{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,14]],"date-time":"2026-10-14T04:09:17Z","timestamp":1791950957554,"version":"4.3.4"},"reference-count":38,"publisher":"American Chemical Society (ACS)","issue":"8","license":[{"start":{"date-parts":[[2026,7,22]],"date-time":"2026-07-22T00:00:00Z","timestamp":1784678400000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-029"},{"start":{"date-parts":[[2026,7,22]],"date-time":"2026-07-22T00:00:00Z","timestamp":1784678400000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-037"},{"start":{"date-parts":[[2026,7,22]],"date-time":"2026-07-22T00:00:00Z","timestamp":1784678400000},"content-version":"stm-asf","delay-in-days":0,"URL":"https:\/\/doi.org\/10.15223\/policy-045"}],"funder":[{"DOI":"10.13039\/501100007280","name":"Srinakharinwirot University","doi-asserted-by":"publisher","award":["102\/2568"],"award-info":[{"award-number":["102\/2568"]}],"id":[{"id":"10.13039\/501100007280","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,8,14]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>To decentralize tuberculosis surveillance in resource-limited and tropical regions, a rapid, robust, and cold-chain-independent near-point-of-care biosensor is urgently required to overcome the instability of conventional liquid-formulation diagnostics. Herein, we developed a ready-to-use lyophilized reaction mix Loop-Mediated Isothermal Amplification (LAMP) matrix integrated with a Lateral Flow Biosensor (LFB) for the instrument-free visual detection of Mycobacterium tuberculosis (M. tuberculosis). The freeze-dried reagents, stabilized with an optimized trehalose-BSA matrix, were hermetically packaged and systematically evaluated for long-term thermal stability across four regulated profiles (\u221220 \u00b0C, 4 \u00b0C, 25 \u00b0C, and 37 \u00b0C) over a 12-month period. Diagnostic viability was further validated in a double-blinded study using 191 clinical sputum specimens against gold-standard culture and line probe assays. The stabilized platform demonstrated an analytical sensitivity down to 102 CFU\/mL and maintained full functional activity for up to 12 months at \u221220 \u00b0C and 4 weeks at 4 \u00b0C, with robust short-term resilience under tropical stress (25 \u00b0C) for up to 1 week. However, the formulation was unable to withstand prolonged exposure to harsh environmental conditions at 37 \u00b0C, resulting in a progressive decline in amplification performance following phase-transition instability. Crucially, clinical validation yielded an outstanding diagnostic sensitivity of 98.92% and a positive predictive value (PPV) of 100.00%. These findings validate the lyophilized LAMP-LFB platform as a highly reliable, sustainable, and distributable molecular diagnostic tool capable of overcoming the cold-chain bottleneck in field-based tuberculosis detection. Although the developed platform shows strong potential for near-point-of-care deployment, further optimization of the lyophilization formulation and process will be necessary to enable prolonged storage and transportation under ambient-temperature conditions.<\/jats:p>","DOI":"10.1021\/acsinfecdis.6c00520","type":"journal-article","created":{"date-parts":[[2026,7,23]],"date-time":"2026-07-23T02:23:31Z","timestamp":1784773411000},"page":"2853-2862","source":"Crossref","is-referenced-by-count":1,"title":["Lyophilized LAMP Assay Combined with Lateral Flow\nBiosensor for Cold-Chain-Free Detection of\n                    <i>Mycobacterium tuberculosis<\/i>"],"prefix":"10.1021","volume":"12","author":[{"given":"Jutturong","family":"Ckumdee","sequence":"first","affiliation":[{"name":"Navamindradhiraj University , , ,","place":["Bangkok, Thailand, 10300"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Somchai","family":"Santiwatanakul","sequence":"additional","affiliation":[{"name":"Srinakharinwirot University , , , ,","place":["Bangkok, Nakhon Nayok Province, Thailand, 26120"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-6422-2659","authenticated-orcid":true,"given":"Nakarin","family":"Vutipow","sequence":"additional","affiliation":[{"name":"Kasetsart University , , ,","place":["Bangkok, Thailand, 10900"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0645-6450","authenticated-orcid":true,"given":"Thongchai","family":"Kaewphinit","sequence":"additional","affiliation":[{"name":"Srinakharinwirot University , , ,","place":["Bangkok, Thailand, 