{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,7]],"date-time":"2026-09-07T11:11:22Z","timestamp":1788779482410,"version":"build-2803163510"},"reference-count":40,"publisher":"Wiley","issue":"20","license":[{"start":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T00:00:00Z","timestamp":1785369600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T00:00:00Z","timestamp":1785369600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"publisher","award":["GM156406"],"award-info":[{"award-number":["GM156406"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Rapid Comm Mass Spectrometry"],"published-print":{"date-parts":[[2026,10,30]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Rationale<\/jats:title>\n                    <jats:p>Multiplexed quantitative proteomics enables simultaneous analysis of multiple biological samples, increasing throughput while reducing instrument time and sample requirements. However, integrating sample multiplexing with data\u2010independent acquisition (DIA) remains challenging. We present a TMTpro plexDIA strategy leveraging MS1\u2010level mass differences between nonisobaric TMTpro reagent variants to enable multiplexed quantification without compromising DIA sensitivity.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Three\n                      <jats:styled-content style=\"fixed-case\">\n                        <jats:italic>Saccharomyces cerevisiae<\/jats:italic>\n                      <\/jats:styled-content>\n                      deletion strains (\n                      <jats:italic>\u0394met6<\/jats:italic>\n                      ,\n                      <jats:italic>\u0394pfk2<\/jats:italic>\n                      , and\n                      <jats:italic>\u0394ura2<\/jats:italic>\n                      ) were labeled with TMTproZero (light), TMTpro16 (standard), and super\u2010heavy TMTpro (heavy), mixed in three permutations, and analyzed by narrow\u2010window DIA (2\u2009\n                      <jats:italic>m<\/jats:italic>\n                      \/\n                      <jats:italic>z<\/jats:italic>\n                      isolation, 300 scan events) on an Orbitrap Astral mass spectrometer. Database searching was performed using FragPipe\/MSFragger with plex\u2010DIA quantification enabled.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>Over 2000 protein groups and over 20\u2009000 precursors were identified per channel per mixture, with closely matched identification rates across all three channels. All nine expected deletion patterns were correctly identified, with channel\u2010specific depletion reproduced consistently across precursor charge states (2+, 3+, and 4+).<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>TMTpro plex\u2010DIA enables accurate, multiplexed quantitative proteomics through MS1\u2010level mass separation of the nonisobaric TMTpro isotopologs. The characteristic deletion patterns observed for each knockout strain serve as intrinsic molecular barcodes, validating sample identity and demonstrating the broad utility of plex\u2010DIA for high\u2010throughput, multiplexed proteomics applications.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1002\/rcm.70146","type":"journal-article","created":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T09:41:23Z","timestamp":1785404483000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Implementation of Nonisobaric TMT Analogs for Accurate Precursor\u2010Level Quantification by plexDIA"],"prefix":"10.1002","volume":"40","author":[{"given":"Ting\u2010Yu","family":"Wei","sequence":"first","affiliation":[{"name":"Department of Cell Biology Harvard Medical School  Boston Massachusetts 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