Magnetic microparticle-based multimer detection system for the detection of prion oligomers in sheep.

Transmissible spongiform encephalopathies (TSEs) are zoonotic fatal neurodegenerative diseases in animals and humans. TSEs are commonly known as bovine spongiform encephalopathy in cattle, scrapie in sheep and goats, chronic wasting disease in cervids, and Creutzfeldt-Jakob disease in humans. The putative transmissible agents are infectious prion proteins (PrP(Sc)), which are formed by the conversion of the normal prion protein on the glycoprotein cell surface in the presence of other PrP(Sc).

Application of capillary immunoelectrophoresis revealed an age- and gender-dependent regulated expression of PrPC in liver.

The cellular prion protein (PrPC) is a glycoprotein, anchored to the plasma membrane and abundantly expressed in the central nervous system. The expression of PrPC in the peripheral tissues is low and only little information is available on its functions in the nonneuronal tissues. The antioxidant function of PrPC during the activation of hepatic stellate cells has already been reported.

Wnt7b is required for epithelial progenitor growth and operates during epithelial-to-mesenchymal signaling in pancreatic development.

Wnt signaling is a well conserved pathway critical for growth, patterning and differentiation of multiple tissues and organs. Previous studies on Wnt signaling in the pancreas have been based predominantly on downstream pathway effector genes such as β-catenin. We here provide evidence that the canonical-pathway member Wnt7b is a physiological regulator of pancreatic progenitor cell growth.

Phenylmethimazole suppresses dsRNA-induced cytotoxicity and inflammatory cytokines in murine pancreatic beta cells and blocks viral acceleration of type 1 diabetes in NOD mice.

Accumulating evidence supports a role for viruses in the pathogenesis of type 1 diabetes mellitus (T1DM). Activation of dsRNA-sensing pathways by viral dsRNA induces the production of inflammatory cytokines and chemokines that trigger beta cell apoptosis, insulitis, and autoimmune-mediated beta cell destruction. This study was designed to evaluate and describe potential protective effects of phenylmethimazole (C10), a small molecule which blocks dsRNA-mediated signaling, on preventing dsRNA activation of beta cell apoptosis and the inflammatory pathways important in the pathogenesis of T1DM.

Notch-mediated post-translational control of Ngn3 protein stability regulates pancreatic patterning and cell fate commitment.

Ngn3 is recognized as a regulator of pancreatic endocrine formation, and Notch signaling as an important negative regulator Ngn3 gene expression. By conditionally controlling expression of Ngn3 in the pancreas, we find that these two signaling components are dynamically linked. This connection involves transcriptional repression as previously shown, but also incorporates a novel post-translational mechanism.

Intraspecies prion transmission results in selection of sheep scrapie strains.

Background: Sheep scrapie is caused by multiple prion strains, which have been classified on the basis of their biological characteristics in inbred mice. The heterogeneity of natural scrapie prions in individual sheep and in sheep flocks has not been clearly defined.

Methodology/principal Findings: In this study, we intravenously injected 2 sheep (Suffolk and Corriedale) with material from a natural case of sheep scrapie (Suffolk breed).

Conditional control of the differentiation competence of pancreatic endocrine and ductal cells by Fgf10.

Fgf10 is a critical component of mesenchymal-to-epithelial signaling during endodermal development. In the Fgf10 null pancreas, the embryonic progenitor population fails to expand, while ectopic Fgf10 expression forces progenitor arrest and organ hyperplasia. Using a conditional Fgf10 gain-of-function model, we observed that the timing of Fgf10 expression affected the cellular competence of the arrested pancreatic progenitors.

Activation of immune cells in bovine mammary gland secretions by zymosan-treated bovine serum.

Mastitis, caused by bacterial infection of the mammary gland, is a major disease of dairy cattle. The greatest risks of intramammary infection occur at the end of lactation and at the initiation of the next lactation when the cow calves. Treating serum with zymosan (yeast cell wall preparation) causes the complement to cleave, allowing this serum to serve as a source of complement fragment 5a (C5a), a potent chemoattractant and activator of the immune system.

