scholarly journals Metabolic Mapping: Quantitative Enzyme Cytochemistry and Histochemistry to Determine the Activity of Dehydrogenases in Cells and Tissues

10.3791/56843 ◽  
2018 ◽  
Author(s):  
Remco J. Molenaar ◽  
Mohammed Khurshed ◽  
Vashendriya V.V. Hira ◽  
Cornelis J.F. Van Noorden
Keyword(s):  
Enzyme Cytochemistry ◽  
Metabolic Mapping ◽  
In Cells

A new cytochemical method: Combined enzyme cytochemistry and immunocytochemistry on ultrathin cryosections

1993 ◽  
Vol 51 ◽  
pp. 326-327
Author(s):  
Toshihiro Takizawa ◽  
John M. Robinson
Keyword(s):  
Cell Types ◽  
Uranyl Acetate ◽  
New Method ◽  
Human Neutrophils ◽  
Enzyme Cytochemistry ◽  
Cytochemical Method ◽  
Antigenic Sites ◽  
Cacodylate Buffer ◽  
Ultrathin Cryosections ◽  
In Cells

Cryo-ultramicrotomy has been a useful method for immunocytochemistry because it can preserve the ultrastructure of cells and also display antigenic sites in cells. Recently, cryo-ultramicrotomy has been successfully applied to enzyme cytochemistry for the demonstration of some phosphatases with lead as the capture metal. However, this application has some problems. The lead reaction product is unstable with uranyl acetate staining, thus leading to poor ultrastructural detail. The purpose of this study is to develop the new method of enzyme cytochemistry using cerium as the capture metal on ultrathin cryosections labeled with immunogold probes. This method should be of importance for demonstrating the ultrastructure localization of certain enzyme and the simultaneous localization of antigens in a variety of cell types.Freshly isolated human neutrophils were fixed with paraformaldehyde and/or glutaraldehyd in 0.1 M cacodylate buffer (pH 7.2) with 5 % sucrose, then pelleted into 10 % gelatin and subsequently infiltrated with 2.3 M sucrose.

Cytoplasmic Tubules in Cells Infected with Laryngotracheitis Virus

1969 ◽  
Vol 27 ◽  
pp. 376-377
Author(s):  
A. M. Watrach
Keyword(s):  
Tissue Culture ◽  
Small Proportion ◽  
Chicken Embryo ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Morphologic Characteristics ◽  
Infectious Laryngotracheitis ◽  
Laryngotracheitis Virus ◽  
In Cells

During a study of the development of infectious laryngotracheitis (LT) virus in tissue culture cells, unusual tubular formations were found in the cytoplasm of a small proportion of the affected cells. It is the purpose of this report to describe the morphologic characteristics of the tubules and to discuss their possible association with the development of virus.The source and maintenance of the strain of LT virus have been described. Prior to this study, the virus was passed several times in chicken embryo kidney (CEK) tissue culture cells.

Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

1971 ◽  
Vol 29 ◽  
pp. 548-549
Author(s):  
Awtar Krishan ◽  
Dora Hsu
Keyword(s):  
Granular Material ◽  
Rna Synthesis ◽  
Culture Medium ◽  
Actinomycin D ◽  
Metabolic Inhibitors ◽  
Protein And Rna Synthesis ◽  
Apparent Reduction ◽  
Plant Alkaloids ◽  
In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

Gastric Ulcers Produced by Cisplatin

1981 ◽  
Vol 39 ◽  
pp. 582-583
Author(s):  
S.K. Aggarwal ◽  
J. San Antonio
Keyword(s):  
Electron Microscopy ◽  
Single Dose ◽  
Side Effects ◽  
Antitumor Agent ◽  
Gastric Ulcers ◽  
Enzyme Cytochemistry ◽  
Intestinal Toxicity ◽  
Ovarian Cancers ◽  
Inner Surface

