Localization of Classical Swine Fever Virus from Chronically Infected Pigs by In Situ Hybridization and Immunohistochemistry

2003 ◽  
Vol 40 (1) ◽  
pp. 107-113 ◽  
Author(s):  
C. Choi ◽  
C. Chae
Keyword(s):  
In Situ Hybridization ◽  
Nucleic Acid ◽  
Mononuclear Cells ◽  
Classical Swine Fever ◽  
Lymphoid Tissues ◽  
Hybridization Technique ◽  
Tissue Sections ◽  
Background Staining ◽  
The Central Nervous System

Classical swine fever (CSF) virus (CSFV) nucleic acid and antigen were detected in 15 pigs with naturally occurring chronic CSF by in situ hybridization and immunohistochemistry. The most consistent and prominent microscopic lesions were perivascular mononuclear cell infiltration and gliosis in the central nervous system of pigs with chronic CSF. Positive cells typically exhibited a dark brown (in situ hybridization) or red (immunohistochemistry) reaction product in the cytoplasm without background staining. A positive signal for both in situ hybridization and immunohistochemistry was detected in mononuclear cells and lymphocytes of lymphoid tissues. Viral nucleic acid was detected in some tissue sections in the absence of viral antigen. The in situ hybridization technique developed in this study was useful for the detection of CSFV RNA in tissues taken from chronically infected pigs and may be a valuable technique for studying the pathogenesis of chronic CSFV infection.

scholarly journals Using a Multiplex Nucleic Acid in situ Hybridization Technique to Determine HCN4 mRNA Expression in the Adult Rodent Brain

2019 ◽  
Vol 12 ◽  
Author(s):  
Julia Oyrer ◽  
Lauren E. Bleakley ◽  
Kay L. Richards ◽  
Snezana Maljevic ◽  
A. Marie Phillips ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Nucleic Acid ◽  
Mrna Expression ◽  
Hybridization Technique ◽  
Rodent Brain

scholarly journals Distribution of Bluetongue Virus in Tissues of Experimentally Infected Pregnant Dogs as Determined by In Situ Hybridization

1996 ◽  
Vol 33 (3) ◽  
pp. 337-340 ◽  
Author(s):  
C. C. Brown ◽  
J. C. Rhyan ◽  
M. J. Grubman ◽  
L. A. Wilbur
Keyword(s):  
In Situ Hybridization ◽  
Nucleic Acid ◽  
Bluetongue Virus ◽  
Mononuclear Cells ◽  
Nonstructural Protein ◽  
The Other ◽  
Post Inoculation ◽  
Viral Nucleic Acid ◽  
Nonstructural Protein 1

Six female dogs (four pregnant and two nonpregnant) were inoculated with bluetongue virus (BTV), serotype 11. Pregnant animals and one nonpregnant dog received 5.5-6.3 log10 of cell culture-adapted virus. The other nonpregnant dog received a modified live vaccine contaminated with bluetongue virus. The nonpregnant animals never became clinically ill and were euthanatized 35 days post-inoculation. Three of the four pregnant dogs aborted, and all four died or were euthanatized 5-10 days post-inoculation. The predominant pathologic feature in the adults was severe pulmonary edema. Various tissues from the bitches and fetuses were examined by in situ hybridization using a digoxigenin-labeled probe corresponding to the nonstructural protein-1 gene of BTV-17. By this technique, viral nucleic acid was detected predominantly in endothelial cells of lung of all four dogs, with lesser amounts in capillaries of uterus, spleen, and kidney in some of the dogs. In two adult dogs, bluetongue viral nucleic acid was detected in mononuclear cells of the periarteriolar lymphoid sheaths of spleen. There was minimal staining of capillaries in placentae in three of the five fetuses examined. There was no viral nucleic acid detected in any of the other fetal tissues.

scholarly journals N-myc proto-oncogene expression during organogenesis in the developing mouse as revealed by in situ hybridization.

1988 ◽  
Vol 107 (4) ◽  
pp. 1325-1335 ◽  
Author(s):  
G Mugrauer ◽  
F W Alt ◽  
P Ekblom
Keyword(s):  
In Situ Hybridization ◽  
Hair Follicles ◽  
Early Differentiation ◽  
Cell Lineages ◽  
Tissue Sections ◽  
Embryonic Tissues ◽  
Differentiated Cells ◽  
The Central Nervous System ◽  
Developing Kidney

The N-myc proto-oncogene is expressed during embryogenesis, suggesting that it plays a role in normal development. Since the myc-family oncogenes have been implicated in the control of cell growth, the embryonic expression may reflect rapid proliferation known to occur in development. Alternatively, N-myc expression may be involved in specific differentiation stages. In many embryonic tissues, early and late differentiation events occur in different locations. By in situ hybridization of tissue sections, we now demonstrate a restricted expression of N-myc mRNA to a few tissues and to areas where the first differentiation stages occur. N-myc expression was most strongly expressed in the developing kidney, hair follicles, and in various parts of the central nervous system. In these tissues, expression was restricted to a few cell lineages. In all lineages, expression was confined to early differentiation stages, and, at onset of overt differentiation, the level of expression decreased dramatically. Several rapidly proliferating tissues showed very little, if any, N-myc expression. In the brain, post-mitotic but not yet differentiated cells expressed high levels of N-myc mRNA. Therefore, N-myc expression is not a simple marker for proliferation in the embryo. Rather, N-myc expression seems to be a feature of early differentiation stages of some cell lineages in kidney, brain, and hair follicles, regardless of the proliferative status of the cell. The results raise the possibility that N-myc may participate in the control of these early differentiation events.

scholarly journals Nonspecific binding of nucleic acid probes to Paneth cells in the gastrointestinal tract with in situ hybridization.

1992 ◽  
Vol 40 (10) ◽  
pp. 1613-1618 ◽  
Author(s):  
K L Garrett ◽  
M D Grounds ◽  
M W Beilharz
Keyword(s):  
In Situ Hybridization ◽  
Gastrointestinal Tract ◽  
Nucleic Acid ◽  
Paneth Cells ◽  
Proteinase K ◽  
Hybridization Technique ◽  
Nonspecific Binding ◽  
Nucleic Acid Probes ◽  
Hybridization Data

Nonspecific binding of a number of unrelated nucleic acid probes to cells in the crypts of Lieberkuhn was observed in the small intestine of mice with the in situ hybridization technique. Hybridization signal was localized to cells which, by virtue of their histological position, represented Paneth cells. This signal could not be removed by RNAse, DNAse, or proteinase K treatment, and was not removed after high-stringency washing conditions. This report indicates that caution must be exercised in the interpretation of in situ hybridization data when looking for nucleic acid sequences in the gastrointestinal tract.

scholarly journals Comparative usefulness of tissue fixatives for in situ viral nucleic acid hybridization.

1985 ◽  
Vol 33 (10) ◽  
pp. 1026-1032 ◽  
Author(s):  
H A McAllister ◽  
D L Rock
Keyword(s):  
In Situ Hybridization ◽  
Nucleic Acid ◽  
Pseudorabies Virus ◽  
Nucleic Acid Hybridization ◽  
Viral Nucleic Acid ◽  
Tissue Sections ◽  
Laboratory Rodents ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded

Traditionally tissues for in situ hybridization of viral nucleic acid have been small pieces obtained from laboratory rodents, and fixatives that are designed for electron microscopy, such as periodate-lysine-paraformaldehyde (PLP) can handle them adequately. However, these fixatives have limited penetrating ability and may produce no appreciable hardening, so alternative fixation methods were evaluated. The intention was to determine whether fixatives adequate for bulky tissues such as whole or halved pig and cow brains would also be compatible with in situ hybridization. Various fixatives were evaluated using a system of intracranial inoculation of BALB/c mice with pseudorabies virus (PRV) followed by in situ hybridization of brain tissue sections with a 35S-labeled PRV DNA probe. Loss of tissue sections was a major problem, particularly with PLP and formalin, but positive results were obtained with five fixatives tested. Cellular morphology was especially good with PLP and with a modification of Carnoy's fluid, MOCA fixative. An incidental but important observation was that formalin is compatible with in situ hybridization. Retroactive studies of viral diseases using routinely processed blocks of tissue (formalin-fixed, paraffin-embedded) are therefore conceivable.

scholarly journals Simultaneous detection of two messenger RNAs in the central nervous system: a simple two-step in situ hybridization procedure using a combination of radioactive and non-radioactive probes.

1991 ◽  
Vol 39 (11) ◽  
pp. 1575-1578 ◽  
Author(s):  
E Normand ◽  
B Bloch
Keyword(s):  
Central Nervous System ◽  
In Situ Hybridization ◽  
Simultaneous Detection ◽  
Messenger Rnas ◽  
Tissue Sections ◽  
System A ◽  
Step Procedure ◽  
Hybridization Procedure ◽  
The Central Nervous System

We present here a method enabling the simultaneous detection of two messenger RNAs in tissue sections by use of a two-step in situ hybridization procedure. Tissue sections were hybridized with a radioactive probe and coated with emulsion. The emulsion was processed for development, fixed, and a second hybridization was performed through the emulsion with a biotinylated probe subsequently revealed with streptavidin-alkaline phosphatase. This procedure allows the detection of two mRNAs without loss of signal, removal of the emulsion, or spurious reaction. The simultaneous detection of oxytocin and vasopressin mRNAs in the hypothalamus, and of dopamine receptor and neuropeptide mRNAs in the striatum, demonstrated the efficiency of the procedure. Such a two-step procedure provides a simple and flexible way to make possible comparative analysis of the localization of two mRNAs within the same tissue section.

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