10110"]},{"name":"Srinakharinwirot University , , ,","place":["Bangkok, Thailand, 10110"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,7,22]]},"reference":[{"key":"2026081410072609300_cit1","doi-asserted-by":"publisher","first-page":"233","DOI":"10.1596\/978-1-4648-0524-0_ch11","article-title":"Tuberculosis","volume-title":"Disease Control Priorities: major Infectious Diseases","author":"Bloom","year":"2017","edition":"3rd"},{"key":"2026081410072609300_cit2","doi-asserted-by":"publisher","first-page":"S7","DOI":"10.1016\/j.ijid.2021.02.107","article-title":"Global Tuberculosis\nReport 2020 \u2013 Reflections on the Global\nTB burden, treatment and prevention efforts","volume":"113","author":"Chakaya","year":"2021","journal-title":"Int. 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J."},{"issue":"9","key":"2026081410072609300_cit6","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0099659","article-title":"Evaluation\nof BACTEC MGIT 960 System for Testing Susceptibility\nof Mycobacterium tuberculosis to First-Line Drugs in China","volume":"9","author":"Zhao","year":"2014","journal-title":"PLoS One"},{"issue":"11","key":"2026081410072609300_cit7","doi-asserted-by":"publisher","first-page":"1005","DOI":"10.1056\/NEJMoa0907847","article-title":"Rapid Molecular Detection of Tuberculosis\nand Rifampin Resistance","volume":"363","author":"Boehme","year":"2010","journal-title":"N. Engl. J. 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Biopharm."},{"issue":"7","key":"2026081410072609300_cit15","doi-asserted-by":"publisher","first-page":"1427","DOI":"10.1002\/jps.20364","article-title":"Mechanism\nof protein stabilization by sugars during freeze-drying and storage:\nNative structure preservation, specific interaction, and\/or immobilization\nin a glassy matrix?","volume":"94","author":"Chang","year":"2005","journal-title":"J. Pharm. Sci."},{"issue":"28","key":"2026081410072609300_cit16","doi-asserted-by":"publisher","first-page":"3747","DOI":"10.1039\/c4cc00540f","article-title":"Simultaneous elimination of carryover\ncontamination\nand detection of DNA with uracil-DNA-glycosylase-supplemented loop-mediated\nisothermal amplification (UDG-LAMP)","volume":"50","author":"Hsieh","year":"2014","journal-title":"Chem. Commun."},{"issue":"1","key":"2026081410072609300_cit17","doi-asserted-by":"publisher","first-page":"18","DOI":"10.3390\/bios3010018","article-title":"Isothermal Amplification\nMethods for the Detection of Nucleic Acids\nin Microfluidic Devices","volume":"3","author":"Zanoli","year":"2013","journal-title":"Biosensors"},{"issue":"4","key":"2026081410072609300_cit18","doi-asserted-by":"publisher","first-page":"2087","DOI":"10.1016\/S0006-3495(96)79407-9","article-title":"Is trehalose\nspecial for preserving\ndry biomaterials?","volume":"71","author":"Crowe","year":"1996","journal-title":"Biophys. J."},{"issue":"2","key":"2026081410072609300_cit19","doi-asserted-by":"publisher","first-page":"1249","DOI":"10.1021\/acs.jctc.9b01071","article-title":"Properties of Aqueous Trehalose Mixtures:\nGlass Transition and Hydrogen Bonding","volume":"16","author":"Olgenblum","year":"2020","journal-title":"J. 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Spectrum."},{"issue":"3","key":"2026081410072609300_cit24","doi-asserted-by":"publisher","first-page":"10","DOI":"10.1128\/JCM.02314-20","article-title":"Xpert MTB\/XDR: A 10-Color Reflex Assay Suitable\nfor Point-of-Care Settings To Detect Isoniazid, Fluoroquinolone, and\nSecond-Line-Injectable-Drug Resistance Directly from Mycobacterium\ntuberculosis-Positive Sputum","volume":"59","author":"Cao","year":"2021","journal-title":"J. Clin. Microbiol."},{"issue":"2","key":"2026081410072609300_cit25","doi-asserted-by":"publisher","first-page":"242","DOI":"10.1016\/S1473-3099(21)00452-7","article-title":"Detection of isoniazid, fluoroquinolone, ethionamide,\namikacin, kanamycin, and capreomycin resistance by the Xpert MTB\/XDR\nassay: A cross-sectional multicentre diagnostic accuracy study","volume":"22","author":"Penn-Nicholson","year":"2022","journal-title":"Lancet Infect. 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Mycobact."},{"issue":"6","key":"2026081410072609300_cit28","doi-asserted-by":"publisher","first-page":"1936","DOI":"10.1128\/JCM.02352-06","article-title":"Operational\nFeasibility\nof Using Loop-Mediated Isothermal Amplification for Diagnosis of Pulmonary\nTuberculosis in Microscopy Centers of Developing Countries","volume":"45","author":"Boehme","year":"2007","journal-title":"J. Clin. Microbiol."