scholarly journals CYTOCHEMISTRY OF CYTOPLASMIC AND INTRANUCLEAR INCLUSIONS INDUCED BY BOVINE PARAINFLUENZA 3 VIRUS (SF-4) IN HUMAN CELL CULTURES

1966 ◽  
Vol 14 (2) ◽  
pp. 187-195 ◽  
Author(s):  
FREDERICK H. KASTEN ◽  
A. E. CHURCHILL
Keyword(s):  
Picric Acid ◽  
Methyl Green ◽  
Bromphenol Blue ◽  
Intranuclear Inclusions ◽  
Fast Green ◽  
Metaphosphoric Acid ◽  
Basic Proteins ◽  
Azure B ◽  
Human Cell Cultures ◽  
Schiff Reaction

Infection of human IIEp-2 cells with myxovirus bovine parainfluenza 3 (SF-4) induces the formation of cytoplasmic and intranuclear inclusions. Both types of inclusions were shown to contain some RNA in late stages (gallocyanin chromalum, azure B bromide, pyronin, RNase), abundant protein at all stages (acidic fast green, mercuric bromphenol blue) primarily with lysine-rich residues (naphthol yellow S, lysine blockage) and lesser amounts of arginine (Sakaguchi) and tyrosine (diazotization coupling). Inclusions failed to stain for histones (alkaline fast green, ammoniacal silver), basic proteins (alkaline Biebrich scarlet, picric acid-eosin). DNA (Feulgen, fluorescent-Feulgen, methyl green, gallocyanin chromalum, azure B bromide) and polysaccharides (fluorescent-PAS, Hale). There was a slight staining for histones with metaphosphoric acid-gallocyanin and basic proteins with picric acid-bromphenol blue or -eosin and a negative reaction for proteins with the fluorescent modification of the N-Bromosuccinimide-Schiff reaction. Acridine orange fluorochroming revealed a false positive reaction for DNA (green fluorescence) which was due to protein.

scholarly journals CHANGES IN NUCLEAR HISTONES DURING FERTILIZATION, AND EARLY EMBRYONIC DEVELOPMENT IN THE PULMONATE SNAIL, Helix aspersa

1960 ◽  
Vol 8 (1) ◽  
pp. 69-81 ◽  
Author(s):  
David P. Bloch ◽  
Howard Y. C. Hew
Keyword(s):  
Developmental Stages ◽  
Picric Acid ◽  
Polar Body ◽  
Helix Aspersa ◽  
Bromphenol Blue ◽  
Fast Green ◽  
Specific Staining ◽  
Trichloracetic Acid ◽  
Basic Proteins ◽  
Snail Helix Aspersa

Calf thymus histories comprising two fractions, one rich in lysine, the other having roughly equal amounts of lysine and arginine, Loligo testes histones rich in arginine, and salmine, are compared with respect to their amino acid compositions, and their staining properties when the proteins are fixed on filter paper. The three types of basic proteins; somatic, arginine-rich spermatid histones, and protamine can be distinguished on the following basis. Somatic and testicular histones stain with fast green or bromphenol blue under the same conditions used for specific staining of histones in tissue preparations. The former histones lose most or all of their stainability after deamination or acetylation. Staining of the arginine-rich testicular histones remains relatively unaffected by this treatment. Protamines do not stain with fast green after treatment with hot trichloracetic acid, but are stained by bromphenol blue or eosin after treatment with picric acid. These methods provide a means for the characterization of nuclear basic proteins in situ. Their application to the early developmental stages of Helix aspersa show the following: After fertilization the protamine of the sperm is lost, and is replaced by faintly basic histones which differ from adult histones in their inability to bind fast green, and from protamines, by both their inability to bind eosin, and their weakly positive reaction with bromphenol blue. These "cleavage" histones are found in the male and female pronuclei, the early polar body chromosomes, and the nuclei of the cleaving egg and morula stages. During gastrulation, the histone complement reverts to a type as yet indistinguishable from that of adult somatic cells.