Application of an immunocapillary electrophoresis assay to the detection of abnormal prion protein in brain, spleen and blood specimens from patients with variant Creutzfeldt-Jakob disease.

Sensitive and specific detection of abnormal prion protein in blood could provide a diagnostic test or screening assay for animal and human prion diseases. Here, the application of an immunocapillary electrophoresis (ICE) method developed for sheep scrapie to brain, spleen and blood from patients with Creutzfeldt-Jakob disease (CJD) is described. The assay involves organic-solvent extraction, a competitive immunoassay using fluorescently labelled synthetic prion protein peptides and polyclonal antibodies specific for those sequences, and analysis by capillary electrophoresis using laser-induced fluorescence detection.

Evaluation of a preclinical blood test for scrapie in sheep using immunocapillary electrophoresis.

An analytical method is described for detection of endogenous disease-associated prion protein in the buffy coat fraction from the blood of sheep infected with scrapie. The method has been improved and evaluated for its performance in the preclinical diagnosis of ovine transmissible spongiform encephalopathies. The test system uses a protocol for sample preparation that includes extraction and concentration and a test method that uses a liquid-phase competitive immunoassay for prion protein.

Rapid PrP(Sc) detection in lymphoid tissue and application to scrapie surveillance of fallen stock in Japan: variable PrP(Sc) accumulation in palatal tonsil in natural scrapie.

Rapid western blot (WB) procedure for an abnormal isoform of prion protein (PrP(Sc) ) detection in lymphoid tissues was established and has been applied to the surveillance of fallen stock. In this program, brain and palatal tonsil were examined by WB and three cases of sheep scrapie were detected. While one clinically scrapie-infected sheep harbored PrP(Sc) in the brain and palatal tonsil, the two sheep in the pre-clinical stage harbored PrP(Sc) in the brain, but not in the palatal tonsil.

Capillary electrophoresis-based noncompetitive immunoassay for the prion protein using fluorescein-labeled protein A as a fluorescent probe.

A novel CE-based noncompetitive immunoassay for prion protein (PrP) was established. Fluorescein isothiocyanate (FITC)-labeled protein A (FITC-PrA) was used as a fluorescent probe to tag monoclonal antibody through noncovalent binding of FITC-PrA to the Fc region of the antibody. The FITC-PrA-Ab was incubated with the analyte, prion protein, under optimized condition, forming the immunocomplex FITC-PrA-Ab-PrP.

Conformational biosensor for diagnosis of prion diseases.

A fluorescence technology to monitor the proliferation of amyloidogenic neurological disorders is proposed. A crude brain homogenate (0.01%) from animals infected with a transmissible spongiform encephalopathy is employed as a catalytic medium initiating conformational changes in 520 nM polypeptide biosensors (Tris/trifluoroethanol 50% mixture at pH 7).

Detection of prion protein using a capillary electrophoresis-based competitive immunoassay with laser-induced fluorescence detection and cyclodextrin-aided separation.

The development of capillary electrophoresis (CE)-based competitive immunoassay for prion protein (PrP) using carboxymethyl beta-cyclodextrin (CM-beta-CD) as a buffer additive is described here. The assay was based on the competitive binding of PrP and a fluorescein-labeled peptide from the prion protein with a limiting amount of specific antibody. The amount of both free and fluorescein-labeled peptide bound to antibody (immunocomplex) were determined by CE with laser-induced fluorescence detection.

Analysis of the performance of antibody capture methods using fluorescent peptides with capillary zone electrophoresis with laser-induced fluorescence.

A method to analyze the performance of an antibody capture method using fluorescent peptides by capillary zone electrophoresis using laser-induced fluorescence (CZE-LIF) for detection has been developed. Fluorescent peptides from the prion protein were synthesized and the corresponding antibodies were produced in rabbits against these peptides. The antibodies were used to capture the fluorescent peptides.

Diagnosis of preclinical and subclinical scrapie in a naturally infected sheep flock utilizing currently available postmortem diagnostic techniques.

