title: Immunohistochemical detection of prion protein in sheep with scrapie.
abstract: Prion protein (PrP), which is involved in the pathogenesis of scrapie, occurs in 2 forms. The form extracted from scrapie brain is protease resistant (PrP-res), whereas PrP from normal brain is protease sensitive (PrP-sen). This study examined whether PrP-res could be detected in brains of sheep with scrapie by immunohistochemistry (IHC). A suitable IHC procedure was developed using brain tissue from hamsters that had been inoculated with the transmissible mink encephalopathy agent. Tissue samples were fixed in PLP (periodate, lysine, paraformaldehyde) that contained paraformaldehyde at a concentration of 0.125%. Before application of the IHC technique, tissue sections were deparaffinized and treated with formic acid to simultaneously enhance PrP-res immunoreactivity and degrade PrP-sen. Primary antibody was obtained from a rabbit immunized to PrP-res extracted from brains of mice with experimentally induced scrapie. Brain from 21 sheep with histopathologically confirmed scrapie were examined by IHC. In all 21 brains, PrP-res was widely distributed throughout the brain stem. Staining was particularly intense in neuronal cell bodies and around blood vessels. The IHC technique successfully detected PrP-res in brain samples that had been frozen or that were severely autolyzed before fixation in PLP. Brains from 11 scrapie-suspect sheep that were not considered histologically positive were also examined by IHC. PrP-res was found in 4 of these brains. Sections of brains from 14 clinically normal sheep did not have detectable PrP-res. Results of this study indicate that IHC detection of PrP-res is equivalent, and perhaps superior, to histopathology for the diagnosis of scrapie in sheep. Furthermore, IHC is applicable to tissues that have autolytic changes or processing artifacts that prevent satisfactory histopathologic evaluation for lesions of scrapie.
authors.affiliation: Virology Cattle Research Unit, USDA, Agricultural Research Service, Ames, IA 50010.
authors.forename: J M
authors.initials: JM
authors.lastname: Miller
authors.forename: A L
authors.initials: AL
authors.lastname: Jenny
authors.forename: W D
authors.initials: WD
authors.lastname: Taylor
authors.forename: R F
authors.initials: RF
authors.lastname: Marsh
authors.forename: R
authors.initials: R
authors.lastname: Rubenstein
authors.forename: R E
authors.initials: RE
authors.lastname: Race
mesh_terms: D000818:Animals; D001921:Brain / Q000382:microbiology; D006224:Cricetinae; D005557:Formaldehyde; D007150:Immunohistochemistry / Q000379:methods*; D008239:Lysine; D010504:Periodic Acid; D011108:Polymers; D018620:PrPSc Proteins; D011328:Prions / Q000032:analysis*; D012608:Scrapie / Q000382:microbiology*; D012756:Sheep
chemical_list: D011108:Polymers; D018620:PrPSc Proteins; D011328:Prions; D010504:Periodic Acid; D005557:Formaldehyde; C009288:metaperiodate; D008239:Lysine; C003043:paraform