{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,14]],"date-time":"2026-10-14T18:42:31Z","timestamp":1792003351705,"version":"4.4.0"},"reference-count":32,"publisher":"Public Library of Science (PLoS)","issue":"6","license":[{"start":{"date-parts":[[2025,6,25]],"date-time":"2025-06-25T00:00:00Z","timestamp":1750809600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>Seeding amplification assays, specifically the Real-Time Quaking-Induced Conversion method (RT-QuIC), have shown great diagnostic potential for \u03b1-synucleinopathies. Numerous research groups have demonstrated the method\u2019s high sensitivity and specificity using cerebrospinal fluid (CSF) samples and various RT-QuIC workflows. However, establishing a uniform and stably performing RT-QuIC protocol remains challenging. To address this, we established an RT-QuIC protocol with a Universal Control Fluid (UCF), which is simple to adopt, performs stably, and allows uniform preparation of both sample and control reactions. Firstly, we adapted and established a published 48-hour RT-QuIC protocol, including the in-house production of recombinant \u03b1-synuclein (rec \u03b1-syn), and evaluated its sensitivity and specificity through a blinded screening of an 81 CSF sample cohort consisting of Parkinson\u2019s disease (PD), dementia with Lewy bodies (DLB), Alzheimer\u2019s disease, motor neuron disease, multiple system atrophy, unidentified neurodegenerative diseases, and healthy controls. Additionally, we tested all CSF samples in three volumes to determine which volume provides the best diagnostic accuracy. The established RT-QuIC performs nearly equally well with 7 \u00b5L and 15 \u00b5L added CSF, resulting in 94% and 94.5% diagnostic accuracy, respectively. Secondly, we developed a UCF solution and tested its performance with the established RT-QuIC protocol. Results indicate that UCF, used in defined volume and concentration, standardizes the preparation of both sample and control reactions without compromising the assay\u2019s diagnostic accuracy and provides a stabilizing environment for the reactions, ensuring higher reproducibility. The established RT-QuIC protocol for pathologic \u03b1-synuclein detection in PD and DLB CSF samples is highly sensitive (92\u201396%) and specific (93\u201396%). Therefore, it is important that its adoption in clinical laboratories is uncomplicated and uniform. RT-QuIC with UCF simplifies, standardizes, and stabilizes the assay\u2019s performance and, thus, could be recommended as a standard protocol for accurate detection of \u03b1-synucleinopathies.<\/jats:p>","DOI":"10.1371\/journal.pone.0326568","type":"journal-article","created":{"date-parts":[[2025,6,25]],"date-time":"2025-06-25T18:15:52Z","timestamp":1750875352000},"page":"e0326568","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":7,"title":["Seeding amplification assay with Universal Control Fluid: Standardized detection of \u03b1-synucleinopathies"],"prefix":"10.1371","volume":"20","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-6574-8093","authenticated-orcid":true,"given":"Remarh","family":"Bsoul","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anja 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