Sensitive and specific detection of sporadic Creutzfeldt-Jakob disease brain prion protein using real-time quaking-induced conversion.
Real-time quaking-induced conversion (RT-QuIC) is an assay in which disease-associated prion protein (PrP) initiates a rapid conformational transition in recombinant PrP (recPrP), resulting in the formation of amyloid that can be monitored in real time using the dye thioflavin T. It therefore has potential advantages over analogous cell-free PrP conversion assays such as protein misfolding cyclic amplification (PMCA). The QuIC assay and the related amyloid seeding assay have been developed largely using rodent-passaged sheep scrapie strains. Given the potential RT-QuIC has for Creutzfeldt-Jakob disease (CJD) research and human prion test development, this study characterized the behaviour of a range of CJD brain specimens with hamster and human recPrP in the RT-QuIC assay. The results showed that RT-QuIC is a rapid, sensitive and specific test for the form of abnormal PrP found in the most commonly occurring forms of sporadic CJD. The assay appeared to be largely independent of species-related sequence differences between human and hamster recPrP and of the methionine/valine polymorphism at codon 129 of the human PrP gene. However, with the same conditions and substrate, the assay was less efficient in detecting the abnormal PrP that characterizes variant CJD brain. Comparison of these QuIC results with those previously obtained using PMCA suggested that these two seemingly similar assays differ in important respects.
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Alexander H
AH
Peden
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Lynne I
LI
McGuire
Bristol Institute for Transfusion Sciences, National Blood Service, Bristol, UK.
Nigel E J
NEJ
Appleford
Bristol Institute for Transfusion Sciences, National Blood Service, Bristol, UK.
Gary
G
Mallinson
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, Hamilton, MT, USA.
Jason M
JM
Wilham
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, Hamilton, MT, USA.
Christina D
CD
Orrú
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, Hamilton, MT, USA.
Byron
B
Caughey
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
James W
JW
Ironside
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Richard S
RS
Knight
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Robert G
RG
Will
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Alison J E
AJE
Green
National CJD Research and Surveillance Unit, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK.
Mark W
MW
Head
D000682:Amyloid / Q000032:analysis; D000818:Animals; D001921:Brain / Q000473:pathology*; D007562:Creutzfeldt-Jakob Syndrome / Q000175:diagnosis*; D006224:Cricetinae; D006801:Humans; D010336:Pathology / Q000379:methods*; D011328:Prions / Q000032:analysis*; D011487:Protein Conformation; D017510:Protein Folding; D013997:Time Factors
D000682:Amyloid; D011328:Prions