bluuebunny/crossref_metadata_2025 huggingface dataset:
THE IN VIVO STAINING OF THE LINING OF MOUSE FORESTOMACH BY PORPHYRINS AND OTHER FLUORESCENT SUBSTANCES
Authors: FIGGE FRANK H. J.
Published: April 2011

1. It was observed that the forestomach in rats and mice became red fluorescent when these animals were maintained on a diet of chlorophyll-containing "Fox Chow".

2. Little or no red fluorescence was observed in the forestomach when mice were fed "Dog Chow", white bread, or peeled boiled potatoes.

3. The materials that gave rise to red fluorescence of the forestomach originated from two main sources:

a. the chlorophyll porphyrin derivatives (chiefly phylloerythrin) resulting from the degradation of chlorophyll in the food, and

b. the porphyrins in the red fluorescent Harderian gland secretions that passed to the stomach via the naso-lacrimal duct, nasopharynx and esophagus.

4. The porphyrins from these sources or porphyrins artificially added to the diet had a tendency to accumulate in the mucus of the forestomach, but not in the mucus of the glandular stomach. Methylcholanthrene, when fed with "Dog Chow", was also observed to accumulate in the forestomach mucus. Other fluorescent substances, when added to the diet, do not stain the mucus of the gastro-intestinal tract so selectively.

5. A pH difference (or double gradient) resulting from the secretion of acid in the glandular (middle) part of the stomach was found to be responsible for this differential diffusion of porphyrin into the mucus of the two parts of the stomach. The effect was thought to be related to the pH solubility curves of the porphyrins and/or to the fact that gastric mucus at its isoelectric point (pH 3) becomes permeable to some substances. When the pH of the alimentary contents was within the range of minimum water solubility of a given porphyrin, the rate of diffusion of the porphyrin into the mucus was greatest.
THE HISTOCHEMICAL LOCALIZATION OF ZINC IN THE DORSOLATERAL PROSTATE OF THE RAT
Authors: RIXON R. H. , WHITFIELD J. F.
Published: April 2011

High concentrations of Zinc in the lateral portions of the dorsolateral prostate of the rat have been associated histochemically with a luminal border organelle characteristic of the epithelium. This border organelle is osmiophilic and argentophilic and contains a high concentration of basophilic material as well as zinc. The organelle is distinct from the Golgi bodies in the epithelial cells. Epithelium from the dorsal portions of the gland lacks this border organelle. High concentrations of zinc and basophilia are also associated with the sub-epithelial stroma in the lateral parts of the prostate and the nucleoli of both parts of the gland. In the dorsal part of the prostate, zinc was detected only in the nucleoli. The possible significance of these results is discussed.
QUANTITATION OF PERIODIC ACID-SCHIFF POSITIVE SUBSTANCES USING RADIOACTIVE SCHIFF REAGENT
Authors: BARKA TIBOR , ORNSTEIN LEONARD
Published: April 2011

A simple method is described which makes it possible to determine the relative amount of Schiff reagent bound to periodic acid oxidized tissue sections. It is based on the application of S35-labeled Schiff reagent. It was found that paraffin embedded tissue sections bind significant amounts of Schiff reagent without any previous treatment, but using appropriate control sections it is possible to determine the relative amounts of aldehyde groups created by periodic acid oxidation as opposed to the amounts in the controls.
ACTION OF PROCAINAMIDE, QUINIDINE AND CHLOROQUINE UPON THE CONDUCTION SYSTEM OF THE DOG'S HEART (HISTOCHEMICAL STUDY)
Authors: SANABRIA A. , CARBONELL L. M. , SOTO R.
Published: April 2011

The effect of quinidine, chloroquine and procainamide on the conducting system of the dog was studied histochemically. There was inhibition of succinic dehydrogenase when chloroquine was administered intravenously and of alpha esterase when procainamide was used.
THE CORRELATION OF ULTRAVIOLET AND FEULGEN ABSORPTION MEASURED IN ASCITES TUMOR CELLS
Authors: BARKA TIBOR , DALLNER GUSTAV
Published: April 2011

The correlation of ultraviolet and Feulgen absorption after ribonuclease, cold perchloric acid, or ribonuclease plus perchloric acid treatments was investigated in fixed ascites tumor cells using microspectrophotometric methods. It was concluded from the measurements that the amount of ribonucleic acid resistant to 24 hour ribonuclease digestion is proportional to the amount of desoxyribonucleic acid revealed by Feulgen staining.
A NITROSOPHENOL REACTION FOR TYROSINE AND RELATED COMPOUNDS IN TISSUE SECTIONS
Authors: GLENNER G. G.
Published: April 2011

1. A method for the histochemical localization of protein-bound tyrosine and other ortho unsubstituted substituted phenols by the formation of a nitrosophenol-metal chelate is described.

2. The histochemical formation of a nitrosophenol confirms the postulated first step in the diazotization-coupling reaction for the histochemical demonstration of protein-bound tyrosine.

3. Though the described method is useful as a confirmatory technique for the localization of protein-bound tyrosine, its sensitivity is markedly less than that of the diazotization-coupling reaction.
A HISTOCHEMICAL STUDY OF ENZYMES IN THE EPIDIDYMIS OF NORMAL, CASTRATED AND HORMONE REPLACED CASTRATED MICE SEPARATED BY ZONE ELECTROPHORESIS IN STARCH GELS
Authors: ALLEN JOHN M. , HUNTER ROBERT L.
Published: April 2011

The ability of aliesterase, alkaline phosphatase, and acid phosphatase of mouse epididymis separated in a starch matrix by electrophoresis to attack cytochemical substrates has been investigated in tissues from normal, vasectomized, castrated, and testosterone propionate treated castrated, animals. Electrophoresis at pH 8.5 resolved 7 enzymes active against α-naphthyl propionate (designated A through G). Castration depressed the activity of A, B, and C; abolished the activity of D, E, and G; and increased the activity of F. Androgen administration restored the activity of all enzymes except that of G. D and E were more active in control animals than in vasectomized animals. Electrophoresis at pH 8.5 resolved 3 enzymes active against naphthol-AS acetate. These were identical to those designated D, E, and F above. Castration depressed the activity of D and E and increased the activity of F. Androgen treatment restored activity to levels in control animals.

