1887

Abstract

Real-time quaking-induced conversion (RT-QuIC) is a rapid, specific and highly sensitive prion seeding activity detection assay that uses recombinant prion protein (rPrP) to detect subinfectious levels of the abnormal isoforms of the prion protein (PrP). Although RT-QuIC has been successfully used to detect PrP in various tissues from humans and animals, including sheep, tissues from goats infected with classical scrapie have not yet been tested. Therefore, the aims of the present study were to (1) evaluate whether prion seeding activity could be detected in the brain tissues of goats with scrapie using RT-QuIC, (2) optimize reaction conditions to improve scrapie detection in goats, and (3) compare the performance of RT-QuIC for the detection of PrP with the more commonly used ELISA and Western blot assays. We further optimized RT-QuIC conditions for sensitive and specific detection of goat scrapie seeding activity in brain tissue from clinical animals. When used with 200 mM sodium chloride, both full-length sheep rPrP substrates (PrP genotypes ARQ and VRQ) provided good discrimination between scrapie-infected and normal goat brain samples at 10 dilution within 15 h. Our findings indicate that RT-QuIC was at least 10 000-fold more sensitive than ELISA and Western blot assays for the detection of scrapie seeding activity in goat brain samples. In addition to WT samples, positive RT-QuIC reactions were also observed with three polymorphic goat brain samples (G/S127, I/M142 and H/R143) tested. Taken together, these findings demonstrate that RT-QuIC sensitively detects prion seeding activity in classical scrapie-infected goat brain samples.

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2016-03-01
2019-10-14
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