scholarly journals Basic Proteins of Plant Nuclei during Normal and Pathological Cell Growth

1959 ◽  
Vol 6 (2) ◽  
pp. 263-276 ◽  
Author(s):  
Ellen Rasch ◽  
John W. Woodard
Keyword(s):  
Animal Species ◽  
Broad Bean ◽  
Higher Plants ◽  
Feulgen Reaction ◽  
Fast Green ◽  
Basic Proteins ◽  
Normal Tissues ◽  
Tissue Sections ◽  
Tumor Tissues ◽  
Sperm Nuclei

Histone proteins were studied by microphotometry of plant tissue sections stained with fast green at pH 8.1. For comparative purposes the Feulgen reaction was used for deoxyribose nuclei acid (DNA); the Sakaguchi reaction for arginine; and the Millon reaction for estimates of total protein. Analysis of Tradescantia tissues indicated that amounts of nuclear histone fell into approximate multiples of the gametic (egg or sperm) quantity except in dividing tissues, where amounts intermediate between multiples were found. In differentiated tissues of lily, corn, onion, and broad bean, histones occurred in constant amounts per nucleus, characteristic of the species, as was found also for DNA. Unlike the condition in several animal species, the basic proteins of sperm nuclei in these higher plants were of the histone type; no evidence of protamine was found. In a plant neoplasm, crown gall of broad bean, behavior of the basic nuclear proteins closely paralleled that of DNA. Thus, alterations of DNA levels in tumor tissues were accompanied by quantitatively similar changes in histone levels to maintain the same Feulgen/fast green ratios found in homologous normal tissues.

scholarly journals ULTRASTRUCTURE AND CYTOCHEMISTRY OF METABOLIC DNA IN TIPULA

1966 ◽  
Vol 30 (1) ◽  
pp. 177-192 ◽  
Author(s):  
A. Lima-de-Faria ◽  
M. J. Moses
Keyword(s):  
Electron Microscope ◽  
Close Contact ◽  
Tritiated Thymidine ◽  
Polytene Chromosomes ◽  
The Body ◽  
Outer Shell ◽  
Synthetic Activity ◽  
Fast Green ◽  
Azure B ◽  
Basic Features

A DNA body is present in the females of the fly Tipula oleracea and is formed in contact with the sex chromosomes in the oogonial interphases. At each oogonial mitosis, the DNA body follows the chromosomes to one anaphase group and is included in one of the telophase nuclei. The body increases appreciably in size during the interphase of meiosis. All oocytes have the body, but only a few nurse cells possess it. The DNA body synthesizes its DNA at a different time than the chromosomes, as is shown by incorporation of tritiated thymidine, and contains 59% of the DNA of the nucleus, as is disclosed by spectrophotometric measurements. At late diplotene the DNA body disintegrates, releasing its DNA into either the nucleus or the cytoplasm. When studied in the electron microscope, the DNA body appears composed of a tight mass of intertwined fibrils. Demonstration that the main mass of the body is composed of DNA is obtained from cytochemical tests which reveal that the DNA body is Feulgen positive, stains green with azure B, incorporates H3-thymidine, and after digestion with DNase is Feulgen negative. The DNA of the body is complexed with histone, like the DNA of the chromosomes, as is revealed by an intense alkaline fast green staining. Electron microscope examination of oocytes reveals that one side of the DNA body is in close contact with the nuclear envelope and that the other side possesses an outer shell composed mainly of particles 150 to 250 A in diameter. Between the outer shell and the chromosomes there is a band of low electron opacity, 4000 to 7000 A thick. In the light microscope, this light band together with the outer shell is Feulgen negative and stains violet with azure B; this is confirmation of the presence of RNA. In the oocytes the nucleoli are found inside the DNA body. These nucleoli have a nucleolonema composed mainly of particles 150 to 250 A. The nucleoli are Feulgen negative, alkaline fast green negative, stain violet with azure B, and do not stain with azure B after RNase digestion, thus confirming their RNA content. The presence of the nucleoli inside the DNA body and of a band of RNA between the body and the chromosomes is indicative of a high RNA synthetic activity. Since the DNA of the body is complexed with histone, as in the chromosomes, and the nucleoli are located inside the body, the simplest interpretation of the DNA body is that it represents hundreds of copies of the operons of the nucleolar organizing region or neighboring regions. The situation found in Tipula has several basic features in common with the polytene chromosomes of other Diptera and with the hundreds of nucleoli present in Triturus oocytes. In all three cases, genes seem to be copied hundreds of times but are kept in different types of packages. A DNA body like the one in Tipula oleracea is found in other species of Diptera and in the Coleoptera. There is no indication, from the present investigation, that the DNA body is in any way associated with a virus.

