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United States Department of Agriculture

Agricultural Research Service

Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES Title: Western-blot detection of PrP**sc in archived paraffin-embedded brainstem from scrapie-affected sheep

Authors
item Kunkle, Robert
item NICHOLSON, ERIC
item LEBEPE-MAZUR, SEMAKALENG
item ORCUTT, DENNIS
item Srinivas, Megan
item GREENLEE, JUSTIN
item ALT, DAVID
item Hamir, Amirali

Submitted to: Journal of Veterinary Diagnostic Investigation
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 10, 2008
Publication Date: July 1, 2008
Citation: Kunkle, R.A., Nicholson, E.M., Lebepe-Mazur, S., Orcutt, D.L., Srinivas, M.L., Greenlee, J.J., Alt, D.P., Hamir, A.N. 2008. Western blot detection of PrP**sc in archived paraffin-embedded brainstem from scrapie-affected sheep. Journal of Veterinary Diagnostic Investigation. 20(4):522-526.

Interpretive Summary: Scrapie is a fatal brain disease of adult sheep and goats, one of a group of mammalian prion diseases known as transmissible spongiform encephalopathies (TSEs). Assays which identify disease-associated prions (PrP**sc) are integral to the diagnosis of scrapie and other TSEs. Results obtained by either immunohistochemistry (IHC) or Western blot (WB) assay are required for the diagnosis of TSE and for presumptive strain typing of the PrP**sc found in affected tissues. Approved or accepted methods for WB diagnosis of TSE requires the use of fresh or frozen non-fixed tissue samples, whereas formalin-fixed, paraffin-embedded tissue is required for the localization of PrP**sc by IHC. Since disparate processing methods are used for WB and IHC, separate tissue samples are collected from the same animal. Occasions arise in which there is either insufficient quantity of tissue available for both IHC and WB or initial tissue processing is incompatible with one of the assays. This communication represents the first description of a method to conduct a WB assay from the same paraffin-embedded brainstem sample used for the IHC diagnosis of sheep scrapie.

Technical 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 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 or Western blot assay are generally adequate for the definitive diagnosis. Approved or accepted methods for Western blot diagnosis of transmissible spongiform encephalopathies requires the use of fresh or frozen non-fixed tissue samples, whereas formalin-fixed, paraffin-embedded tissue is required for the localization of PrP**Sc by immunohistochemistry. Since 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 due to the vagaries of sampling, especially in case material containing moderate to low levels of PrP**Sc. This communication describes a method to conduct a Western blot assay from the same paraffin-embedded brainstem sample used for the immunohistochemical diagnosis of experimentally-induced sheep scrapie.

Last Modified: 7/28/2014
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