title: Detection of PrP(Sc) in formalin-fixed, paraffin-embedded tissue by Western blot differentiates classical scrapie, Nor98 scrapie, and bovine spongiform encephalopathy.
abstract: Transmissible, spongiform encephalopathies including bovine spongiform encephalopathy (BSE) and scrapie are fatal neurodegenerative disorders associated with the presence of an infectious abnormal isoform of normal mammalian proteins called prions. Identification of the prion protein associated with scrapie (PrP(Sc)) in the central nervous system is typically based upon immunoassays including immunohistochemistry (IHC) using formalin-fixed tissues or Western blot (WB) assays using fresh and/or frozen, non-formalin-fixed tissues. Each assay can discriminate between BSE, classical scrapie, and a previously reported strain of scrapie recently identified in the United States named Nor98 scrapie. Different tissue samples are required from the same animal to run these 2 different immunoassays. This may result in inconsistent test results for the same animal. Sampling problems such as collecting insufficient volumes of fresh tissue or less than optimal anatomic location of brainstem for IHC can affect the ability of the test procedures to offer definitive and discriminatory results. Recently, a WB method using formalin-fixed, paraffin-embedded (FFPE) tissue to identify PrP(Sc) was developed that successfully identified PrP(Sc) in sheep affected by classical scrapie. In the current study, the use of this technique to produce discriminatory results identifying classical BSE in bovine tissue and both classical and Nor98 scrapie in ovine tissue using paraffin-embedded brain samples is described. Protein-banding patterns from WB using FFPE tissue were similar to protein-banding patterns produced by WB assays utilizing fresh tissues from the same animals, and results correlated well with the IHC PrP(Sc)-positive staining present in the cerebellum and obex regions of brain samples from these animals.
authors.affiliation: Animal and Plant Health Inspection Services, National Veterinary Services Laboratories, Pathobiology Laboratory, U.S. Department of Agriculture, Ames, IA 50010, USA. Christina.M.Loiacono@aphis.usda.gov
authors.forename: Christina M
authors.initials: CM
authors.lastname: Loiacono
authors.forename: Nadine
authors.initials: N
authors.lastname: Beckwith
authors.forename: Robert A
authors.initials: RA
authors.lastname: Kunkle
authors.forename: Dennis
authors.initials: D
authors.lastname: Orcutt
authors.forename: S Mark
authors.initials: SM
authors.lastname: Hall
mesh_terms: D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000473:pathology; D002417:Cattle; D002531:Cerebellum / Q000473:pathology; D003937:Diagnosis, Differential; D016643:Encephalopathy, Bovine Spongiform / Q000175:diagnosis* / Q000473:pathology; D007118:Immunoassay / Q000379:methods; D007150:Immunohistochemistry; D018620:PrPSc Proteins / Q000032:analysis*; D012608:Scrapie / Q000175:diagnosis* / Q000473:pathology; D012756:Sheep
chemical_list: D018620:PrPSc Proteins