scholarly journals A CYTOCHEMICAL STUDY OF CYTOPLASMIC BASIC PROTEINS IN THE ASCIDIAN OOCYTE

1965 ◽  
Vol 25 (2) ◽  
pp. 319-326 ◽  
Author(s):  
Richard Davenport ◽  
Janice C. Davenport
Keyword(s):  
Positive Charge ◽  
Amino Groups ◽  
Acid Dyes ◽  
Fast Green ◽  
Cytochemical Study ◽  
Basic Proteins ◽  
Protein Molecules ◽  
Nucleoprotein Complex ◽  
High Concentrations ◽  
Net Positive Charge

The cytoplasm of young oocytes of the ascidians contains high concentrations of proteins which are stainable with alkaline fast green at pH 8.1 and above. These proteins cannot be stained even with acid dyes at low pH unless RNA is removed. Deamination and formalin blockage of amino groups is incapable of destroying the net positive charge on these protein molecules in the presence of RNA, but these treatments destroy the charge if RNA is removed. It is therefore concluded that basic proteins and RNA exist as a nucleoprotein complex in the ribosomes of these young oocytes. The detectable RNA of the mature oocytes and unfertilized eggs shows no evidence of being associated with basic proteins.

scholarly journals STRUCTURE OF SOME STRIGEATAS' ORGAN OF BRANDES

2019 ◽  
pp. 399-403
Author(s):  
Nacheva ◽  
Manikovskaya
Keyword(s):  
Periodic Acid ◽  
Secretory Activity ◽  
Bromphenol Blue ◽  
Main Function ◽  
Periodic Acid Schiff ◽  
Alaria Alata ◽  
Sudan Black B ◽  
Glandular Cells ◽  
Secondary Function ◽  
Schiff Reaction

The purpose of the research: the analysis of histology and histochemistry of the Brandes organ of Strigea strigis and Alaria alata. The trematode marites of Strigea striges and Alaria alata were the material. Fixation of the material was carried out in 10% neutral formalin. The treatment of the specimens was carried out using the conventional histological and histochemical methods: with hematoxylin-eosin according to the method of Van Gizon, Mallory, with Sudan black B, sulema-Bromphenol blue according to Bonhage, with periodic acid Schiff reaction by Mac-Manus, alcian blue according to Stedman and Mowry and with toluidine blue. Histochemical reactions were performed with appropriate controls. The studies have shown that the structure of the Brandes organ of Strigea strigis and Alaria alata differs by their constituents and morphology of glandular cells. The histochemical reactions are similar. The cells of the glandular complex show bromophenolophilia, toluudinophilia and fuchsinophilia in periodic acid Schiff reaction speaks about the glycoprotein nature of the secreted substances. Bromphenophilia and sudanophilia of glandular cells cytoplasm indicate the presence of lipoprotein substances in them. The Brandes of S. strigis and A. alata is a morphofunctional unit, to which the principle of multi-functionality is inherent. It performs the main function – digestion of food components by means of developed glandular structures and specialized secretory activity. Its ability to fix the helminth tightly in the endostatin can be considered a secondary function of the organ.

