Rapid and sensitive detection of miRNA via light scatter-aided emulsion-based isothermal amplification using a custom low-cost device
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Rapid_and_Sensitive_Detection .pdf
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2025-06-10
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Final Accepted Manuscript
Author
Hertenstein, TylerTang, Yisha
Day, Alexander S
Reynolds, Jocelyn
Viboolmate, Patrick V
Yoon, Jeong-Yeol

Affiliation
Department of Biomedical Engineering, The University of ArizonaDepartment of Chemical and Environmental Engineering, The University of Arizona
Issue Date
2023-06-10
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Elsevier LtdCitation
Hertenstein, T., Tang, Y., