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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Virus and Prion Research » Research » Publications at this Location » Publication #363307

Research Project: Pathobiology, Genetics, and Detection of Transmissible Spongiform Encephalopathies

Location: Virus and Prion Research

Title: PAD-bead enrichment enhances detection of PrPSc using real-time quaking-induced conversion

item Hwang, Soyoun
item Dassanayake, Rohana
item Nicholson, Eric

Submitted to: Bioscience Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/4/2019
Publication Date: 12/13/2019
Citation: Hwang, S., Dassanayake, R.P., Nicholson, E.M. 2019. PAD-bead enrichment enhances detection of PrPSc using real-time quaking-induced conversion. Bioscience Reports. 12:806.

Interpretive Summary: Prion diseases are fatal neurodegenerative diseases that affect a wide range of livestock and wildlife. The disease process occurs through the misfolding of a normally occuring protein. Detection of this misfolded protein is the only known means by which a prion disease can be diagnosed. A recently developed approach for the detection of this misfolded protein uses a technique referred to as Real-time quaking induced conversion (RT-QuIC). RT-QuIC amplifies the amount of misfolded protein available for detection but can be inhibited by naturally occurring contaminants in the tissue samples used for the technique. In this work we present a method to clean the samples prior to use and show that this sample cleanup technique enhances the detection and reduces the time required to assess the results.

Technical Abstract: Scrapie is a transmissible spongiform encephalopathy (TSE) that naturally occurs in sheep and goats. This fatal neurodegenerative disease results from misfolding of the normal cellular prion protein (PrPC) to a pathogenic prion protein form (PrPSc). This pathogenic form, PrPSc, accumulates in the brain and lymphoid tissues. The presence of PrPSc can be detected by an in vitro conversion assay known as real-time quaking induced conversion (RT-QuIC). RT-QuIC has been used to detect PrPSc in a variety of biological tissues from brains to fluids. While this technique is both rapid and sensitive enhancing the detection of prions would be valuable in the diagnostic laboratories. In this study, we assessed whether PrPSc detection sensitivity of RT-QuIC can be increased by enriching PrPSc in scrapie tissue homogenates using commercially available aggregated protein binding ligands coated magnetic beads (PAD-Beads). Coupling the RT-QuIC to PAD-Bead based cleanup allowed detection of PrPSc rapidly and without dilutions of scrapie sheep brain homogenates prior to RT-QuIC. The PAD-Beads sample pretreatment step prior to RT-QuIC is a useful enhancement in the diagnosis of TSEs.