Electrophoresis at pH 8.5 resolved one enzyme active against α-naphthyl phosphate at pH 8.0 (alkaline phosphatase). This enzyme was more active in tissues from vascetomized than from control animals. Castration abolished its activity and androgen treatment restored it to levels seen in control animals.

Electrophoresis at pH 8.5 resolved one enzyme active against α-naphthyl phosphate at pH 6.0 (acid phosphatase). The activity of this enzyme was similar in tissues from normal, vascetomized, castrated, and androgen treated castrated animals.
HISTOCHEMICAL DEMONSTRATION OF CYTOCHROME OXIDASE WITH NEW AMINE REAGENTS
Authors: BURSTONE M. S.
Published: April 2011

A series of aryl amines were employed as new histochemical reagents for the demonstration of cytochrome oxidase. The amines, upon oxidation, form indamine-azine type dyes which are capable of chelating with a number of metals.

The most useful substrate consisted of a mixture of N-phenyl- p-phenylenediamine (I) and p-methoxy- p-aminodiphenylamine (II) or 3-amino-9-ethyl-carbazole (V).

The reaction was inhibited by pretreatment of sections with saline, and augmented by the addition of cytochrome c. Heat, cyanide, and sulfide were inhibitory.

Although raw-frozen sections were employed, mitochondrial localizations were obtained in specific instances in selected areas of certain tissues.

The possibility of demonstrating other oxidative enzymes with these substrates warrants further study.
PRESENCE OF TRIMETAPHOSPHATASE IN THE INTESTINAL MUCOSA AND PROPERTIES OF THE ENZYME
Authors: BERG GEORGE G. , GORDON L. HEICKLEN
Published: April 2011

Trimetaphosphatase was shown to be present in intestine. Enzyme extracted from rat duodenum had optimum activity at pH 4, and was not activated by divalent cations. There was no inhibition by Mg++, weak inhibition by Ca++, Zn++, Co++, and Mn++, and strong inhibition by VO++ and possibly by Be++, Fe++, and Cu++. The enzyme was relatively resistant to organic solvents, formaldehyde, and KCN, and readily inactivated by NaF and KSCN. The bulk of the enzyme was bound to a sedimentable fraction, the remainder was soluble in water. Histochemical localization in the rat was in the lamina propria mucosae of the entire small intestine. In the mouse the enzyme was localized in the cytoplasm of the mucous epithelium of the small intestine.
ALKYLATION OF AMINES: A STUDY OF METHYLATION AND ITS EFFECTS ON AMINES
Authors: TERNER JACOB Y. , CLARK GEORGE
Published: April 2011

The effect of normal methylation procedures upon tissue was reexamined. It was demonstrated that the methyl alcohol and HCl prevent the oxidative destruction of amines by either chloramine T or nitrous acid while effecting in no way their salt forming properties. It was shown that the ethyl chloride in methanol will give much the same effect. The authors conclude that methylation at higher temperatures alkylates amines.
GLYCOGEN FRACTIONS AND THEIR ROLE IN THE HISTOCHEMICAL DETECTION OF GLYCOGEN
Authors: KUGLER J. H. , WILKINSON W. J. C.
Published: April 2011

Glycogen fractions have been investigated in guinea pig liver by the anthrone technique and histochemically by Gomori's hexamine silver method.

It is concluded that it is only the trichloroacetic acid soluble fraction of tissue glycogen that is demonstrated by this histochemical technique.
THE HISTOCHEMICAL DEMONSTRATION OF α-d-GLUCOSIDASE IN MAMMALIAN TISSUES
Authors: RUTENBURG ALEXANDER M. , GOLDBARG JULIUS A. , RUTENBURG SELMA H. , LANG RUTH T.
Published: April 2011

A histochemical method for the demonstration of α-d-glucosidase is presented. The procedure and the problems encountered in its development are discussed. Surveys of mammalian tissues showed that this enzyme is generally demonstrable by this method only in the mucosa of the duodenum and upper jejunum and in the kidney cortex. Starvation resulted in decreased enzymatic activity in the intestinal mucosa, whereas increased activity was observed after the ingestion of food.
STUDIES ON STREPTOCOCCAL HYALURONIDASE AND ANTIHYALURONIDASE. III. THE PRODUCTION AND CELLULAR LOCALIZATION OF HYALURONIDASE FOLLOWING STREPTOCOCCAL INFECTION
Authors: EMMART E. W. , TURNER W. A.
Published: April 2011

I. Streptococcal hyaluronidase and the bacterium Streptococcus hemolyticus, Group C, strain 7, have been identified within the same tissue section by tagging the former with its specific antibody coupled to yellow-green fluorescein and the latter with Group C antisera coupled to acid rhodamine B (C.I. No. 45100).

