scholarly journals Effect of Postmortem Degradation on the Preservation of Viral Particles and Rabies Antigens in Mice Brains. Light and Electron Microscopic Study

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 938
Author(s):  
Jeison Monroy-Gómez ◽  
Gerardo Santamaría ◽  
Ladys Sarmiento ◽  
Orlando Torres-Fernández
Keyword(s):  
Electron Microscopy ◽  
Rabies Virus ◽  
Immunoelectron Microscopy ◽  
Degradation Process ◽  
Control Group ◽  
Direct Immunofluorescence ◽  
Electron Microscopic ◽  
Viral Antigens ◽  
Viral Particles ◽  
Rabies Diagnosis

Rabies diagnosis is mainly made on fresh brain tissue postmortem by means of the direct immunofluorescence test. However, in some cases, it is not possible to use this technique, given that the affected nervous tissue goes through a postmortem degradation process, due to problems in the handling and transport of the samples. For this reason, the preservation in time of the rabies virus inclusions was assessed, as well as the immunoreactivity and the ultrastructure of viral particles in tissue with postmortem degradation. Brains of mice inoculated with rabies virus and control mice were processed for conventional histology, immunohistochemistry, electron microscopy, and immunoelectron microscopy in different postmortem times. In the processed tissues for hematoxylin and eosin, the presence of eosinophilic inclusions was not observed beyond 12 h postmortem. Surprisingly, the immunoreactivity of the viral antigens increased with time, at least until 30 h postmortem. It was possible to easily recognize the viral particles by means of conventional electron microscopy until 12 h postmortem. Immunoelectron microscopy allowed us to identify the presence of viral antigens disseminated in the neuronal cytoplasm until 30 h postmortem, but immunoreactive viral particles were not observed. The rabies infection did not cause histological or ultrastructural alterations different from those in the control group as a result of the postmortem degradation. In conclusion, the immunohistochemistry is a reliable test for rabies diagnosis in samples with postmortem degradation and that have been fixed with aldehydes.

scholarly journals Identification of Tomato Infecting Viruses That Co-Isolate with Nanovesicles Using a Combined Proteomics and Electron-Microscopic Approach

Nanomaterials ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1922
Author(s):  
Ramila Mammadova ◽  
Immacolata Fiume ◽  
Ramesh Bokka ◽  
Veronika Kralj-Iglič ◽  
Darja Božič ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Mosaic Virus ◽  
Protein Identification ◽  
Plant Viruses ◽  
Size Exclusion ◽  
Tomato Mosaic Virus ◽  
High Yield ◽  
Plant Resources ◽  
Electron Microscopic ◽  
Viral Particles

Plant-derived nanovesicles (NVs) have attracted interest due to their anti-inflammatory, anticancer and antioxidative properties and their efficient uptake by human intestinal epithelial cells. Previously we showed that tomato (Solanum lycopersicum L.) fruit is one of the interesting plant resources from which NVs can be obtained at a high yield. In the course of the isolation of NVs from different batches of tomatoes, using the established differential ultracentrifugation or size-exclusion chromatography methods, we occasionally observed the co-isolation of viral particles. Density gradient ultracentrifugation (gUC), using sucrose or iodixanol gradient materials, turned out to be efficient in the separation of NVs from the viral particles. We applied cryogenic transmission electron microscopy (cryo-TEM), scanning electron microscopy (SEM) for the morphological assessment and LC–MS/MS-based proteomics for the protein identification of the gradient fractions. Cryo-TEM showed that a low-density gUC fraction was enriched in membrane-enclosed NVs, while the high-density fractions were rich in rod-shaped objects. Mass spectrometry–based proteomic analysis identified capsid proteins of tomato brown rugose fruit virus, tomato mosaic virus and tomato mottle mosaic virus. In another batch of tomatoes, we isolated tomato spotted wilt virus, potato virus Y and southern tomato virus in the vesicle sample. Our results show the frequent co-isolation of plant viruses with NVs and the utility of the combination of cryo-TEM, SEM and proteomics in the detection of possible viral contamination.

scholarly journals PREPARATION AND USE OF SOLUBLE FERRITIN-ANTIFERRITIN COMPLEXES AS A SPECIFIC MARKER FOR IMMUNOELECTRON MICROSCOPY

