title: Western blot detection of PrP Sc in archived paraffin-embedded brainstem from scrapie-affected sheep.
abstract: Scrapie is a naturally occurring fatal neurodegenerative disease of adult sheep and goats, one of a group of mammalian diseases known as transmissible spongiform encephalopathies (TSE) or prion diseases. Immunoassays that identify disease-associated prion protein (PrP Sc) are integral to the diagnosis of scrapie and other prion diseases. Results obtained by either immunohistochemistry (IHC) or Western blot (WB) assay are generally adequate for the definitive diagnosis. Approved or accepted methods for WB diagnosis of TSEs requires the use of fresh or frozen nonfixed tissue samples, whereas formalin-fixed, paraffin-embedded tissue is required for the localization of PrP Sc by IHC. Because disparate processing methods are used for these accepted diagnostic techniques, separate tissue samples are collected from the same animal. Occasions arise in which there is either insufficient quantity of tissue available to complete analysis by both techniques or initial tissue processing is incompatible with one of the assays. Also, results between the assays may differ because of the vagaries of sampling, especially in case material that contains moderate-to-low levels of PrP Sc. The present article describes a method to conduct a WB assay from the same paraffin-embedded brainstem sample used for the IHC diagnosis of experimentally induced sheep scrapie.
authors.affiliation: National Animal Disease Center, US Department of Agriculture, Animal Research Service, Ames, IA 50010, USA. Robert.Kunkle@ars.usda.gov
authors.forename: Robert A
authors.initials: RA
authors.lastname: Kunkle
authors.forename: Eric M
authors.initials: EM
authors.lastname: Nicholson
authors.forename: Semakaleng
authors.initials: S
authors.lastname: Lebepe-Mazur
authors.forename: Dennis L
authors.initials: DL
authors.lastname: Orcutt
authors.forename: Megan L
authors.initials: ML
authors.lastname: Srinivas
authors.forename: Justin J
authors.initials: JJ
authors.lastname: Greenlee
authors.forename: David P
authors.initials: DP
authors.lastname: Alt
authors.forename: Amir N
authors.initials: AN
authors.lastname: Hamir
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000662:veterinary*; D001933:Brain Stem / Q000378:metabolism*; D016612:Paraffin Embedding; D018620:PrPSc Proteins / Q000302:isolation & purification*; D012608:Scrapie / Q000473:pathology*; D012680:Sensitivity and Specificity; D012756:Sheep; D013048:Specimen Handling
chemical_list: D018620:PrPSc Proteins