scholarly journals THE NUCLEOLI IN MITOTIC DIVISIONS OF MAMMALIAN CELLS IN VITRO

1965 ◽  
Vol 26 (2) ◽  
pp. 539-553 ◽  
Author(s):  
T. C. Hsu ◽  
Frances E. Arrighi ◽  
Robert R. Klevecz ◽  
B. R. Brinkley
Keyword(s):  
Electron Microscopy ◽  
Protein Synthesis ◽  
Mammalian Cells ◽  
Fluorescent Microscopy ◽  
Chinese Hamster ◽  
Methyl Green ◽  
Azure B ◽  
Nucleolar Material ◽  
Cell Strains

In a number of mammalian cell strains nucleoli persisted through mitosis. This phenomenon was especially pronounced in several cell lines derived from Chinese hamster tissues. All the methods employed, including radioautography with tritiated uridine, cytochemical stains (methyl green-pyronin and azure B), fluorescent microscopy (coriphosphine O), ribonuclease digestion, and electron microscopy, demonstrated that the bodies identified as persistent nucleoli in the mitotic stages had the same characteristics as did the nucleoli in the interphase. Persistent nucleoli may attach to the chromosomes or may be free in the cytoplasm. In cells where no persistent nucleoli as such were noted, nucleolar material was observed to attach to the chromosomes in shapeless masses which moved with the chromosomes during anaphase. At least a portion of the nucleolar material was included in the daughter nuclei, presumably for immediate use for protein synthesis after cell division.

scholarly journals EVIDENCE OF DIFFERENCES IN THE DESOXYRIBONUCLEOPROTEIN COMPLEX OF RAPIDLY PROLIFERATING AND NON-DIVIDING CELLS

1955 ◽  
Vol 1 (6) ◽  
pp. 531-550 ◽  
Author(s):  
David P. Bloch ◽  
Gabriel C. Godman
Keyword(s):  
Methyl Green ◽  
Feulgen Staining ◽  
Fast Green ◽  
Residual Protein ◽  
Interphase Cells ◽  
Dividing Cells ◽  
The Difference ◽  
Cytophotometric Analysis ◽  
Two Stages ◽  
Increased Susceptibility

1. Quantitative cytophotometric analysis of the interphase cells of a rapidly proliferating differentiated tissue such as liver of new born rat, indicates that these cells can be separated into two groups on the basis of their staining characteristics after methanol fixation. 2. These groups are thought to correspond to two stages of interphase. The first, called "autosynthetic interphase," comprises cells which are duplicating chromosomal material in preparation for mitosis, and shows parallel increases in the methyl green and Feulgen staining of DNA and the fast green staining of histone from the diploid (2 C) to double these values (4 C). 3. The second group is designated the "heterosynthetic interphase," during which the cell ceases proliferating and functions in a manner commensurate with its state of differentiation. In this stage Feulgen staining indicates the diploid chromosomal complement, but there is a decreased capacity of the DNA to bind methyl green and of the histone to bind fast green. 4. The difference between the methyl green binding of the heterosynthetic and autosynthetic 2 C cells is due to the presence of a protein in the former which presumably inhibits staining by competing with the dye for binding sites on the DNA. The effect of this inhibition can be removed by extracting the protein, or by blocking the protein basic groups. 5. The decreased fast green staining of histone in the heterosynthetic cells can be minimized by prolonged fixation with formaldehyde. It is thought to stem either from a similar type of inhibition, or from an increased susceptibility of the histone to loss from the cell during this stage. 6. While histone inhibits methyl green staining of DNA in all cells, the differences between the staining properties of the autosynthetic and heterosynthetic interphase cells are believed to be due to another protein, whose properties appear similar to those of the chromosomal "residual protein." It is concluded that a complex of DNA and residual protein existing during the heterosynthetic interphase is dissociated during the autosynthetic interphase.

