{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,31]],"date-time":"2026-08-31T14:02:03Z","timestamp":1788184923149,"version":"build-2803163510"},"reference-count":30,"publisher":"Wiley","issue":"12","license":[{"start":{"date-parts":[[2024,9,18]],"date-time":"2024-09-18T00:00:00Z","timestamp":1726617600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000265","name":"Medical Research Council","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100000265","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2024,12]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Low availability of routine nucleic acid amplification testing (NAAT) during infection outbreaks, especially in less resourced environments, was highlighted by the Covid pandemic. One of the barriers lies with the supply chain and cost of the active diagnostic ingredients (ADIs) that are the reagents for NAATs. This work explores a novel synthesis method to produce a key NAAT reagent, namely the 2\u2032\u2010deoxynucleoside 5\u2032\u2010triphosphate (dNTPs), via a reusable enzyme bioreactor, that can be integrated into a NAAT workflow. A self\u2010immobilizing R5\u2010silaffin kinase fusion enzyme was designed for immobilization on silica, converting dNMPs to their respective dNTP ADIs for PCR in a R5\u2010kinase mini\u2010bioreactor, designed to be implemented in a reusable device, stable over 2 months, when stored at 4\u00b0C. The performance is demonstrated for PCR reactions of the lambda genome and showed successful amplification up to 7.5\u2009kb. In comparison with commercial dNTPs, in <jats:italic>Plasmodium malariae<\/jats:italic> NAATs, a high linear correlation was shown between the <jats:italic>C<jats:sub>t<\/jats:sub><\/jats:italic> value and the log(Copy Number), with lower incidence of false positives than with the commercial dNTPs. Overall a pathway to generate deoxynucleotides from monophosphate precursors was demonstrated, and an immobilized enzyme mini\u2010bioreactor investigated as a proof\u2010of\u2010principle for work\u2010flow integration with NAAT in low\u2010resource research and diagnostics labs.<\/jats:p>","DOI":"10.1002\/bit.28837","type":"journal-article","created":{"date-parts":[[2024,9,18]],"date-time":"2024-09-18T03:49:36Z","timestamp":1726631376000},"page":"3834-3847","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Active diagnostic ingredients (ADIs) for PCR: A mini\u2010bioreactor producing dNTPs with silica immobilized R5\u2010kinases"],"prefix":"10.1002","volume":"121","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0354-8824","authenticated-orcid":false,"given":"Anna R.","family":"Bird","sequence":"first","affiliation":[{"name":"Chemical Engineering and Biotechnology University of Cambridge Cambridge UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9572-9854","authenticated-orcid":false,"given":"Elizabeth A. 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