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immunochemical tests (FITs) for detection of fecal occult blood in human medicine. 7 – 10 FITs use an antibody against human globin to detect blood and therefore are unaffected by diet or medications. 11 These tests detect smaller quantities of hemoglobin than

Open access
in American Journal of Veterinary Research

SUMMARY

A pig rhabdomyosarcoma cell line (PRUM59) was established, and the immuno(histo)chemical and cytogenetic characterization of these cells was determined. At various swine farms in the Netherlands, pigs were observed that had solitary or multiple skin nodules, which were diagnosed as rhabdomyosarcomas. Cells of a tumor derived from a 3.5-week-old female pig were cultured for immunochemical and cytogenetic analyses. The cell line had characteristic features of undifferentiated muscle cells, similar to those observed in tumor tissue sections; they contained titin, a high-molecular weight protein specific for striated muscle, as dot-like aggregates and as filaments, desmin filaments and cross-striations, smooth muscle actin stress fibers, and vimentin filaments. The cells stained positively for striated muscle actin and tropomyosin as well. The immunohistochemical staining results were supported by results of immunoblotting experiments. Karyotyping of the cells revealed a deletion of a major part of Xq24-qter, a part of the long arm of 1 of the 2 X chromosomes. The other X chromosome and all autosomes appeared to be normal.

Free access
in American Journal of Veterinary Research

SUMMARY

Eleven adult goats and 32 adult outbred mice were inoculated iv with Cowdria ruminantium-infected blood (Kwanyanga isolate), monitored clinically, then serially euthanatized. Predominant clinical signs of disease in goats were depression, head tremors, seizures, and dyspnea. In mice, dyspnea and depression were the only clinical signs of disease noticed. Tissues were examined histologically and immunohistochemically for C ruminantium colonies or antigen. In goats, C ruminantium was detected only in endothelial cells of the brain, even though gross and microscopic lesions were confined to the thorax. In mice, C ruminantium was detected only in endothelial cells of the heart and lungs.

Free access
in American Journal of Veterinary Research

Abstract

Objectives

To develop methods to isolate, culture, and characterize equine hoof endothelial cells (EC) and keratinocytes.

Sample Population

Cells harvested from the forelimbs of 8 horses.

Procedure

EC were obtained via catheters placed in the palmar digital arteries of the disarticulated lower portion of the forelimbs from 4 horses that had been heparinized prior to euthanasia. Phosphate-buffered saline solution was used to remove and discard RBC from blood vessels, and collagenase was used to loosen and flush EC from the vasculature. Hoof keratinocytes were obtained from 4 recently euthanatized horses by use of dispase/trypsin dissociation of the coronary band epidermis. Use of an extracellular matrix gel as a culture flask attachment factor was important to the success of hoof keratinocyte cultures.

Results

EC from the palmar digital arteries were successfully cultured and characterized by in vitro morphology, uptake of a fluorescence-labeled acetylated-low density lipoprotein, and lack of expression of von Willebrand factor and smooth muscle actin. Hoof keratinocytes were characterized by morphology in culture and expression of keratin proteins, as determined by immunochemical reaction. Keratinocyte cultures were also positive for vimentin expression.

Conclusions

Culture techniques to isolate and characterize hoof cells should aid investigators in their study of equine hoof pathobiologic features, especially as it relates to laminitis. (Am J Vet Res 1999;60:128–132)

Free access
in American Journal of Veterinary Research

SUMMARY

C-reactive protein (crp) was isolated from equine serum by use of calcium-dependent affinity chromatography conjugated pneumococcal C-polysaccharide, anion exchange chromatography, and gel filtration. It was identified as genuine crp by its immunochemical cross-reactivity with anti-human crp, its homology with human crp in amino acid composition, and its pentameric structure as revealed by electron microscopy. Purified equine crp had a molecular weight of approximately 118,000 and was composed of 5 identical, nonglycosylated and noncovalently associated subunits with molecular weight of approximately 23,000 each. Equine crp migrated in the region between β- and γ-globulin by results of immunoelectrophoresis, and its isoelectric point was about 7.0.

In horses, increased crp concentration was associated with clinical pneumonitis, enteritis, and arthritis, compared with values obtained in clinically normal horses by use of single radial immunodiffusion method. After im administration of turpentine oil or castration, serum crp concentration increased to 6 times higher than baseline values. Results indicate that crp may be an acute-phase reactant protein in horses.

Free access
in American Journal of Veterinary Research

Abstract

Objective—To compare hepatic metabolism of pyrrolizidine alkaloids (PAs) between sheep and cattle and elucidate the protective mechanism of sheep.

Sample Population—Liver microsomes and cytosol from 8 sheep and 8 cattle.

Procedure—The PA senecionine, senecionine N-oxide (nontoxic metabolite) and 6,7-dihydro-7-hydroxy- 1-hydroxymethyl-5H-pyrrolizine (DHP; toxic metabolite) were measured in microsomal incubations. The kcat (turnover number) was determined for DHP and N-oxide formation. Chemical and immunochemical inhibitors were used to assess the role of cytochrome P450s, flavin-containing monooxygenases (FMOs), and carboxylesterases in senecionine metabolism. The CYP3A, CYP2B, and FMO concentrations and activities were determined, in addition to the role of glutathione (GSH) in senecionine metabolism.

Results—DHP concentration did not differ between species. Sheep formed more N-oxide, had higher N-oxide kcat, and metabolized senecionine faster than cattle. The P450 concentrations and isoforms had a large influence on DHP formation, whereas FMOs had a large influence on N-oxide formation. In cattle, CYP3A played a larger role in DHP formation than in sheep. FMO activity was greater in sheep than in cattle. Addition of GSH to in vitro microsomal incubations decreased DHP formation; addition of cytosol decreased N-oxide formation.

Conclusions and Clinical Relevance—Hepatic metabolism differences alone do not account for the variation in susceptibility seen between these species. Rather, increased ruminal metabolism in sheep appears to be an important protective mechanism, with hepatic enzymes providing a secondary means to degrade any PAs that are absorbed from the rumen. (Am J Vet Res 2004;65:1563–1572)

Full access
in American Journal of Veterinary Research

-derived porphyrins by use of spectrophotometry, 2,3 and species-specific immunochemical assays. 4–6 In addition, colorless chromogens that become colored when oxidized (eg, guaiac and tetramethylbenzidine) have long been used in tests that make use of the

Full access
in American Journal of Veterinary Research

difficulties associated with isolation of L borgpetersenii serovar hardjo from either tissue or urine obtained from vaccinated cattle despite histopathologic, immunochemical, and molecular evidence that organisms were present. Results reported for these

Full access
in American Journal of Veterinary Research

-shaped mesenchymal cells, and prominent blastemal cells. Some canine nephroblastomas are immunochemically reactive for the human Wilms tumor marker WT1 in tissue sections, 1 but immunocytochemical staining for WT1 in canine nephroblastomas has not been reported. A

Full access
in Journal of the American Veterinary Medical Association

. Combiplate 8, Labsystems Oy, Atlsinki, Finland e. Goat anti-dog IgA (Fc) antibodies, Nordic Immunochemicals, Tilberg, The Netherlands f. Superblock in PBS, Pierce Chemical Co, Rockford, Ill g. Dog IgA ELISA quantitation kit, Bethyl, Montgomery, Tex h

Full access
in American Journal of Veterinary Research