},{"issue":"1","key":"2026081410072609300_cit29","doi-asserted-by":"publisher","first-page":"268","DOI":"10.1186\/s12879-019-3881-y","article-title":"Diagnostic\naccuracy of TB-LAMP for pulmonary tuberculosis: A systematic review\nand meta-analysis","volume":"19","author":"Shete","year":"2019","journal-title":"BMC Infect. Dis."},{"key":"2026081410072609300_cit30","doi-asserted-by":"publisher","first-page":"107108","DOI":"10.1016\/j.microc.2021.107108","article-title":"Identification of S315T mutation\nin katG gene using probe-free exclusive mismatch primers for a rapid\ndiagnosis of isoniazid-resistant Mycobacterium tuberculosis by real-time\nloop-mediated isothermal amplification","volume":"175","author":"Khumwan","year":"2022","journal-title":"Microchem.\nJ."},{"issue":"3","key":"2026081410072609300_cit31","doi-asserted-by":"publisher","first-page":"626","DOI":"10.1111\/jam.13647","article-title":"Loop-mediated\nisothermal amplification (LAMP): A versatile technique for detection\nof micro-organisms","volume":"124","author":"Wong","year":"2018","journal-title":"J. Appl. Microbiol."},{"issue":"5","key":"2026081410072609300_cit32","doi-asserted-by":"publisher","first-page":"877","DOI":"10.1038\/nprot.2008.57","article-title":"Loop-mediated isothermal\namplification (LAMP) of gene sequences and simple visual detection\nof products","volume":"3","author":"Tomita","year":"2008","journal-title":"Nat. Protoc."},{"key":"2026081410072609300_cit33","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1155\/2013\/926230","article-title":"Detection ofMycobacterium\ntuberculosisby\nUsing Loop-Mediated Isothermal Amplification Combined with a Lateral\nFlow Dipstick in Clinical Samples","volume":"2013","author":"Kaewphinit","year":"2013","journal-title":"BioMed. Res.\nInt."},{"key":"2026081410072609300_cit34","doi-asserted-by":"publisher","first-page":"192","DOI":"10.3389\/fmicb.2017.00192","article-title":"Loop-Mediated\nIsothermal Amplification Label-Based Gold Nanoparticles Lateral Flow\nBiosensor for Detection of Enterococcus faecalis and Staphylococcus\naureus","volume":"8","author":"Wang","year":"2017","journal-title":"Front. Microbiol."},{"issue":"2","key":"2026081410072609300_cit35","doi-asserted-by":"publisher","first-page":"59","DOI":"10.2144\/000114253","article-title":"Visual Detection of Isothermal Nucleic Acid Amplification\nUsing pH-Sensitive\nDyes","volume":"58","author":"Tanner","year":"2015","journal-title":"BioTechniques"},{"issue":"2","key":"2026081410072609300_cit36","doi-asserted-by":"publisher","first-page":"199","DOI":"10.1248\/cpb.52.199","article-title":"Protection of Protein Secondary Structure\nby Saccharides of Different Molecular Weights during Freeze-Drying","volume":"52","author":"Izutsu","year":"2004","journal-title":"Chem. Pharm. Bull."},{"issue":"1","key":"2026081410072609300_cit37","doi-asserted-by":"publisher","first-page":"24","DOI":"10.1002\/pro.3","article-title":"Effect of trehalose\non protein structure","volume":"18","author":"Jain","year":"2009","journal-title":"Protein\nSci."},{"issue":"19","key":"2026081410072609300_cit38","doi-asserted-by":"publisher","first-page":"3957","DOI":"10.1093\/nar\/25.19.3957","article-title":"Betaine\nImproves the PCR Amplification of GC-Rich DNA Sequences","volume":"25","author":"Henke","year":"1997","journal-title":"Nucleic Acids Res."}],"container-title":["ACS Infectious Diseases"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/acsinfecdis.6c00520","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/aidcbc\/article-pdf\/12\/8\/2853\/66177858\/acsinfecdis.6c00520.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/aidcbc\/article-pdf\/12\/8\/2853\/66177858\/acsinfecdis.6c00520.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T14:22:01Z","timestamp":1786717321000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/aidcbc\/article\/12\/8\/2853\/5228606\/Lyophilized-LAMP-Assay-Combined-with-Lateral-Flow"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,22]]},"references-count":38,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2026,7,22]]},"published-print":{"date-parts":[[2026,8,14]]}},"URL":"https:\/\/doi.org\/10.1021\/acsinfecdis.6c00520","relation":{},"ISSN":["2373-8227"],"issn-type":[{"value":"2373-8227","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,22]]}}}