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Isolation of Proteinase K-Sensitive Prions Using Pronase E and Phosphotungstic Acid

D'Castro, L; Wenborn, A; Gros, N; Joiner, S; Cronier, S; Collinge, J; Wadsworth, JDF; (2010) Isolation of Proteinase K-Sensitive Prions Using Pronase E and Phosphotungstic Acid. PLOS ONE , 5 (12) , Article e15679. 10.1371/journal.pone.0015679. Green open access

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Abstract

Disease-related prion protein, PrPSc, is classically distinguished from its normal cellular precursor, PrPC, by its detergent insolubility and partial resistance to proteolysis. Molecular diagnosis of prion disease typically relies upon detection of protease-resistant fragments of PrPSc using proteinase K, however it is now apparent that the majority of disease-related PrP and indeed prion infectivity may be destroyed by this treatment. Here we report that digestion of RML prion-infected mouse brain with pronase E, followed by precipitation with sodium phosphotungstic acid, eliminates the large majority of brain proteins, including PrPC, while preserving >70% of infectious prion titre. This procedure now allows characterization of proteinase K-sensitive prions and investigation of their clinical relevance in human and animal prion disease without being confounded by contaminating PrP

Type: Article
Title: Isolation of Proteinase K-Sensitive Prions Using Pronase E and Phosphotungstic Acid
Open access status: An open access version is available from UCL Discovery
DOI: 10.1371/journal.pone.0015679
Publisher version: http://dx.doi.org/10.1371/journal.pone.0015679
Language: English
Additional information: © 2010 D'Castro et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. This research was funded by the UK Medical Research Council (http://www.mrc.ac.uk/index.htm). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors have read the journal's policy and have the following conflicts. J.C. is a Director and J.C. and J.D.F.W. are shareholders and consultants of D-Gen Limited, an academic spin-out company working in the field of prion disease diagnosis, decontamination, and therapeutics. D-Gen markets the ICSM35 and ICSM18 antibodies used in this study. This does not alter the authors' adherence to all PLoS ONE policies on sharing data and materials.
Keywords: TRANSMISSIBLE MINK ENCEPHALOPATHY, SPONGIFORM ENCEPHALOPATHY, STRAIN VARIATION, TRANSGENIC MICE, MOLECULAR-BASIS, DISEASE, PROTEASE, SCRAPIE, PRPSC, THERMOLYSIN
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Institute of Prion Diseases
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > UCL Institute of Prion Diseases > MRC Prion Unit at UCL
URI: https://discovery.ucl.ac.uk/id/eprint/1301571
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