2. The methods used above suggest a means of studying the absorption and activity of other antigenic enzymes and toxins elaborated during the course of infection.
AZO-COUPLING RATE OF ENTEROCHROMAFFIN WITH VARIOUS DIAZONIUM SALTS
Authors: LILLIE R. D. , GRECO-HENSON JACQUELINE P. , CASON JOSEPH C.
Published: April 2011

The azo-coupling of enterochromaffin with some 30 diazomium salts has been explored, in regard to rate, concentration and pH. A technic of enterochromaffin cell counting in consecutive serial sections was employed and the counts related to those obtained in immediately adjacent serial sections reacted with the ferric ferri-cyanide reagent of Golodetz and Unna (6) as modified by Lillic and Burtner (8). This procedure enabled the equation of a considerable series of experiments with different diazotates on a succession of individual animals.

Structure of the diazonium salt proved to have a considerable influence on the azo coupling rate. Of the nitroanilines, nitroanisidines and nitrotoluidines, those with the nitro group para to the diazonium radical coupled rapidly, those with ortho nitro groups slowly. Also p-anisidine was more rapid than the ortho isomer.

In the specific cases of p-nitroaniline and of di- o-anisidinse fresh diazotates were found to be more rapid and effective couplers than commercial stabilized salts in similar concentrations. It is probable that other comparisons of the same sort would give similar results.

While azo coupling increased in rapidity with higher diazo concentrations, increasing density of background reaction decreased the demonstrability of enterochromaffin, so that optimal concentrations were often 1-10 mM, but lower with "rapid" couplers or higher with slow.

The second diazonium group of fast blue B is probably but little utilized in azo coupling with enterochromaffin under histochemical conditions, since di- o-anisidine treated with one molar equivalent of nitrous acid appeared almost if not quite as effective as when two molar equivalents were used.

Fast blue B appears to fall into an intermediate group of diazotates, being neither as fast as the p-nitro derivatives nor as slow as fast blue RR, BB or VB, for example.
MODIFICATIONS OF HISTOCHEMICAL TECHNIQUES FOR THE DEMONSTRATION OF CYTOCHROME OXIDASE
Authors: BURSTONE M. S.
Published: April 2011

Four reagents were employed in conjunction with p-aminodiphenylamine as new histochemical substrates for cytochrome oxidase. The most useful

[See figures in PDF file]

substrates were 8-amino-1,2,3,4-tetrahydroquinone (I) and 8-hydroxy-1,4-naphthoquinone (III). (I) forms blue dyes and is recommended for general use, as well as for tissues of low activity (human gingiva). With the latter, the reaction may be augmented by the addition of cytochrome c.

The dye produced from (III) has a strong affinity (substantivity) for tissue elements and resists dehydration with solvents.

The new-substrates were useful in visualizing cytochrome oxidase activity in the digestive tract including oral mucosa, central nervous system, muscle and bone.
CYTOLOGICAL LOCALIZATION OF CHOLINESTERASE IN CULTURED SKELETAL MUSCLE CELLS
Authors: ENGEL W. KING
Published: April 2011

The location of acetyl and butyryl thiocholinesterase activity was studied in cultured chick embryo skeletal muscle cells which were growing without being innervated. Muscle cells in all three stages of development, i.e., the single myoblast, the plurinucleated myoblastic strap, and the multinucleated myocytic strap, showed sites of thiocholinesterase activity as small brown granules located diffusely in the cytoplasm and accentuated in the perinuclear areas. In the well differentiated myocytic straps, thiocholinesterase was localized along the Z lines. No enzyme activity was found at the sarcolemma or in the nuclei. No thiocholinesterase-containing motor end-plate structures were found in any stage of development.
DISTRIBUTION OF ZINC IN BONE TISSUE
Authors: HAUMONT S.
Published: April 2011

High concentrations of zinc have been found by a histochemical method at the sites of calcification. Zinc has been detected in the developing osteons (Haversian systems) of compact bone exactly at the border line between calcified and uncalcified tissue (Fig. 1), in the cartilaginous partitions of hypertrophic cells (Fig. 3), and in the endochondral bone which has just been deposited in the metaphysis (Fig. 4).
HISTOCHEMICAL DEMONSTRATION OF SUCCINIC DEHYDROGENASE IN THE FAT BODY OF THE DESERT LOCUST AND SOME GRASSHOPPERS
Authors: GEORGE J. C. , HEGDEKAR B. M.
Published: April 2011

Succinic dehydrogenase activity in the whole fat body of the desert locust and three species of grasshoppers has been demonstrated histochemically. The localization of the enzyme has been observed to be confined to the periphery of the cells. Fat solvents like ethyl acetate, ether, 3:1 ether-acetone mixture, or acetone alone, have been found to inhibit the enzyme, as was also the case with frozen sections or air dried material. Failure to demonstrate the activity of the enzyme in the latter case may be attributed to the possible release of an inhibitor, as has been already suggested by earlier authors in the case of homogenization.
THE DISTRIBUTION OF RIBONUCLEASE ACTIVITY IN RAT LIVER DURING AZO-DYE CARCINOGENESIS
Authors: AMANO HARUKO , DAOUST R.
Published: April 2011

The distribution of ribonuclease activity in the liver of azo-dye fed animals was investigated histochemically using the substrate film method.

The ribonuclease activity of liver parenchyma was lower than normal in cirrhotic and neoplastic rat livers while high activities were observed in the bile duct and connective tissue trabeculae. Intense reactions were also given by the necrotic regions.

