Ultrasensitive human prion detection in cerebrospinal fluid by real-time quaking-induced conversion (original) (raw)

The development of technologies for the in vitro amplification of the abnormal conformers of prion protein (PrP Sc ) has generated the potential for a novel diagnostic assay for prion disease. Previously, we developed a new PrP Sc amplification assay designated quaking-induced conversion (QUIC), which involves intermittent, automated shaking of the substrate, soluble recombinant PrP. We further improved the rapidity and practicality of this method by combining it with thioflavin T fluorescence to monitor the amyloid fibril formation. This assay, termed "real-time QUIC (RT-QUIC)", allows within 48 h, the detection of ≥1 fg of PrP Sc in diluted Creutzfeldt-Jakob disease (CJD) brain homogenate. Moreover, we assessed the technique first in a series of Japanese subjects, and then in a blind study of 30 cerebrospinal fluid specimens from Australia, which achieved greater than 80% sensitivity and 100% specificity. These findings indicate the promising enhanced diagnostic capacity of RT-QUIC in the ante-mortem evaluation of suspected CJD. 4 Transmissible spongiform encephalopathies or prion diseases are characteristically associated with the accumulation of PrP Sc in the central nervous system through auto-catalytic conversion of normal cellular PrP (PrP c ) into replicate misfolded isomers 1,2 . Despite occasional reported exceptions 3,4 , PrP Sc remains the best characterized and most reliable marker of prion disease.

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