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Previously, we identified a novel immune biomarker <jats:italic>IFI27<\/jats:italic>, validated with routine RT-qPCR methods employed in a research setting, that discriminates between influenza and bacteria in patients with suspected respiratory infection. This study aimed to assess the <jats:bold>\n              <jats:italic>In<\/jats:italic>\n            <\/jats:bold>\n            <jats:italic>Signia<\/jats:italic> method, which employs a novel gene normalization technique to yield a variable transcript analysis (VITA) index. The VITA index measures gene expression relative to a non-transcribed region of DNA, such that it is independent of sample quality or quantity. We compared <jats:italic>IFI27<\/jats:italic> gene expression measured by the <jats:bold>\n              <jats:italic>In<\/jats:italic>\n            <\/jats:bold>\n            <jats:italic>Signia<\/jats:italic> assay to that of the research assay in blood samples collected from patients with respiratory diseases and SARS-CoV-2 vaccinated individuals. The study found a strong correlation and acceptable agreement between traditional \u0394Cq methods and <jats:bold>\n              <jats:italic>In<\/jats:italic>\n            <\/jats:bold>\n            <jats:italic>Signia<\/jats:italic> for <jats:italic>IFI27<\/jats:italic> levels in the higher range (log(\u0394Cq)<jats:sub>Research<\/jats:sub>\u00a0&gt;\u00a01), but not for <jats:italic>IFI27<\/jats:italic> expression levels below this range, likely due to the different normalization strategies. Notably the <jats:bold>\n              <jats:italic>In<\/jats:italic>\n            <\/jats:bold>\n            <jats:italic>Signia<\/jats:italic> assay was more sensitive in detecting viral infection among hospital patients. These findings suggest that the <jats:bold>\n              <jats:italic>In<\/jats:italic>\n            <\/jats:bold>\n            <jats:italic>Signia<\/jats:italic> assay, which supports high throughput workflows, may be used for the rapid detection of viral infection in patients with respiratory symptoms.<\/jats:p>","DOI":"10.1038\/s41598-025-04688-9","type":"journal-article","created":{"date-parts":[[2025,7,1]],"date-time":"2025-07-01T17:44:32Z","timestamp":1751391872000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Evaluating the InSignia IFI27 expression assay for detecting viral respiratory infection compared to a traditional gene normalisation assay"],"prefix":"10.1038","volume":"15","author":[{"given":"Tiana M.","family":"Pelaia","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Karan","family":"Kim","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicole","family":"Lima","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Claire","family":"Gibbs","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lee M.","family":"Smith","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sally","family":"Teoh","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ya","family":"Wang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Colin","family":"Denver","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elisa","family":"Mokany","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alison","family":"Todd","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sam","family":"Orde","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Benjamin","family":"Tang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Anthony","family":"McLean","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Maryam","family":"Shojaei","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2025,7,1]]},"reference":[{"key":"4688_CR1","doi-asserted-by":"publisher","first-page":"1100352","DOI":"10.3389\/fgene.2023.1100352","volume":"14","author":"M Tsakiroglou","year":"2023","unstructured":"Tsakiroglou, M., Evans, A. & Pirmohamed, M. 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