The decrease in ribonuclease activity of parenchymal cells was found to take place at early stages of 4-dimethylaminoazobenzene feeding, that is before hyperplastic cells are transformed into cancerous cells. No similar variation could be observed in the liver of animals fed the control basal diet nor in normal regenerating liver following partial hepatectomy.
STUDIES OF SIALIC ACID-CONTAINING MUCINS IN BOVINE SUBMAXILLARY AND RAT SUBLINGUAL GLANDS
Authors: QUINTARELLI G. , TSUIKI S. , HASHIMOTO Y. , PIGMAN W. , DELLOVO MARIE C.
Published: April 2011

Sialic acid present in some salivary mucins could be removed from tissue sections by mild acid hydrolysis. Bovine submaxillary glands, incubated in acid medium prior to staining, did not show any metachromasia or alcianophilia, whereas the periodic acid Schiff reaction was diminished but not eliminated. When influenza virus neuraminidase was utilized, the results were essentially the same as those obtained after mild hydrolysis. Chemical studies confirmed the histochemical results. It was found that when submaxillary mucin underwent acid hydrolysis, sialic acid was easily split off without further breaking of the mucin molecule. When proteolytic enzymes were employed, the acini could not be stained by any of these methods. These results with proteolytic enzymes are interpreted as due to a fragmentation and solubilization of the mucin molecule.

Different results were obtained for rat sublingual glands. Acid hydrolysis removed the safranin O metachromasia as well as the alcianophilic material, but the time and temperatures needed were different from those for bovine submaxillary tissue. The periodic acid Schiff staining properties did not change. The same difference was noted when proteolysis was utilized, and the time required for complete digestion of the acinar content was three times greater than for bovine tissue. Furthermore, influenza virus neuraminidase had no effect on the rat gland sections.

The present data suggest that the use of mild acid hydrolysis and of proteolysis applied to tissue sections containing high amounts of sialic acid can be a useful tool in practical histochemical techniques for studies on sialomucins.
INVESTIGATIONS ON THE STRUCTURE OF THE ENTEROCHROMAFFIN SUBSTANCE
Authors: LILLIE R. D.
Published: April 2011

Summarizing, a catechol structure for enterochromaffin seems more indicated than a hydroquinone structure. Any aminophenol struture, primary or secondary, seems contraindicated. The resorcinol hypothesis seems untenable. Serotonin is chemically and sometimes histochemically demonstrable in carcinoid tumors, but its presence in normal enterochromaffin cells in demonstrable amounts seems contraindicated. The azo positive phenol in the enterochromaffin cells appears to be a distinct substance.
STUDIES OF INDOPHENOL BLUE SYNTHESIS: I. THE ROLE OF FREE RADICAL FORMATION BY HEART MUSCLE PARTICULATES DURING THE "G" NADI REACTION
Authors: PERSON PHILIP , FINE ALBERT
Published: April 2011

A study of the "G" Nadi reaction mediated by Keilin and Hartree beef heart muscle preparations has revealed that of the two Nadi reagents, i.e., dimethylparaphenylenediamine and α-naphthol, only the diamine is enzymatically oxidized by the terminal respiratory enzymes of the heart particulates. In both spectrophotometric and manometric experiments, enzymatic oxidation of α-naphthol by the heart muscle enzymes could not be detected. Oxidation of the diamine resulted in formation of a stable free radical and polymers with characteristic absorption spectra. The free radical and polymers, upon formation, rapidly reacted with the α-naphthol in a non-enzymatic oxidative coupling to form indophenol blue. Incubation of dimethylparaphenylenediamine alone with heart muscle particles resulted in production of an insoluble blue dye in the particles. Upon extraction with 95% ethanol, the pigment, which is not indophenol blue, exhibited a broad absorption maximum at 460-470 mµ.
THE LOCALIZATION OF N-ACETYL-β-GLUCOSAMINIDASE IN TISSUES
Authors: PUGH DOREEN , WALKER P. G.
Published: April 2011

α-Naphthyl N-acetyl-β-glucosaminide has been prepared and used in a simultaneous azo-dye coupling method for the demonstration of N-acetyl-β-glucosaminidase activity in formalin fixed frozen sections

The effect of fixation on the activity of the enzyme has been invesitgated. In addition the effects of varying the substrate concentration, amount of diazonium salt and pH of the incubating medium are described

N-Acetylglucosaminonolactone, a specific inhibitor of N-acetyl-β-glucosaminidase has been used in experiments designed to test the specificity of the reaction.

The relative activity of the enzyme in rat tissue homogenates has been measured and its distribution in rat tissues and human cartilage and synovial membrane is described

A satisfactory localization of N-acetyl-β-glucosaminidase is possible by the technique described. Factors affecting the histochemical localization are discussed in relation to the results obtained with other enzymes
HISTOLOGICAL LOCALIZATION IN RAT TISSUES OF INTRAVENOUSLY INJECTED THYROTROPHIN LABELED WITH A FLUORESCENT DYE
Authors: MANCINI R. E. , VILAR O. , DELLACHA J. M. , DAVIDSON O. W. , CASTRO A.
Published: April 2011

Several Armour thyrotrophin preparations were labeled with different amounts of a fluorescent dye, acid rhodamine B (Lissamine-Rhodamine B 200, Imperial Chemical Industries, Ltd.) following Chadwick's method with slight modifications. The native thyrotrophin lost from 12 to 34% of its biological activity after labeling, proved by Gilliland's method. A portion of the same labeled thyrotrophin was denaturated by heating in acetic acid medium. Different lots of rats were intravenously injected with native labeled thyrotrophin, denaturated labeled thyrotrophin and fluorescent dye solution respectively.

