دورية أكاديمية
Paper-based loop-mediated isothermal amplification and CRISPR integrated platform for on-site nucleic acid testing of pathogens.
| العنوان: | Paper-based loop-mediated isothermal amplification and CRISPR integrated platform for on-site nucleic acid testing of pathogens. |
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| المؤلفون: | Sen A; DNAiTECH Ltd, Marlborough Research Center, 2650 State Highway 1, Grovetown, Blenheim, Marlborough, 7202, New Zealand., Masetty M; Department of Chemical and Environmental Engineering, University of Cincinnati, Cincinnati, OH, 45221, USA., Weerakoon S; Department of Chemical and Environmental Engineering, University of Cincinnati, Cincinnati, OH, 45221, USA., Morris C; DNAiTECH Ltd, Marlborough Research Center, 2650 State Highway 1, Grovetown, Blenheim, Marlborough, 7202, New Zealand., Yadav JS; Department of Environmental & Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45221, USA., Apewokin S; Division of Infectious Diseases, College of Medicine, University of Cincinnati, Cincinnati, OH, 45221, USA., Trannguyen J; Division of Infectious Diseases, College of Medicine, University of Cincinnati, Cincinnati, OH, 45221, USA., Broom M; DNAiTECH Ltd, Marlborough Research Center, 2650 State Highway 1, Grovetown, Blenheim, Marlborough, 7202, New Zealand. Electronic address: Murray@dnaitech.com., Priye A; Department of Chemical and Environmental Engineering, University of Cincinnati, Cincinnati, OH, 45221, USA; Digital Futures, University of Cincinnati, Cincinnati, OH, 45221, USA. Electronic address: piryah@uc.edu. |
| المصدر: | Biosensors & bioelectronics [Biosens Bioelectron] 2024 Aug 01; Vol. 257, pp. 116292. Date of Electronic Publication: 2024 Apr 17. |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Elsevier Advanced Technology Country of Publication: England NLM ID: 9001289 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1873-4235 (Electronic) Linking ISSN: 09565663 NLM ISO Abbreviation: Biosens Bioelectron Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: Oxford : Elsevier Advanced Technology Original Publication: [Barking, Essex, England] : Elsevier Applied Science, 1989- |
| مواضيع طبية MeSH: | Nucleic Acid Amplification Techniques*/methods , Nucleic Acid Amplification Techniques*/instrumentation , Biosensing Techniques*/methods , Paper* , SARS-CoV-2*/genetics , SARS-CoV-2*/isolation & purification , COVID-19*/diagnosis , COVID-19*/virology , Molecular Diagnostic Techniques*/methods , Molecular Diagnostic Techniques*/instrumentation , CRISPR-Cas Systems*/genetics, Humans ; Limit of Detection ; Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; Equipment Design ; COVID-19 Nucleic Acid Testing/methods ; COVID-19 Nucleic Acid Testing/instrumentation ; Escherichia coli/genetics ; Escherichia coli/isolation & purification ; CRISPR-Associated Proteins/genetics ; Smartphone |
| مستخلص: | We report the development and initial validation of a paper-based nucleic acid testing platform that integrates Loop-mediated isothermal amplification (LAMP) with clustered regularly interspaced short palindromic repeats (CRISPR) technology, referred to as PLACID (Paper-based LAMP-CRISPR Integrated Diagnostics). LAMP eliminates the need for thermal cycling, resulting in simplified instrumentation, and the CRISPR-associated protein (Cas 12a) system eliminates false positive signals from LAMP products, resulting in highly selective and sensitive assays. We optimized the assay to perform both amplification and detection entirely on paper, eliminating the need for complex fluid handling steps and lateral flow assay transfers. Additionally, we engineered a smartphone-operated system that includes a low-powered, non-contact IR heating chamber to actuate paper-based LAMP and CRISPR reactions and enable the detection of fluorescent signals from the paper. The platform demonstrates high specificity and sensitivity in detecting nucleic acid targets with a limit of detection of 50 copies/μL. We integrate an equipment-free sample preparation separation technology designed to streamline the preparation of crude samples prior to nucleic acid testing. The practical utility of our platform is demonstrated by the successful detection of spiked SARS-CoV-2 RNA fragments in saliva, E. Coli in soil, and pathogenic E. Coli in clinically fecal samples of infected patients. Furthermore, we demonstrate that the paper-based LAMP CRISPR chips employed in our assays possess a shelf life of several weeks, establishing them as viable candidates for on-site diagnostics. Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.) |
| فهرسة مساهمة: | Keywords: COVID-19; CRISPR/Cas12a; Loop-mediated isothermal amplification (LAMP); Molecular diagnostics; Nucleic acid amplification tests; Paper-based diagnostics; SARS-CoV-2; Smartphone diagnostics |
| المشرفين على المادة: | 0 (CRISPR-Associated Proteins) |
| SCR Protocol: | LAMP assay |
| تواريخ الأحداث: | Date Created: 20240423 Date Completed: 20240508 Latest Revision: 20240508 |
| رمز التحديث: | 20240509 |
| DOI: | 10.1016/j.bios.2024.116292 |
| PMID: | 38653014 |
| قاعدة البيانات: | MEDLINE |
Full Text Finder | تدمد: | 1873-4235 |
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| DOI: | 10.1016/j.bios.2024.116292 |