1974 ◽  
Vol 22 (1) ◽  
pp. 35-39 ◽  
Author(s):  
AMERICO A. MARUCCI ◽  
ROBERT M. DOUGHERTY ◽  
HENRY S. DISTEFANO
Keyword(s):  
Electron Microscopy ◽  
Tissue Culture ◽  
Immunoelectron Microscopy ◽  
Immune Complexes ◽  
Avian Leukosis Virus ◽  
Specific Marker ◽  
Viral Antigens ◽  
Soluble Immune Complexes

Soluble immune complexes of ferritin and chicken antiferritin have been prepared. This has been used with unlabeled chicken antibody to avian leukosis virus and rabbit antichicken globulin to demonstrate the presence of specific viral antigens in tissue culture by electron microscopy.

Fluorescent and Immuno-Electron Microscopy of Detergent Extracted Cytoskeletal and Associated Antigens

1985 ◽  
Vol 43 ◽  
pp. 582-583
Author(s):  
E.J. Basgall ◽  
M.M. Soong ◽  
W.A.F. Tompkins
Keyword(s):  
Electron Microscopy ◽  
Cultured Cells ◽  
Leukemia Virus ◽  
Protein A ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Simple Method ◽  
Viral Antigens ◽  
Associated Proteins ◽  
Nih 3T3

Triton X-100 detergent extraction has been shown to be a relatively simple method for revealing the internal architecture of cultured cells. This method has proven valuable in studies of cytoskeleton-associated proteins and their functions in regulating cell activities. Exposed cytoskeletal elements can be examined using either transmission or high-resolution scanning electron microscopy. Immunochemical studies at the electron microscopic level have been simplified by the availability of Protein A-colloidal gold as a high affinity marker for immunoglobulins, specifically IgG.Several investigators have reported an association between the cytoskeleton and viral antigens. Evidence has indicated that, in some systems, the cytoskeleton plays a significant role in virus infection. Immunofluorescent studies on canine distemper virus infected, extracted cells have revealed altered cytoskeletal staining patterns, as compared to non-infected controls. In our laboratory, immuno-electronmicroscopy studies of extracted NIH/3T3 cells infected with Maloney-murine leukemia virus have indicated an association between cytoskeletal actin and the viral antigens gp70 and pl5E.

Electron Microscopic Demonstration of Adenovirus in Appendix Vermiformis in a Case of Ileocecal Intussusception

PEDIATRICS ◽  
1973 ◽  
Vol 51 (3) ◽  
pp. 566-570
Author(s):  
Eduardo J. Yunis ◽  
Yoshie Hashida
Keyword(s):  
Electron Microscopy ◽  
Electron Microscopic ◽  
Simple Method ◽  
Appendix Vermiformis ◽  
Viral Particles ◽  
History Of ◽  
Ileocecal Intussusception ◽  
Electron Microscopic Demonstration ◽  
Hematoxylin Eosin ◽  
Selection Of

Viral particles identical to those of adenovirus were demonstrated in the intranuclear inclusion bodies of the epithelial cells of an appendix vermiformis removed from a boy with a recent history of intussusception. Viral particles were demonstrated by electron microscopy following reprocessing of a formaldehyde-fixed, paraffin embedded and hematoxylin-eosin stained section. This simple method permits precise selection of the tissue for electron microscopy and in instances such as this gives sufficient tissue preservation to demonstrate viral particles. Our findings support the idea that some cases of intussusception may be the result of the adenovirus infection.

scholarly journals Diagnosis and Analysis of a Recent Case of Human Rabies in Canada

2002 ◽  
Vol 13 (2) ◽  
pp. 129-133 ◽  
Author(s):  
Lindsay D Elmgren ◽  
Susan A Nadin-Davis ◽  
Frances T Muldoon ◽  
Alexander I Wandeler
Keyword(s):  
Polymerase Chain Reaction ◽  
Rabies Virus ◽  
False Negative ◽  
Strain Identification ◽  
Direct Immunofluorescence ◽  
Product Analysis ◽  
Chain Reaction ◽  
Food Inspection ◽  
Polymerase Chain ◽  
Rabies Diagnosis