It was observed: 1) Native labeled thyrotrophin declined rapidly in the circulation, though it remained longer than denaturated labeled thyrotrophin. 2) Fluorescence corresponding to native thyrotrophin was seen for a very short time in the thyroid gland and longer in the connective tissue of ocular structures, interstitium of skeletal muscles, perivisceral adipose tissue and mast cells. The Kupffer cells and other macrophages as well as the proximal tubule cells showed the longest accumulation. 3) Fluorescence corresponding to denatured labeled thyrotrophin was not observed in the thyroid gland; it was seen with low intensity in the connective and adipose tissues as well as in the mast cells. It was observed with higher intensity in the macrophages, convoluted tubule cells and it was seen to be eliminated through the epithelium of the small intestine. 4) Rats injected with acid rhodamine B solution eliminated the dye through the urinary and bile systems. In these animals fluorescence was seen only in the macrophages and proximal tubule cells, though in lesser amount compared with the animal injected with labeled hormone.
BIOCHEMICAL AND HISTOCHEMICAL IDENTIFICATION OF SIALIC ACID CONTAINING MUCINS OF RODENT VAGINA AND SALIVARY GLANDS
Authors: WARREN LEONARD , SPICER SAMUEL S.
Published: April 2011

The mucins elaborated in the mucification reaction of the vagina of the rat and mouse, like salivary mucins, show the histochemical properties of a nonsulfated acid mucopolysaccharide. In the mouse these have been identified as sialomucins on tile basis of the elimination of basophilia from tissue sections by specific digestion with purified sialidase. However, the removal of sialic acid effected by sialidase is not accompanied by an alteration in periodic acid Schiff staining and hence extensive degradation of the polymer does not take place.

Sialidase alters the staining of mouse sialomucins quickly in the sublingual gland, relatively slowly in the vagina and even more slowly in the submaxillary gland. Digestion with the enzyme fails to alter staining reactions in the siahic acid-rich, histochemically similar mucin in the salivary gland and vagina of the pregnant rat, and fails to release sialic acid either from histologic sections or homogenates of these rat organs.

Changes in the sialic acid level in the vagina during the oestrous cycle parallel the cyclic variations of the amount of mucin observed histochemically. Both are consistently elevated in pregnancy. The sialic acid concentration is low and mucin is absent in oestradiol treated pre-ovulatory mice, but these increase with subsequent progesterone treatment.

Identification of the sialic acid by means of paper chromatography reveals that the rat and mouse vagina and sublingual and submaxillary glands all contain N-acetylneuraminie acid and that mouse vagina and rat submaxillary gland also contain N-glycolylneuraminic acid. An un-identified form of neuraminic acid with rapid mobility in two solvents is present in the rat submaxillary gland.
THE USE OF SODIUM CHLORIDE-CONTAINING FIXATIVES IN MINIMIZING CELLULAR DISTORTION IN HISTOLOGICAL AND CYTOCHEMICAL PREPARATIONS
Authors: BIRGE WESLEY J. , TIBBITTS F. DONALD
Published: April 2011

The principal objective of this study was to determine the relative effect of the concentration of small, mobile ions in fixing solutions on cellular shrinkage and distortion occurring during fixation or in the subsequent processing of tissues by the paraffin method.

Samples of rat liver tissue were fixed in neutral 20% formalin, neutral 20% formalin/0.7% sodium chloride, Bouin's fluid, and Bouin's fluid/0.7% sodium chloride. By a comparison of nuclear and cytoplasmic volumes, it was noted that in the case of formalin fixation, the addition of 0.7% sodium chloride reduced nuclear and cytoplasmic shrinkage by 18-26%. In Bouin fixation the addition of 0.7% sodium chloride reduced nuclear shrinkage by 7-10% and cytoplasmic shrinkage by approximately 21%.

It was also shown that when the sodium chloride is omitted, neutral 20% formalin is significantly less effective than Boumin's fluid in stabilizing the cytoplasm of liver parenchymal cells against shrinkage.

In those instances in which tissue shrinkage was minimized by the addition of sodium chloride to the fixing medium, the final preparations revealed less tissue distortion and a more adequate preservation of fine detail. Some mention is also made of differences incurred in staining reactions of liver tissue following the use of sodium chloride containing fixatives.
A METHOD OF DEMONSTRATING CALCIUM IN TISSUE SECTIONS USING CHLORANILIC ACID
Authors: CARR LAWRENCE B. , RAMBO OSCAR N. , FEICHTMEIR THOMAS V.
Published: April 2011

A rapid and simple method for selective staining of calcium in histologic sections depends on the formation of an insoluble, colored and doubly refractile complex between the calcium cation and chloranilic acid derived from aqueous solution. The method is described.
A CYTOCHEMICAL STUDY OF GOLGI ASSOCIATED THIAMINE PYROPHOSPHATASE IN THE EPIDIDYMIS OF THE MOUSE
Authors: ALLEN JOHN M. , SLATER JUDITH J.
Published: April 2011

The properties and distribution of an enzyme in the epididymis of the mouse hydrolyzing thiamine pyrophosphate were investigated using cytochemical and quantitative methods. Cytochemical study indicated thiamine pyrophosphatase to be restricted to the Golgi regions of the epithelial cells lining the epididymal canal. Activity due to this enzyme was easily separated from that due to alkaline phosphatase (which also hydrolyzed thiamine pyrophosphate) on the basis of differences in localization and on the greater sensitivity of the latter enzyme to cysteine. The thiamine pyrophosphatase present in the epididymis of the mouse had optimal activity in the region of pH 9.5. In the acid range, activity was greatly restricted but the distribution pattern did not differ from that observed in the alkaline range nor did it resemble the pattern of activity associated with acid phosphatase activity in this tissue. Quantitative determinations supported the cytochemical investigations in terms of a pH optimum in the alkaline range and the resistance of thiamine pyrophosphatase to cysteine inhibition.
THE EFFECT OF HORMONE TREATMENT ON THE PHOSPHORYLASE ACTIVITY OF THE RABBIT UTERUS
Authors: BO WALTER J.
Published: April 2011

Glycogen and phosphorylase activity were identified in the uterus of hormone treated rabbits. Glycogen granules were very abundant in both layers of the myometrium following estrogen stimulation. In the ovariectomized, progesterone or estrogen-progesterone treated animals only a small number of glycogen granules was detected in the myometrium.