Scrapie is a naturally occurring transmissible encephalopathy of sheep and goats. Currently available methods for diagnosis are the presence of characteristic histopathologic changes and detection of an abnormal form of prion protein (PrPres) in the brains of affected animals. This study documents preclinical and subclinical scrapie in a flock of 16 sheep utilizing histopathology, immunohistochemistry (IHC), western blot, and electron microscopy (for scrapie-associated fibrils) for confirmation of the disease.

Use of capillary electrophoresis and fluorescent labeled peptides to detect the abnormal prion protein in the blood of animals that are infected with a transmissible spongiform encephalopathy.

Transmissible spongiform encephalopathies in humans and in animals are fatal neuro-degenerative diseases with long incubation times. The putative cause of these diseases is a normal host protein, the prion protein, that becomes altered. This abnormal prion protein is found mostly in the brains of infected individuals in later stages of the disease, but also can be found in lymphoid and other tissues in lower amounts.

Capillary isoelectric focusing of the scrapie prion protein.

Prion diseases or transmissible spongiform encephalopathies belong to a group of neurodegenerative diseases that infect both animals and humans. These diseases are associated with an accumulation of fibrils in the brains of infected individuals. These fibrils are composed of an abnormal isoform of a host-encoded glycoprotein that is characterized by its insolubility and partial resistance to proteases.

A diagnostic test for scrapie-infected sheep using a capillary electrophoresis immunoassay with fluorescent-labeled peptides.

Scrapie in sheep and goats is the prototype of transmissible spongiform encephalopathies found in humans and animals. A feature of these diseases is the accumulation of rod-shaped fibrils in the brain that form from an aggregated protein. This protein (PrPSC) is a protease-resistant form of a normal host cell protein.

Use of capillary sodium dodecyl sulfate gel electrophoresis to detect the prion protein extracted from scrapie-infected sheep.

Scrapie in sheep and in goats is the prototype of a group of transmissible spongiform encephalopathies (TSE). A feature of these diseases is the accumulation in the brain of rod shaped fibrils that form from an aggregated protein that is a protease-resistant form of a modified normal host cell protein. In this study, we compared SDS gel capillary electrophoresis to conventional SDS-PAGE and Western blot to detect the monomer of this aggregated protein.

Improvements in a competition assay to detect scrapie prion protein by capillary electrophoresis.

Scrapie in sheep and goats is the prototype of transmissible spongiform encephalopathies found in humans and animals. A feature of these diseases is the accumulation of rod-shaped fibrils in the brain that form from an aggregated protein. This protein is a protease-resistant form of a normal host cell protein.

Capillary electrophoresis of the scrapie prion protein from sheep brain.

Scrapie in sheep and goats causes a progressive, degenerative disease of the central nervous system and is the prototype of other transmissible spongiform encephalopathies (TSE) found in humans and in animals. In samples of TSE-affected brains, unique rod-shaped structures are found and are infectious. These rods are composed of a protease-resistant, post-translationally modified cellular protein (PrPsc) that has a molecular mass of ca.

Characterization by capillary electrophoresis of the surface glycoproteins of ovine lentiviruses before and after treatment with glycosidic enzymes.

Ovine lentiviruses are a group of viruses that infect sheep and goats. These viruses contain a surface glycoprotein (SU) that is very similar among the viral strains. Sera from infected animals react equally well with SU from each strain.

Antigenic relatedness between ovine progressive pneumonia virus (OPPV) and HIV-1.

The antigenic relatedness between ovine progressive pneumonia virus (OPPV), a lentivirus that infects sheep, and human immunodeficiency virus (HIV-1) was examined by immunoblot analysis. Fourteen of 20 sheep sera, that were positive for OPPV antibodies on an immunodiffusion test, reacted with HIV-1 p24 on a commercial blot of HIV-1 and cell lysate proteins (Virostat, Portland, Maine). Sheep OPPV antisera did not bind to cellular antigens on a negative control blot of cellular proteins.

Seroprevalence of ovine progressive pneumonia virus in various domestic and wild animal species, and species susceptibility to the virus.

Ovine progressive pneumonia is caused by a lentivirus of known infectivity only for sheep and goats. Virus susceptibility of 11 other species of animals was examined. Species included cattle, chickens, deer, dogs, goats, hamsters, horses, mice, pigs, rabbits, and rats.