BACKGROUND: On September 30, 2000, staff at the Canadian Food Inspection Agency's Centre of Expertise for Rabies, located at the Animal Diseases Research Institute in Ottawa, Ontario, diagnosed rabies in a child from Quebec. This was the first case of rabies in a human in Canada in 15 years and in 36 years in the province of Quebec. After spending a week in intensive care in a Montreal hospital, the nine-year-old boy succumbed to this nearly always fatal disease. The boy had been exposed to a bat in late August 2000, while vacationing with his family in the Quebec countryside.METHODS: Antemortem specimens taken from the patient were sent to the Canadian Food Inspection Agency laboratory for rabies diagnosis. Samples included saliva, eye secretions, corneal impressions, cerebral spinal fluid and skin. Specimens were examined by direct immunofluorescence microscopy, and results were confirmed using molecular biological techniques. Virus strain identification was performed by both genetic methods and phenotypic analysis with monoclonal antibodies.RESULTS: Initial results from direct immunofluorescence staining indicated that rabies virus was present in the skin biopsy specimen but not in the corneal impressions. This diagnosis of rabies was confirmed by polymerase chain reaction product analysis from several of the submitted specimens. Virus isolation from postmortem samples was not possible because fresh brain tissue was not available. Rabies virus was isolated from saliva and was determined to be similar to a variant that circulates in populations of silver-haired bats.INTERPRETATION: Intravitam diagnosis of rabies in humans is very dependent on the samples submitted for diagnosis and the method used for testing. Upon first examination, only skin specimens were positive for rabies virus antigen; using polymerase chain reaction analysis, only saliva yielded positive results for rabies virus genome. Extensive testing and retesting of specimens submitted for rabies diagnosis in humans must be done to avoid false negative results.

Localization of immunolabels by electron microscopy and image averaging

1983 ◽  
Vol 41 ◽  
pp. 282-283
Author(s):  
J. Frank ◽  
P.-Y. Sizaret ◽  
A. Verschoor ◽  
J. Lamy
Keyword(s):  
Electron Microscopy ◽  
Single Particle ◽  
Attachment Site ◽  
Uranyl Acetate ◽  
Limulus Polyphemus ◽  
Resolution Limit ◽  
Electron Microscopic ◽  
Image Averaging ◽  
Top View ◽  
Better Than

The accuracy with which the attachment site of immunolabels bound to macromolecules may be localized in electron microscopic images can be considerably improved by using single particle averaging. The example studied in this work showed that the accuracy may be better than the resolution limit imposed by negative staining (∽2nm).The structure used for this demonstration was a halfmolecule of Limulus polyphemus (LP) hemocyanin, consisting of 24 subunits grouped into four hexamers. The top view of this structure was previously studied by image averaging and correspondence analysis. It was found to vary according to the flip or flop position of the molecule, and to the stain imbalance between diagonally opposed hexamers (“rocking effect”). These findings have recently been incorporated into a model of the full 8 × 6 molecule.LP hemocyanin contains eight different polypeptides, and antibodies specific for one, LP II, were used. Uranyl acetate was used as stain. A total of 58 molecule images (29 unlabelled, 29 labelled with antl-LPII Fab) showing the top view were digitized in the microdensitometer with a sampling distance of 50μ corresponding to 6.25nm.

Immunocytochemistry. A Review of Methodology for Electron Microscopic Applicaiton

1974 ◽  
Vol 32 ◽  
pp. 328-329
Author(s):  
Joseph E. Mazurkiewicz
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Ultrastructural Level ◽  
Electron Microscopic ◽  
Biological Interest ◽  
Investigative Approach ◽  
Immunologic Activity ◽  
Dense Label

Immunocytochemistry is a powerful investigative approach in which one of the most exacting examples of specificity, that of the reaction of an antibody with its antigen, isused to localize tissue and cell specific molecules in situ. Following the introduction of fluorescent labeled antibodies in T950, a large number of molecules of biological interest had been studied with light microscopy, especially antigens involved in the pathogenesis of some diseases. However, with advances in electron microscopy, newer methods were needed which could reveal these reactions at the ultrastructural level. An electron dense label that could be coupled to an antibody without the loss of immunologic activity was desired.