In the uteri of the rabbits treated with estrogen, phosphorylase activity was very intense in both layers of the myometrium. Only a trace of the enzyme was observed in the muscle layers of the animals treated with progesterone or with estrogen and progesterone. In the ovariectomized rabbits phosphorylase was observed in very small amounts in the outer part of the longitudinal layer.
THE HISTOCHEMICAL DEMONSTRATION OF PROTEASE BY A GELATIN-SILVER FILM SUBSTRATE
Authors: ADAMS C. W. M. , TUQAN N. A.
Published: April 2011

A method is described for the histochemical demonstration of proteolytic enzymes by digestion of a gelatin-silver film. The effect of inhibitors, fixation and heat upon pancreatic proteases and peripheral nerve cathepsin is discussed
HISTOCHEMISTRY OF MYELIN. III. PERIPHERAL NERVE CATHEPSIN
Authors: ADAMS C. W. M. , BAYLISS OLGA B.
Published: April 2011

Some of the properties of peripheral nerve cathepsin were investigated by histochemical and biochemical means. This cathepsin appears to be a constituent of the myelin sheath but its presence in the axon as well cannot be excluded. The enzyme is thermostable, sulphydryl-dependant and resistant to chloroform, methanol and acetone. It is mainly associated with the lipid and proteoplipid fractions of the nerve and this distribution supports the view that the enzyme is a constituent of the myclin sheath itself.
THE HISTOCHEMICAL DEMONSTRATION OF HISTAMINE IN MAST CELLS
Authors: LAGUNOFF DAVID , PHILLIPS MICHAEL , BENDITT EARL P.
Published: April 2011

The use of formaldehyde vapor fixation following freeze-drying of tissues permits the histochemical demonstration of histamine in rat mast cells by several color producing reactions and fluorescence. Formaldehyde vapor fixed, frozendried mast cells develop an orange color on reaction with diazotized p-bromoaniline; the cells reduce both ferric ferricyanide and methenamine silver, and fluoresce yellow-green in ultraviolet light. These reactions are consistent with the presence of a tetrahydropyridoiminazole formed in rat mast cells by the reaction of the contained histamine with the applied formaldehyde.
STUDIES OF ACID PHOSPHATASE. I. ELECTROPHORETIC SEPARATION OF ACID PHOSPHATASES OF RAT LIVER ON POLYACRYLAMIDE GELS
Authors: BARKA TIBOR
Published: April 2011

The technique of Polyacrylamide gel electrophoresis for acid phosphatases is described. This permits the separation of the soluble acid phosphatases of rat liver into three fractions.
ON THE MECHANISM OF RESORCIN-FUCHSIN STAINING
Authors: PUCHTLER HOLDE , SWEAT FAYE , BATES R. , BROWN J. H.
Published: April 2011

Acetylation, sulfation and phosphorylation induce binding of resorcin-fuchsin by glycogen, basement membranes, reticulum fibers, collagen, and other tissue structures containing polysaccharides as demonstrated by the periodic acid Schiff reaction. These structures do not stain with resorcin-fuchsin in control section without pretreatment. It is concluded that the binding of resorcin-fuchsin is due to the introduction of ester groups.

Extraction procedures, designed to remove dyes held by salt-like or ionic linkage, indicate binding of resorcin-fuchsin by non-ionic bonds.

According to data on the dyeing of the polysaccharide ester cellulose acetate in textile dyeing, cellulose acetate dyes are adsorbed by hydrogen-bonding between the carbonyl oxygen of the ester group and phenolic hydroxyl groups of the dye. It seems possible that in resorcin-fuchsin a phenolic hydroxyl group of the resorcinol moiety of the dye is free to react.

The intense staining of agar, esterified glycogen, collagen and reticulum fibers with resorcin-fuchsin implies that binding of this dye does not convey any information concerning the protein moiety of tissue structures. Thus, resorcin-fuchsin can not be considered specific for elastic fibers in the chemical sense of the term.
THE ACTION OF HISTOLOGICAL FIXATIVES ON TISSUE LIPIDS. COMPARISON OF THE ACTION OF SEVERAL FIXATIVES USING PAPER CHROMATOGRAPHY
Authors: HESLINGA F. J. M. , DEIERKAUF F. A.
Published: April 2011

A simple procedure to obtain information on the influence of fixatives on tissue lipids is described. The action of several histological fixatives on tissue lipids was investigated by extraction of the lipids from fixed tissues followed by analysis of the extracts by paper chromatography. Primary fixatives as well as fixative mixtures were investigated.

Formaldehyde reacts very fast with phosphatidylethanolamine; other lipids are attacked gradually.

The greater part of the lipids become insoluble under influence of dichromate and chromic acid. The still extractable part consists mainly of oxidation products.

Osmium tetroxide immobilises most lipids in fresh and in formaldehyde fixed tissues.

Mercuric chloride is shown to react with several lipid components, possibly by formation of addition complexes with highly unsaturated fatty acid chains.