Cisplatin (cis-dichlorodiammineplatinum(II)) a potent antitumor agent is now available for the treatment of testicular and ovarian cancers. It is however, not free from its serious side effects including nephrotoxicity, gastro intestinal toxicity, myelosuppression, and ototoxicity. Here we now report that the drug produces peculiar bloating of the stomach in rats and induces acute ulceration.Wistar-derived rats weighing 200-250 g were administered cisplatin(9 mg/kg) ip as a single dose in 0.15 M NaCl. After 3 days the animals were sacrificed by decapitation. The stomachs were removed, the contents analyzed for pepsin and acidity. The inner surface was examined with a dissecting microscope after a moderate stretching for ulcers. Affected areas were fixed and processed for routine electron microscopy and enzyme cytochemistry.The drug treated animals kept on food and water consistently showed bloating and lesions (Fig. 1) with a frequency of 6-70 ulcers in the rumen section of the stomachs.

Detection of Nucleic Acids in Cells or Tissue Sections Using In situ Polymerase Chain Reaction (PCR)

1996 ◽  
Vol 54 ◽  
pp. 16-17
Author(s):  
J. R. Hully ◽  
K. R. Luehrsen ◽  
K. Aoyagi ◽  
C. Shoemaker ◽  
R. Abramson
Keyword(s):  
Nucleic Acids ◽  
Viral Infections ◽  
Anatomical Location ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Cellular Source ◽  
Gene Transcripts ◽  
Initial Description ◽  
In Cells

The development of PCR technology has greatly accelerated medical research at the genetic and molecular levels. Until recently, the inherent sensitivity of this technique has been limited to isolated preparations of nucleic acids which lack or at best have limited morphological information. With the obvious exception of cell lines, traditional PCR or reverse transcription-PCR (RT-PCR) cannot identify the cellular source of the amplified product. In contrast, in situ hybridization (ISH) by definition, defines the anatomical location of a gene and/or it’s product. However, this technique lacks the sensitivity of PCR and cannot routinely detect less than 10 to 20 copies per cell. Consequently, the localization of rare transcripts, latent viral infections, foreign or altered genes cannot be identified by this technique. In situ PCR or in situ RT-PCR is a combination of the two techniques, exploiting the sensitivity of PCR and the anatomical definition provided by ISH. Since it’s initial description considerable advances have been made in the application of in situ PCR, improvements in protocols, and the development of hardware dedicated to in situ PCR using conventional microscope slides. Our understanding of the importance of viral latency or viral burden in regards to HIV, HPV, and KSHV infections has benefited from this technique, enabling detection of single viral copies in cells or tissue otherwise thought to be normal. Clearly, this technique will be useful tool in pathobiology especially carcinogenesis, gene therapy and manipulations, the study of rare gene transcripts, and forensics.

Integrated morphometrical and electron energy loss image analysis in cytochemistry

1989 ◽  
Vol 47 ◽  
pp. 402-403
Author(s):  
W.C. de Bruijn ◽  
A.A.W. de Jong ◽  
C.W.J. Sorber
Keyword(s):  
Image Analysis ◽  
Acid Phosphatase ◽  
Chemical Analysis ◽  
Active Form ◽  
List Type ◽  
Type Number ◽  
Enzyme Cytochemistry ◽  
Related Data ◽  
Endogenous Peroxidase ◽  
Electron Energy Loss

One aspect of enzyme cytochemistry is, whether all macrophage lysosomal hydrolytical enzymes are present in an active form, or are activated upon stimulation. Integrated morphometrical and chemical analysis has been chosen as a tool to illucidate that cytochemical problem. Mouse peritoneal resident macrophages have been used as a model for this complicated integration of morphometrical and element-related data. Only aldehyde-fixed cells were treated with three cytochemical reactions to detect different enzyme activities within one cell (for details see [1,2]). The enzyme-related precipitates anticipated to be differentiated, were:(1).lysosomal barium and sulphur from aryl sulphatase activity,(2).lysosomal cerium and phosphate from acid phosphatase activity and(3).platinum/di-amino-benzidine( D A B) complex from endogenous peroxidase activity.

