyl ester. The activity of trypsin against the carbobenzyloxy ester is greater than that previously described biochemically for other synthetic substrates. The inhibition of the amidase and esterase activity of trypsin and the tryptic enzyme in human mast cells by ε-aminocaproic acid derivatives is characterized and evidence is presented to show that this enzymic activity in human mast cells is more closely related to trypsin than to plasmin (fibrinolysin).
HISTOCHEMICAL STUDIES ON THE ROLE OF THE MAST CELL IN CALCERGY
Authors: SELYE HANS , GABBIANI GIULIO , SERAFIMOV NIKOLA
Published: April 2011

Experiments were performed on rats, using special histochemical stains for the demonstration of metachromatic materials, lead, calcium, phosphate and carbonate, to study calcergy, the induction of soft-tissue calcification by topical treatment with certain compounds such as lead acetate.

In simple calcergy, when lead acetate is directly injected into the subcutis, it is seen to impregnate the collagen fibers in the injection site without noteworthy participation of the mast cells. The lead-treated area secondarily attracts calcium, phosphate and possibly carbonate.

In mastocalcergy, the lead acetate is injected intravenously and local calcification is produced by the simultaneous subcutaneous injection of a mast-cell discharger such as polymyxin. Here, the mast cells in the treated area first show degranulation without any mineralization, but this is soon followed by lead uptake on the mast-cell granules with secondary attraction of calcium, phosphate and possibly carbonate. Finally, the discharged and calcified mast-cell granules disintegrate into a dustlike, fine precipitate which is transferred to the circumjacent collagen fibers where it initiates an intense process of mineralization, eventually leading to the complete petrification of the polymyxin-injected area.

Apparently, in addition to its already well established functions of histamine, heparin and serotonin production, the mast cell also plays a role in mineralization.