The fixative mixtures additively display the actions of the individual components; the chemically reactive components tend to dominate the results.
MICRODETERMINATION OF GLYCOGEN WITH ANTHRONE REAGENT
Authors: TEMPLETON MCCORMICK
Published: April 2011

With samples of embryonic skeletal muscle, 20-40 µg wet weight, it was possible to measure 0.08 µg of glycogen within 10% in a final volume of 98.4 µl. Commercial anthrone was washed, dissolved in absolute alcohol, filtered and recrystallized. Glycogen was also repurified. For the working reagent, 20 mg of anthrone was dissolved in 10 ml of 77% (v/v) sulfuric acid. A sample was dispersed in 10 µl water in a small test tube, 90 µl anthrone reagent was added, promptly mixed and the tube plugged with a polyethylene stopper. Triplicate 10 µl portions of water and of dilutions of glycogen were treated the same as the sample. All tubes were heated at 100°C for 15 minutes then cooled and optical density read at 630 mµ in a spectrophotometer. The extinction coefficient for a 1% glycogen solution was 490 ± 3% with 1 cm cell length. Recovery experiments were good. Proteins and pentoses did not interfere significantly. Methods for distinguishing between mono- and polysaccharides are discussed.
PHOSPHORYLASE AND AMYLO-1,6-TRANSGLUCOSIDASE IN RODENT ORAL MUCOUS MEMBRANE
Authors: KLINGSBERG JULES , BUTCHER EARL O.
Published: April 2011

Fresh frozen sections of rodent palate and gingiva were stained for phosphorylase and amylo-1,6-transglucosidase by the method of Takeuchi. Although phosphorylase has been reported to be lacking in rodent skin it appears to be present in rodent oral mucous membrane. The activity appears to be of a low intensity in this region since consistent staining was obtained only with thick sections (70 µ). Straight chain and branching enzymes were found to be more abundant in the palate than the gingiva, and the observed activity diminished with increasing age. Straight chain enzyme activity appeared abundant in the basal cells of the stratum Malpighii although stainable glycogen is usually lacking in these cells.
Regulatory effects of TLR2 on megakaryocytic cell function
Authors: Beaulieu Lea M. , Lin Elaine , Morin Kristine M. , Tanriverdi Kahraman , Freedman Jane E.
Published: April 2011

TLR2, a functional, inflammatory-related receptor, is known to be expressed on megakaryocytes and platelets and to lead to infection and immune-mediated activation of platelets; however, the role of this receptor in megakaryocytes is not understood. Using Meg-01 cells and mouse megakaryocytes, we found that NFκB, ERK-MAPK, and PI3K/Akt pathways, known downstream pathways of TLRs, are activated by Pam3CSK4, a TLR2-specific ligand. In addition, transcription factors associated with megakaryocyte maturation, GATA-1, NF-E2, and mammalian target of rapamycin (mTOR), are all increased in the presence of Pam3CSK4. The effect of Pam3CSK4 on megakaryocyte maturation was verified by the increase in DNA content and adhesion to extracellular matrix proteins by TLR2-dependent stimulation. In addition, TLR2 stimulation resulted in an increase in reactive oxygen species (ROS) production. Gene expression and protein levels of GP1b, CD41, MCP-1, COX2, NFκB1, and TLR2 were up-regulated in megakaryocytes after TLR2 stimulation through NFκB, PI3K/Akt, and ERK-MAPK pathways. Treatment of wild-type mice with Pam3CSK4 resulted in a return to normal platelet levels and an increase in megakaryocyte maturation, which did not occur in the TLR2−/− mice. Therefore, inflammation, through TLR2, can increase maturation and modulate the phenotype of megakaryocytes, contributing to the interrelationship between inflammation and hemostasis.
Modification of PCV-2 virulence by substitution of the genogroup motif of the capsid protein
Authors: Allemandou Aude , Grasland Béatrice , Hernandez-Nignol Anne-Cécile , Kéranflec'h André , Cariolet Roland , Jestin André
Published: April 2011

Porcine circovirus type 2 (PCV-2) is the causal agent of the post-weaning multisystemic wasting syndrome (PMWS). PCV-2 are small single-stranded circular DNA viruses clustered into two main genogroups: PCV-2a and PCV-2b. Each genogroup present a specific highly-conserved motif of six amino acids (between amino acids 86 and 91) in the PCV-2 capsid protein. The aim of this study was to verify whether the motif located in the capsid protein and specific to each PCV-2 genogroup contributes to virulence. Two parental DNA clones, PCV-2a and PCV-2b, were constructed as well as two mutants DNA clones, PCV-2a/motif 2b and PCV-2b/motif 2a by exchanging the capsid motif of each genogroup. The four DNA clones were characterized in vitro as well as in vivo. Cells transfected by the four DNA clones produced infectious viruses. In specific-pathogen-free piglets transfected by the four infectious DNA clones, PCV-2b/motif 2a virulence was not attenuated while the PCV-2a/motif 2b virulence was drastically reduced compared to their parent virulence. These results suggest that the amino acids between positions 86 and 91 of the capsid protein are determinant for the virulence of isolates. However, the environment of this motif seems also involved.
Glycoproteomic characterization of carriers of the CD15/Lewisx epitope on Hodgkin's Reed-Sternberg cells
Authors: Powlesland Alex S , Barrio Maria Marcela , Mordoh José , Hitchen Paul G , Dell Anne , Drickamer Kurt , Taylor Maureen E
Published: April 2011

Background

The Lewisx trisaccharide, also referred to as the CD15 antigen, is a diagnostic marker used to distinguish Hodgkin's lymphoma from other lymphocytic cancers. However, the role of such fucosylated structures remains poorly understood, in part because carriers of Lewisx structures on Hodgkin's Reed-Sternberg cells have not been identified.

