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PEDS Advance Access published online on July 1, 2009
Protein Engineering Design and Selection, doi:10.1093/protein/gzp031
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Published by Oxford University Press. [2009] All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Human variant Creutzfeldt–Jakob disease and sheep scrapie PrPres detection using seeded conversion of recombinant prion protein

Christina D. Orrú1,2,{dagger}, Jason M. Wilham1,{dagger}, Andrew G. Hughson1,{dagger}, Lynne D. Raymond1, Kristin L. McNally1, Alex Bossers3, Ciriaco Ligios4 and Byron Caughey1,5

1Laboratory of Persistent Viral Diseases, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, 903 S. 4th St., Hamilton, MT 59840, USA 2Department of Biomedical Sciences and Technologies, University of Cagliari, SS554, Km 4550, Monserrato 09042, Italy 3Central Veterinary Institute of Wageningen UR (CVI) Department of Bacteriology and TSEs, 8200 AB Lelystad, The Netherlands 4 Experimental Zoo-prophylactic Institute of Sardinia, Sassari 07100, Italy

5 To whom correspondence should be addressed. E-mail: bcaughey{at}nih.gov

The pathological isoform of the prion protein (PrPres) can serve as a marker for prion diseases, but more practical tests are needed for preclinical diagnosis and sensitive detection of many prion infections. Previously we showed that the quaking-induced conversion (QuIC) assay can detect sub-femtogram levels of PrPres in scrapie-infected hamster brain tissue and distinguish cerebral spinal fluid (CSF) samples from normal and scrapie-infected hamsters. We now report the adaptation of the QuIC reaction to prion diseases of medical and agricultural interest: human variant Creutzfeldt-Jakob disease (vCJD) and sheep scrapie. PrPres-positive and -negative brain homogenates from humans and sheep were discriminated within 1–2 days with a sensitivity of 10–100 fg PrPres. More importantly, in as little as 22 h we were able to distinguish CSF samples from scrapie-infected and uninfected sheep. These results suggest the presence of prions in CSF from scrapie-infected sheep. This new method enables the relatively rapid and sensitive detection of human CJD and sheep scrapie PrPres and may facilitate the development of practical preclinical diagnostic and high-throughput interference tests.

Keywords: cerebral spinal fluid/CJD/diagnostics/prion/scrapie

Received June 2, 2009; revised June 2, 2009; accepted June 3, 2009.

{dagger} These authors contributed equally to this study.


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