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SCRAPIE , 



SCRAPIE STUDIES ON SHEEP FROM A DEPOPULATED OKLAHOMA FLOCK. Proctor, S. J. and Jenkins, S. J. 

 (Project Report) 



Introduction 



Scrapie was diagnosed in a small flock (24) of Oklahoma sheep. Many of the remaining sheep in the herd were 

 losing wool. Flock depopulation was initiated and brain tissue was obtained to determine if other sheep in the herd 

 were. infected with scrapie. Three different fixatives were compared on these brains. Previous studies suggested that 

 formalin or iormal-glutaraldehyde were preferred over Zenker's or glutaraldehyde fixatives. 



Materials and Methods 



After euthanasia, the brain was removed from each sheep and the frontal lobes were removed and frozen. The 

 remainder of the brain was divided along the median plane, and each half was randomly placed in one of three 

 fixatives: 



A. Neutral-buffered 10 percent formolsaline (phosphate buffered). 



B. 10 percent formalin in 0.067 M phosphate buffer plus 7.5 percent sucrose. 



C. 1.25 percent glutaraldehyde and 5 percent formalin at 440 mOsM of phosphate buffer and saline. 

 Paraffin and frozen sections were prepared from the medulla, pons, mesencephalon, and thalamus and stained 



with azure -eosin-hematozylin and CajaFs gold-sublimate technique, respectively. 



Results 



Typical scrapie lesions could not be found in any brain sections examined. An occasional single vacuolated 

 neuron was found in a few sections; but since these vacuoles were located in the periphery of the cytoplasm and did 

 not push the nucleus eccentrically, they were not considered indicative of scrapie. 



When stained with azure-eosin-hematoxylin, there was no significant increased preservation of tissue with 

 fixatives B or C when compared to A (our routine fixative). Fixative, C (glutaraldehyde-formalin) stained poorly with 

 Gajal's gold-sublimate. There was no significant difference between A and B— the formalin fixatives. 



Discussion 



No lesions indicative of scrapie could be found in any of the sheep. Most of the vacuoles found were peripheral 

 and could have been due to handling of the brain after death. More centrally located vacuoles were found 

 predominantly in the older sheep (7-10 years old). No more than four vacuoles in one section or total brain were 

 observed. 



Although there was an increased number of astrocytes seen in many of the brains, the astrocytic processes 

 were thin, short, and uniform. They did not resemble the large irregular astrocytic processes caused by scrapie. There 

 was no indication of an increased number of reactive astrocytes with increased age. 



Preservation and stainability of microscopic sections were used as criteria to evaluate each fixative. Tissues 

 held in fixative A (our routine fixative) were preserved and stained as well as or better than tissues in the other two 

 fixatives. It was decided to continue to use "A" as the fixative of choice. 



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