{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T20:50:27Z","timestamp":1792788627937,"version":"4.5.0"},"update-to":[{"DOI":"10.1371\/journal.ppat.1012554","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2024,10,4]],"date-time":"2024-10-04T00:00:00Z","timestamp":1728000000000}}],"reference-count":65,"publisher":"Public Library of Science (PLoS)","issue":"9","license":[{"start":{"date-parts":[[2024,9,20]],"date-time":"2024-09-20T00:00:00Z","timestamp":1726790400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/publicdomain\/zero\/1.0\/"}],"funder":[{"DOI":"10.13039\/100006492","name":"Division of Intramural Research, National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["ZIAAI001086"],"award-info":[{"award-number":["ZIAAI001086"]}],"id":[{"id":"10.13039\/100006492","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000038","name":"Natural Sciences and Engineering Research Council of Canada","doi-asserted-by":"publisher","award":["Discovery Grant 2022-03744"],"award-info":[{"award-number":["Discovery Grant 2022-03744"]}],"id":[{"id":"10.13039\/501100000038","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Interdisciplinary Theoretical and Mathematical Sciences programme at RIKEN"},{"DOI":"10.13039\/100000049","name":"National Institute on Aging","doi-asserted-by":"publisher","award":["P30 AG072976"],"award-info":[{"award-number":["P30 AG072976"]}],"id":[{"id":"10.13039\/100000049","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Indiana Alzheimer\u2019s Disease Research Center","award":["AGO5131"],"award-info":[{"award-number":["AGO5131"]}]}],"content-domain":{"domain":["www.plospathogens.org"],"crossmark-restriction":false},"short-container-title":["PLoS Pathog"],"abstract":"<jats:p>\n                    Disease associated pathological aggregates of alpha-synuclein (\u03b1Syn\n                    <jats:sup>D<\/jats:sup>\n                    ) exhibit prion-like spreading in synucleinopathies such as Parkinson\u2019s disease (PD) and dementia with Lewy bodies (DLB). Seed amplification assays (SAAs) such as real-time quaking-induced conversion (RT-QuIC) have shown high diagnostic sensitivity and specificity for detecting proteopathic \u03b1Syn\n                    <jats:sup>D<\/jats:sup>\n                    seeds in a variety of biospecimens from PD and DLB patients. However, the extent to which relative proteopathic seed concentrations are useful as indices of a patient\u2019s disease stage or prognosis remains unresolved. One feature of current SAAs that complicates attempts to correlate SAA results with patients\u2019 clinical and other laboratory findings is their quantitative imprecision, which has typically been limited to discriminating large differences (e.g. 5\u201310 fold) in seed concentration. We used end-point dilution (ED) RT-QuIC assays to determine \u03b1Syn\n                    <jats:sup>D<\/jats:sup>\n                    seed concentrations in patient biospecimens and tested the influence of various assay variables such as serial dilution factor, replicate number and data processing methods. The use of 2-fold versus 10-fold dilution factors and 12 versus 4 replicate reactions per dilution reduced ED-RT-QuIC assay error by as much as 70%. This enhanced assay format discriminated as little as 2-fold differences in \u03b1Syn\n                    <jats:sup>D<\/jats:sup>\n                    seed concentration besides detecting ~2-16-fold seed reductions caused by inactivation treatments. In some scenarios, analysis of the data using Poisson and midSIN algorithms provided more consistent and statistically significant discrimination of different seed concentrations. We applied our improved assay strategies to multiple diagnostically relevant PD and DLB antemortem patient biospecimens, including cerebrospinal fluid, skin, and brushings of the olfactory mucosa. Using ED \u03b1Syn RT-QuIC as a model SAA, we show how to markedly improve the inter-assay reproducibility and quantitative accuracy. Enhanced quantitative SAA accuracy should facilitate assessments of pathological seeding activities as biomarkers in proteinopathy diagnostics and prognostics, as well as in patient cohort selection and assessments of pharmacodynamics and target engagement in drug trials.\n                  <\/jats:p>","DOI":"10.1371\/journal.ppat.1012554","type":"journal-article","created":{"date-parts":[[2024,9,20]],"date-time":"2024-09-20T13:37:55Z","timestamp":1726839475000},"page":"e1012554","update-policy":"https:\/\/doi.org\/10.1371\/journal.ppat.corrections_policy","source":"Crossref","is-referenced-by-count":32,"title":["Enhanced quantitation of pathological \u03b1-synuclein in patient biospecimens by RT-QuIC seed amplification assays"],"prefix":"10.1371","volume":"20","author":[{"given":"Ankit","family":"Srivastava","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qinlu","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christina D.","family":"Orr\u00f9","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manel","family":"Fernandez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yaroslau","family":"Compta","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bernardino","family":"Ghetti","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gianluigi","family":"Zanusso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wen-Quan","family":"Zou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3928-6101","authenticated-orcid":true,"given":"Byron","family":"Caughey","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Catherine A. 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