{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,29]],"date-time":"2026-10-29T01:59:21Z","timestamp":1793239161733,"version":"4.5.0"},"reference-count":30,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2016,5,23]],"date-time":"2016-05-23T00:00:00Z","timestamp":1463961600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Foundation Alliance Biosecure"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Rapid and sensitive detection of prions is important in managing prion diseases. The real-time quaking-induced conversion (RT-QuIC) assay for prion seeding activity has been applied to many prion diseases and provides for specific antemortem diagnostic testing. We evaluated RT-QuIC\u2019s long-term consistency and varied multiple reaction parameters. Repeated assays of a single scrapie sample using multiple plate readers and recombinant prion protein (rPrPSen) substrates gave comparable results. N-terminal truncated hamster rPrPSen (residues 90\u2013231) hastened both prion-seeded and prion-independent reactions but maintained a clear kinetic distinction between the two. Raising temperatures or shaking speeds accelerated RT-QuIC reactions without compromising specificity. When applied to nasal brushings from Creutzfeldt-Jakob disease patients, higher temperatures accelerated RT-QuIC kinetics, and the use of hamster rPrPSen (90\u2013231) strengthened RT-QuIC responses. Elongation of shaking periods reduced scrapie-seeded reaction times, but continuous shaking promoted false-positive reactions. Furthermore, pH 7.4 provided for more rapid RT-QuIC reactions than more acidic pHs. Additionally, we show that small variations in the amount of sodium dodecyl sulfate (SDS) significantly impacted the assay. Finally, RT-QuIC performed in multiplate thermoshakers followed by fluorescence readings in separate plate readers enhanced assay throughput economically. Collectively, these results demonstrate improved speed, efficacy and practicality of RT-QuIC assays and highlight variables to be optimized for future applications.<\/jats:p>","DOI":"10.3390\/v8050140","type":"journal-article","created":{"date-parts":[[2016,5,24]],"date-time":"2016-05-24T09:05:05Z","timestamp":1464080705000},"page":"140","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":85,"title":["Factors That Improve RT-QuIC Detection of Prion Seeding Activity"],"prefix":"10.3390","volume":"8","author":[{"given":"Christina","family":"Orr\u00fa","sequence":"first","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew","family":"Hughson","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5059-7672","authenticated-orcid":false,"given":"Bradley","family":"Groveman","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katrina","family":"Campbell","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kelsie","family":"Anson","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matteo","family":"Manca","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Allison","family":"Kraus","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Byron","family":"Caughey","sequence":"additional","affiliation":[{"name":"Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Hamilton, 59840 MT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2016,5,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"543","DOI":"10.1146\/annurev-micro-092412-155735","article-title":"Prions and the potential transmissibility of protein misfolding diseases","volume":"67","author":"Kraus","year":"2013","journal-title":"Annu. 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