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Multiplex molecular point-of-care testing (mPOCT) enables automated and rapid differential diagnosis for timely intervention. This study independently evaluated the clinical performance of Coyote FlashDetect LyocartE SARS-CoV-2\/FluA\/FluB and FlashDetect LyocartE RSV\/MP\/ADV multiplex mPOCT assays using the Sansure as the primary comparator, followed by a subset-based, target-specific paired comparison with Cepheid mPOCT assay for selected respiratory pathogens (FluA, FluB, and RSV). Specifically, 642 and 427 throat swab specimens were used to evaluate the Coyote SARS-CoV-2\/FluA\/FluB and Coyote RSV\/MP\/ADV multiplex mPOCT assays against Sansure assays, respectively. Coyote assays demonstrated high concordance with Sansure assays, with overall percent agreement (OPA) values ranging from 93.8% to 100.0% for six pathogens covered by two Coyote assays. Most discrepancies with Sansure assays occurred in samples with cycle threshold (Ct) values \u226535, especially among FluA samples. Moreover, 414, 414, and 215 specimens were allocated for FluA, FluB, and RSV, respectively, to conduct target-specific paired comparison between Coyote and Cepheid. Each Coyote assay also demonstrated high concordance with the Cepheid assays, with PPA values of 96.0%, 100.0%, and 98.3% for FluA, FluB, and RSV, respectively. These results support the use of Coyote mPOCT assays in specific settings. Coyote assays demonstrated a high degree of concordance for shared targets with comparator methods, supporting their use as a rapid and user-friendly diagnostic approach for multiplex respiratory pathogen detection.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>This study highlights the value of multiplex molecular point-of-care tests. The POCT assays demonstrate high concordance with the laboratory-based lab tests in this retrospective specimen cohort, offering a viable, rapid, and user-friendly alternative for multiplex respiratory pathogen detection and hold the promise to be used in primary healthcare settings and public health surveillance.<\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.03236-25","type":"journal-article","created":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T13:03:11Z","timestamp":1789650191000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Multicenter evaluation of Coyote FlashDetect LyocartE multiplex assays for six respiratory pathogens: a comparative analysis with Sansure and Cepheid assays"],"prefix":"10.1128","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3936-9580","authenticated-orcid":false,"given":"Cong","family":"Liu","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02jwb5s28","id-type":"ROR","asserted-by":"publisher"}],"name":"National Center for Clinical Laboratories, Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing Hospital\/ National Center of Gerontology","place":["Beijing, P. 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