scholarly journals REPOPULATION OF THE POSTMITOTIC NUCLEOLUS BY PREFORMED RNA

1972 ◽  
Vol 53 (3) ◽  
pp. 611-623 ◽  
Author(s):  
Stephanie Gordon Phillips
Keyword(s):  
Actinomycin D ◽  
Chinese Hamster ◽  
Mitotic Cell ◽  
Fast Green ◽  
Azure B ◽  
Mouse Cells ◽  
Nucleolar Rna ◽  
L929 Mouse ◽  
Nucleolar Components ◽  
Mitotic Cells

This study is concerned with the fate of the nucleolar contents, particularly nucleolar RNA, during mitosis Mitotic cells harvested from monolayer cultures of Chinese hamster embryonal cells, KB6 (human) cells, or L929 (mouse) cells were allowed to proceed into interphase in the presence or absence (control) of 0.04–0 08 µg/ml of actinomycin D, a concentration which preferentially inhibits nucleolar (ribosomal) RNA synthesis 3 hr after mitosis, control cells had large, irregularly shaped nucleoli which stained intensely for RNA with azure B and for protein with fast green. In cells which had returned to interphase in the presence of actinomycin D, nucleoli were segregated into two components easily resolvable in the light microscope, and one of these components stained intensely for RNA with azure B. Both nucleolar components stained for protein with fast green In parallel experiments, cultures were incubated with 0.04–0 08 µg/ml actinomycin D for 3 hr before harvesting of mitotic cells, then mitotic cells were washed and allowed to return to interphase in the absence of actinomycin D. 3 hr after mitosis, nuclei of such cells were devoid of large RNA-containing structures, though small, refractile nucleolus-like bodies were observed by phase-contrast microscopy or in material stained for total protein. These experiments indicate that nucleolar RNA made several hours before mitosis persists in the mitotic cell and repopulates nucleoli when they reform after mitosis

scholarly journals CHANGING NUCLEAR HISTONE PATTERNS DURING DEVELOPMENT I. FERTILIZATION AND EARLY CLEAVAGE IN THE CRAB, EMERITA ANALOGA

1968 ◽  
Vol 16 (7) ◽  
pp. 473-479 ◽  
Author(s):  
JACK C. VAUGHN
Keyword(s):  
Developmental Stages ◽  
Sperm Nucleus ◽  
Bromphenol Blue ◽  
Early Cleavage ◽  
Fast Green ◽  
Adult Type ◽  
Emerita Analoga ◽  
Cleavage Stages ◽  
Nuclear Histone ◽  
Early Developmental

Application of cytochemical techniques to the early developmental stages of the decapod crab, Emerita analoga, shows the following. After fertilization, the sperm nucleus, which apparently contains no basic proteins prior to this stage, becomes associated with a class of weakly basic histones, which differ from adult type histones in their apparent inability to bind alkaline fast green and from protamines by their inability to bind bromphenol blue following acetylation. This class of histones only persists until the late pronucleus stage, by which time the chromosomes contain a class of histones that are indistinguishable from adult histones by qualitative cytochemical techniques. No further changes in the nuclear histones are detected in the zygote or early cleavage stages. These changing histone patterns during early crab development are discussed with reference to other similar studies in other organisms.

The indicator system of the Sabin-Feldman phenomenon in Toxoplasma

Parasitology ◽  
1960 ◽  
Vol 50 (1-2) ◽  
pp. 13-20
Author(s):  
C. Kulasiri
Keyword(s):  
Toluidine Blue ◽  
Nile Blue ◽  
Neutral Red ◽  
Indicator System ◽  
Alkaline Solutions ◽  
Methyl Green ◽  
Celestine Blue ◽  
Cresyl Blue ◽  
Azure B ◽  
Saline Solutions

Azure A, azure B and Nile blue in both neutral and alkaline solutions were found to be suitable for use as indicator dyes in the Sabin-Feldman test. Toluidine blue, brilliant cresyl blue and thionine were also suitable as neutral solutions. Azure C, phloxine, phloxine B, neutral red, methyl green, pyronine Y, eosin and celestine blue were found to be unsuitable. Azure B and toluidine blue in saline solutions were serviceable up to 5 months after preparation. The significance of these findings is discussed.

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