Journal Article Rapid and Quantitative Assay of Amyloid-Seeding Activity in Human Brains Affected with Prion Diseases

Takatsuki, Hanae  ,  Satoh, Katsuya  ,  Sano, Kazunori  ,  Fuse, Takayuki  ,  Nakagaki, Takehiro  ,  Mori, Tsuyoshi  ,  Ishibashi, Daisuke  ,  Mihara, Ban  ,  Takao, Masaki  ,  Iwasaki, Yasushi  ,  Yoshida, Mari  ,  Atarashi, Ryuichiro  ,  Nishida, Noriyuki

10 ( 6 )  , p.e0126930 , 2015-06-12 , Public Library of Science
Description
The infectious agents of the transmissible spongiform encephalopathies are composed of amyloidogenic prion protein, PrPSc. Real-time quaking-induced conversion can amplify very small amounts of PrPSc seeds in tissues/body fluids of patients or animals. Using this in vitro PrP-amyloid amplification assay, we quantitated the seeding activity of affected human brains. End-point assay using serially diluted brain homogenates of sporadic Creutzfeldt-Jakob disease patients demonstrated that 50% seeding dose (SD50) is reached approximately 1010/g brain (values varies 108.79-10.63/g). A genetic case (GSS-P102L) yielded a similar level of seeding activity in an autopsy brain sample. The range of PrPSc concentrations in the samples, determined by dot-blot assay, was 0.6-5.4 μg/g brain; therefore, we estimated that 1 SD50 unit was equivalent to 0.06-0.27 fg of PrPSc. The SD50 values of the affected brains dropped more than three orders of magnitude after autoclaving at 121°C. This new method for quantitation of human prion activity provides a new way to reduce the risk of iatrogenic prion transmission.
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http://naosite.lb.nagasaki-u.ac.jp/dspace/bitstream/10069/35721/1/PLoS10_126930.pdf

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