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Ultrasensitive RT-QuIC Seed Amplification Assays for Disease-Associated Tau, α-Synuclein, and Prion Aggregates

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Protein Misfolding Diseases

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1873))

Abstract

The abnormal assembly of tau, α-synuclein (αSyn), or prion protein into oligomers and multimers underpins the molecular pathogenesis of multiple neurodegenerative diseases. Such pathological aggregates can often grow by seeded polymerization mechanisms. We and others have taken advantage of these mechanisms to amplify seeding activities in vitro and devise ultrasensitive, specific and quantitative assays for these etiological biomarkers. Real-time quaking-induced conversion (RT-QuIC) assays are performed in multiwell plates with fluorescent readouts, facilitating efficient throughput. Prion RT-QuIC assays on cerebrospinal fluid (CSF) samples are being widely used for antemortem diagnosis of human prion diseases. Recently, we have also described a tau RT-QuIC prototype that has been optimized for Pick disease (with predominant 3R tau pathology) that detects 3R tau seeds in postmortem CSF, and brain tissue dilutions as extreme as a billion-fold. αSyn RT-QuIC prototypes have also been developed, providing ~92% diagnostic sensitivity and 100% specificity for Parkinson’s disease and dementia with Lewy bodies using antemortem CSF. Here we provide detailed protocols for our 3R tau and αSyn RT-QuIC assays and refer the reader to published up-to-date protocols for prion RT-QuIC assays (Orru et al. Methods Mol Biol 1658:185–203, 2017; Schmitz et al. Nat Protoc 11:2233–2242, 2016).

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Acknowledgments

This work was supported by the Intramural Research Program of the NIAID.

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Authors and Affiliations

  1. Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT, USA

    Eri Saijo, Bradley R. Groveman, Allison Kraus, Michael Metrick, Christina D. Orrù, Andrew G. Hughson & Byron Caughey

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  1. Eri Saijo
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  2. Bradley R. Groveman
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  3. Allison Kraus
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  4. Michael Metrick
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  5. Christina D. Orrù
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  6. Andrew G. Hughson
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  7. Byron Caughey
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Corresponding author

Correspondence to Byron Caughey .

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Editors and Affiliations

  1. BioISI – Biosystems and Integrative Sciences Institute, Faculty of Sciences University of Lisbon, Lisbon, Portugal

    Cláudio M. Gomes

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Saijo, E. et al. (2019). Ultrasensitive RT-QuIC Seed Amplification Assays for Disease-Associated Tau, α-Synuclein, and Prion Aggregates. In: Gomes, C. (eds) Protein Misfolding Diseases. Methods in Molecular Biology, vol 1873. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8820-4_2

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  • DOI: https://doi.org/10.1007/978-1-4939-8820-4_2

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  • Publisher Name: Humana Press, New York, NY

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