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We developed a multi-layered, paper-based microfluidic and miniaturized electrophoresis system for rapid nucleic acid extraction, separation, amplification, and detection, designed for resource-limited settings. Constructed from acrylic, transparency film, pressure-sensitive adhesion, and Whatman paper using a CO2 laser, the setup simplifies traditional methods and eliminates the need for complex equipment. DNA extraction and purification are achieved using Zweifach\u2013Fung bifurcation and Fahraeus effect principles, with detection via a hydrogel-assisted colorimetric isothermal reverse transcriptase-loop-mediated isothermal amplification technique. The system accurately identified the SARS-CoV-2 N-gene and \u03b2-actin human gene, validated by a compact electrophoresis setup. In clinical validation with 12 patient specimens, the system demonstrated a positive predictive agreement of 83.0% and a negative predictive agreement of 100%. The system achieves a limit of detection of 1 copy\/\u03bcl and can potentially transform nucleic acid detection assays in healthcare settings. This study addresses key challenges in nucleic acid detection, such as ensuring sample quality and quantity, reducing reliance on sophisticated equipment, preventing contamination, simplifying procedures, and providing rapid and accurate diagnostics for emerging pathogens.<\/jats:p>","DOI":"10.1063\/5.0211812","type":"journal-article","created":{"date-parts":[[2024,12,4]],"date-time":"2024-12-04T18:11:08Z","timestamp":1733335868000},"update-policy":"https:\/\/doi.org\/10.1063\/aip-crossmark-policy-page","source":"Crossref","is-referenced-by-count":2,"title":["Miniaturized electrophoresis: An integrated microfluidic cartridge with functionalized hydrogel-assisted LAMP for sample-to-answer analysis of nucleic acid"],"prefix":"10.1063","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3558-9201","authenticated-orcid":false,"given":"Natish","family":"Kumar","sequence":"first","affiliation":[{"name":"Department of Chemical Engineering, Indian Institute of Technology 1 , Jammu 181221,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-7691-495X","authenticated-orcid":false,"given":"Monika","family":"Kumari","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering, Indian Institute of Technology 1 , Jammu 181221,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9555-920X","authenticated-orcid":false,"given":"Devtulya","family":"Chander","sequence":"additional","affiliation":[{"name":"Fermentation and Microbial Biotechnology Division, CSIR\u2014Indian Institute of Integrative Medicine 2 , Canal Road, Jammu 180001,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8160-2518","authenticated-orcid":false,"given":"Sandeep","family":"Dogra","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Government Medical College 3 , Jammu 180001,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5514-5608","authenticated-orcid":false,"given":"Asha","family":"Chaubey","sequence":"additional","affiliation":[{"name":"Fermentation and Microbial Biotechnology Division, CSIR\u2014Indian Institute of Integrative Medicine 2 , Canal Road, Jammu 180001,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8124-0441","authenticated-orcid":false,"given":"Ravi Kumar","family":"Arun","sequence":"additional","affiliation":[{"name":"Department of Chemical Engineering, Indian Institute of Technology 1 , Jammu 181221,","place":["India"]}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"317","published-online":{"date-parts":[[2024,12,4]]},"reference":[{"key":"2024120418110318700_c1","doi-asserted-by":"publisher","first-page":"112592","DOI":"10.1016\/j.bios.2020.112592","article-title":"Advances in point-of-care nucleic acid extraction technologies for rapid diagnosis of human and plant diseases","volume":"169","year":"2020","journal-title":"Biosens. 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