High time resolution enzyme cytochemistry of rod outer segment

1990 ◽  
Vol 48 (3) ◽  
pp. 878-879
Author(s):  
Takuma Saito ◽  
Toshihiro Takizawa
Keyword(s):  
Enzyme Activities ◽  
Outer Segment ◽  
Rapid Change ◽  
High Time Resolution ◽  
Visual Cell ◽  
Enzyme Cytochemistry ◽  
Activation Effect ◽  
Rod Outer Segment ◽  
Enzymatic Cascades ◽  
Rapid Drop

Cells and tissues live on a number of dynamic metabolic pathways, which are made up of sequential enzymatic cascades.Recent biochemical and physiological studies of vision research showed the importance of cGMP metabolism in the rod outer segment of visual cell, indicat ing that the photon activated rhodopsin exerts activation effect on the GTP binding protein, transducin, and this act ivated transducin further activates phosphodiesterase (PDEase) to result in a rapid drop in cGMP concentration in the cytoplasm of rod outer segment. This rapid drop of cGMP concentration exerts to close the ion channel on the plasma membrane and to stop of inward current brings hyperpolarization and evokes an action potential.These sequential change of enzyme activities, known as cGMP cascade, proceeds quite rapidly within msec order. Such a rapid change of enzyme activities, such as PDEase in rod outer segment, was not a matter of conventional histochemical invest igations.

Absence of temporal ordering of apoptotic features in heat-shock treated leukemia and lymphoma cell lines

1996 ◽  
Vol 54 ◽  
pp. 758-759
Author(s):  
D. W. Fairbain ◽  
M.D. Standing ◽  
K.L. O'Neill
Keyword(s):  
Heat Shock ◽  
Cell Lines ◽  
Chromatin Condensation ◽  
Membrane Integrity ◽  
Resistant Cell ◽  
Membrane Blebbing ◽  
Late Loss ◽  
Induced Apoptosis ◽  
Dying Cells ◽  
In Cells

Apoptosis is a genetically defined response to physiological stimuli that results in cellular suicide. Features common to apoptotic cells include chromatin condensation, oligonucleosomal DNA fragmentation, membrane blebbing, nuclear destruction, and late loss of ability to exclude vital dyes. These characteristics contrast markedly from pathological necrosis, in which membrane integrity loss is demonstrated early, and other features of apoptosis, which allow a non-inflammatory removal of dead and dying cells, are absent. Using heat shock-induced apoptosis as a model for examining stress response in cells, we undertook to categorize a variety of human leukemias and lymphomas with regard to their response to heat shock. We were also interested in determining whether a common temporal order was followed in cells dying by apoptosis. In addition, based on our previous results, we investigated whether increasing heat load resulted in increased apoptosis, with particular interest in relatively resistant cell lines, or whether the mode of death changed from apoptosis to necrosis.

Umatilla Virus Association with Fibrils and Tubules in Cultured Cells

1980 ◽  
Vol 38 ◽  
pp. 476-477
Author(s):  
Neil M. Foster ◽  
Ruth D. Breckon
Keyword(s):  
Bluetongue Virus ◽  
Linear Array ◽  
Cultured Cells ◽  
Intimate Association ◽  
Infected Cells ◽  
Dark Staining ◽  
In Cells

Macrotubules have been described1 in cells infected with Umatilla virus (UMAV), an orbivirus for which bluetongue virus (BTV) is the protype. Macrotubules, often in linear array, were observed in the cytoplasm and in intimate association with viroplasms of infected cells. Macrotubules had outside and inside diameters of 20 and 15 nm and many had dark-staining centers with diameters similar to the interiors of the tubules. UMAV was 60 nm and the RNA core was 30 nm in diameter. This report describes the association of UMAV with macrotubules and two types of microtubules.

Photo-tethered molecular beacons for superior light-induction

2021 ◽  
Author(s):  
Robin Klimek ◽  
Mantian Wang ◽  
Vivien R. McKenney ◽  
Erin M. Schuman ◽  
Alexander Heckel
Keyword(s):  
Molecular Beacons ◽  
Light Induction ◽  
In Cells

Photolabile circularization of molecular beacons via backbone phosphates leads to superior probes to study spatiotemporal aspects of RNA in cells.

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