Methods

GalMBP, an engineered carbohydrate-recognition protein that binds selectively to oligosaccharides with paired terminal galactose and fucose residues, has been used in conjunction with proteomic and glycomic analysis to identify glycoprotein carriers of Lewisx and related glycan structures in multiple Hodgkin's Reed-Sternberg cell lines.

Results

Multiple glycoproteins that bind to GalMBP and carry CD15/Lewisx have been identified in a panel of six Reed-Sternberg cell lines. The most commonly identified Lewisx-bearing glycoproteins are CD98hc, which was found in all six cell lines tested, and intercellular adhesion molecule-1 and DEC-205, which were detected in five and four of the lines, respectively. Thus, several of the most prominent cell adhesion molecules on the lymphomas carry this characteristic glycan epitope. In addition, the Hodgkin's Reed-Sternberg cell lines can be grouped into subsets based on the presence or absence of less common Lewisx-bearing glycoproteins.

Conclusions

CD98 and intercellular adhesion molecule-1 are major carriers of CD15/Lewisx on Reed-Sternberg cells. Binding of DC-SIGN and other glycan-specific receptors to the Lewisx epitopes on CD98 and intercellular adhesion molecule-1 may facilitate interaction of the lymphoma cells with lymphocytes and myeloid cells in lymph nodes.
Evolution of Parallel Spindles Like genes in plants and highlight of unique domain architecture#
Authors: Cigliano Riccardo Aiese , Sanseverino Walter , Cremona Gaetana , Consiglio Federica M , Conicella Clara
Published: April 2011

Background

Polyploidy has long been recognized as playing an important role in plant evolution. In flowering plants, the major route of polyploidization is suggested to be sexual through gametes with somatic chromosome number (2n). Parallel Spindle1 gene in Arabidopsis thaliana (AtPS1) was recently demonstrated to control spindle orientation in the 2nd division of meiosis and, when mutated, to induce 2n pollen. Interestingly, AtPS1 encodes a protein with a FHA domain and PINc domain putatively involved in RNA decay (i.e. Nonsense Mediated mRNA Decay). In potato, 2n pollen depending on parallel spindles was described long time ago but the responsible gene has never been isolated. The knowledge derived from AtPS1 as well as the availability of genome sequences makes it possible to isolate potato PSLike (PSL) and to highlight the evolution of PSL family in plants.

Results

Our work leading to the first characterization of PSLs in potato showed a greater PSL complexity in this species respect to Arabidopsis thaliana. Indeed, a genomic PSL locus and seven cDNAs affected by alternative splicing have been cloned. In addition, the occurrence of at least two other PSL loci in potato was suggested by the sequence comparison of alternatively spliced transcripts.

Phylogenetic analysis on 20 Viridaeplantae showed the wide distribution of PSLs throughout the species and the occurrence of multiple copies only in potato and soybean.

The analysis of PSLFHA and PSLPINc domains evidenced that, in terms of secondary structure, a major degree of variability occurred in PINc domain respect to FHA. In terms of specific active sites, both domains showed diversification among plant species that could be related to a functional diversification among PSL genes. In addition, some specific active sites were strongly conserved among plants as supported by sequence alignment and by evidence of negative selection evaluated as difference between non-synonymous and synonymous mutations.

Conclusions

In this study, we highlight the existence of PSLs throughout Viridaeplantae, from mosses to higher plants. We provide evidence that PSLs occur mostly as singleton in the analyzed genomes except in soybean and potato both characterized by a recent whole genome duplication event. In potato, we suggest the candidate PSL gene having a role in 2n pollen that should be deeply investigated.

We provide useful insight into evolutionary conservation of FHA and PINc domains throughout plant PSLs which suggest a fundamental role of these domains for PSL function.
Composite transcriptome assembly of RNA-seq data in a sheep model for delayed bone healing
Authors: Jäger Marten , Ott Claus-Eric , Grünhagen Johannes , Hecht Jochen , Schell Hanna , Mundlos Stefan , Duda Georg N , Robinson Peter N , Lienau Jasmin
Published: April 2011

Background

The sheep is an important model organism for many types of medically relevant research, but molecular genetic experiments in the sheep have been limited by the lack of knowledge about ovine gene sequences.

Results

Prior to our study, mRNA sequences for only 1,556 partial or complete ovine genes were publicly available. Therefore, we developed a composite de novo transcriptome assembly method for next-generation sequence data to combine known ovine mRNA and EST sequences, mRNA sequences from mouse and cow, and sequences assembled de novo from short read RNA-Seq data into a composite reference transcriptome, and identified transcripts from over 12 thousand previously undescribed ovine genes. Gene expression analysis based on these data revealed substantially different expression profiles in standard versus delayed bone healing in an ovine tibial osteotomy model. Hundreds of transcripts were differentially expressed between standard and delayed healing and between the time points of the standard and delayed healing groups. We used the sheep sequences to design quantitative RT-PCR assays with which we validated the differential expression of 26 genes that had been identified by RNA-seq analysis. A number of clusters of characteristic expression profiles could be identified, some of which showed striking differences between the standard and delayed healing groups. Gene Ontology (GO) analysis showed that the differentially expressed genes were enriched in terms including extracellular matrix, cartilage development, contractile fiber, and chemokine activity.

Conclusions

Our results provide a first atlas of gene expression profiles and differentially expressed genes in standard and delayed bone healing in a large-animal model and provide a number of clues as to the shifts in gene expression that underlie delayed bone healing. In the course of our study, we identified transcripts of 13,987 ovine genes, including 12,431 genes for which no sequence information was previously available. This information will provide a basis for future molecular research involving the sheep as a model organism.