{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T12:06:31Z","timestamp":1790165191217,"version":"4.1.0"},"reference-count":45,"publisher":"Wiley","license":[{"start":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T00:00:00Z","timestamp":1790121600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T00:00:00Z","timestamp":1790121600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/100000038","name":"U.S. Food and Drug Administration","doi-asserted-by":"publisher","award":["75F40122C00200"],"award-info":[{"award-number":["75F40122C00200"]}],"id":[{"id":"10.13039\/100000038","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Biotech &amp; Bioengineering"],"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>Continuous downstream processing of messenger ribonucleic acid (mRNA) therapeutics requires purification strategies that are scalable, robust, and capable of removing product\u2010 and process\u2010related impurities from in vitro transcription mixtures. Precipitation\u2010based purification coupled with tangential flow filtration (TFF) has recently been demonstrated as a chromatography\u2010free route for continuous mRNA purification. However, implementation of this approach requires real\u2010time monitoring strategies that can distinguish the product\u2010rich precipitated phase from the impurity\u2010rich liquid phase, as well as phase\u2010dependent transport behavior across the integrated process. In this work, we develop a phase\u2010resolved process analytical technology (PAT) and residence time distribution (RTD) framework for continuous mRNA purification by PEG\/NaCl precipitation and sequential TFF. Inline UV\u2013Vis spectroscopy and LED\u2010based optical density measurements were used concurrently to separate nucleic\u2010acid absorbance from turbidity\u2010driven responses associated with suspended mRNA precipitates. RTD experiments with dissolved and precipitated tracers revealed phase\u2010dependent transport behavior in both the tubular precipitator and TFF module, which was quantified using axial dispersion and tanks\u2010in\u2010series models. During integrated operation, inline PAT captured startup, steady\u2010state, and washout behavior, while stable TMP indicated operation without observable membrane fouling. This work demonstrates that precipitation\u2010based continuous mRNA purification can be strengthened through phase\u2010resolved monitoring and RTD\u2010based process understanding, providing a framework for robust development of continuous downstream processes for nucleic acid therapeutics.<\/jats:p>","DOI":"10.1002\/bit.70372","type":"journal-article","created":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T11:48:12Z","timestamp":1790164092000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Process Analytical Technology (PAT) for Continuous Purification of mRNA via Precipitation and Tangential Flow Filtration"],"prefix":"10.1002","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-8536-2120","authenticated-orcid":false,"given":"Rajan R.","family":"Bhawnani","sequence":"first","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9336-2953","authenticated-orcid":false,"given":"Maria del Carme Pons","family":"Royo","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rohan","family":"Kadambi","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew","family":"Hatas","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Steve","family":"Burcat","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-9131-1958","authenticated-orcid":false,"given":"Tyler","family":"Arnold","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hasan","family":"Al\u2010Mahayni","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4304-3484","authenticated-orcid":false,"given":"Richard D.","family":"Braatz","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7468-8093","authenticated-orcid":false,"given":"Allan S.","family":"Myerson","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering Massachusetts Institute of Technology Cambridge Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2026,9,23]]},"reference":[{"key":"e_1_2_8_2_1","doi-asserted-by":"crossref","first-page":"1215","DOI":"10.1002\/bit.23023","article-title":"Real Time Monitoring of Multiple Parameters in Mammalian Cell Culture Bioreactors Using an In\u2010Line Raman Spectroscopy Probe","volume":"108","author":"Abu\u2010Absi N. 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