Ultrastructural investigation of spontaneous pituitary gonadotroph cell adenomas in aging Sprague-Dawley rats

1983 ◽  
Vol 41 ◽  
pp. 702-703
Author(s):  
D. J. McComb ◽  
J. Beri ◽  
F. Zak ◽  
K. Kovacs
Keyword(s):  
Electron Microscopy ◽  
Animal Model ◽  
Epoxy Resin ◽  
Immunoelectron Microscopy ◽  
Tumor Type ◽  
Gonadal Function ◽  
Sprague Dawley ◽  
Male And Female ◽  
Reticulin Fibers ◽  
Sprague Dawley Rats

Gonadotroph cell adenomas of the pituitary are infrequent in human patients and are not invariably associated with altered gonadal function. To date, no animal model of this tumor type exists. Herein, we describe spontaneous gonadotroph cell adenomas in old male and female Sprague-Dawley rats by histology, immunocytology and electron microscopy.The material consisted of the pituitaries of 27 male and 38 female Sprague Dawley rats, all 26 months of age or older, removed at routine autopsy. Sections of formal in-fixed, paraffin-embedded tissue were stained with hematoxylin-phloxine-saffron (HPS), the PAS method and the Gordon-Sweet technique for the demonstration of reticulin fibers. For immunostaining, sections were exposed to anti-rat β-LH, anti-ratβ-TSH, anti-rat PRL, anti-rat GH and anti-rat ACTH 1-39. For electron microscopy, tissue was fixed in 2.5% glutaraldehyde, postfixed in 1% OsO4 and embedded in epoxy-resin. Tissue fixed in 10% formalin, embedded in epoxy resin without osmification, was used for immunoelectron microscopy.

An Electron Microscopic Study of an Avian Reovirus that Causes Arthritis

1971 ◽  
Vol 29 ◽  
pp. 254-255
Author(s):  
E, R. Walker ◽  
N. O. Olson ◽  
M. H. Friedman
Keyword(s):  
Electron Microscopy ◽  
Viral Genome ◽  
Inner Core ◽  
Avian Reovirus ◽  
Electron Microscopic ◽  
Outer Coat ◽  
Acid Type ◽  
Chicken Eggs ◽  
Icosahedral Particle ◽  
Mature Virus

An unidentified virus, responsible for an arthritic-like condition in chickens was studied by electron microscopy and other methods of viral investigation. It was characterized in chorio-allantoic membrane (CAM) lesions of embryonating chicken eggs and in tissue culture as to: 1) particle size; 2) structure; 3) mode of replication in the cell; and 4) nucleic acid type.The inoculated virus, coated and uncoated, is first seen in lysosomal-like inclusions near the nucleus; the virions appear to be uncoated in these electron dense inclusions (Figure 1), Although transfer of the viral genome from these inclusions is not observable, replicating virus and mature virus crystals are seen in the cytoplasm subsequent to the uncoating of the virions.The crystals are formed in association with a mass of fibrils 50 to 80 angstroms in diameter and a ribosome-studded structure that appears to be granular endoplasmic reticulum adapted to virus replication (Figure 2). The mature virion (Figure 3) is an icosahedral particle approximately 75 millimicrons in diameter. The inner core is 45 millimicrons, the outer coat 15 millimicrons, and the virion has no envelope.

A Comparative Study of Fractographic Replicas

1974 ◽  
Vol 32 ◽  
pp. 566-567
Author(s):  
Loren Anderson ◽  
Pat Pizzo ◽  
Glen Haydon
Keyword(s):  
Electron Microscopy ◽  
Electron Microscopic Examination ◽  
Direct Examination ◽  
Electron Microscopic ◽  
Reliable Technique ◽  
Service Failures ◽  
Transmission Electron ◽  
Mode Of Failure ◽  
Scanning Electron Microscopic ◽  
Scanning Electron

Transmission electron microscopy of replicas has long been used to study the fracture surfaces of components which fail in service. Recently, the scanning electron microscope (SEM) has gained popularity because it allows direct examination of the fracture surface. However, the somewhat lower resolution of the SEM coupled with a restriction on the sample size has served to limit the use of this instrument in investigating in-service failures. It is the intent of this paper to show that scanning electron microscopic examination of conventional negative replicas can be a convenient and reliable technique for determining mode of failure.

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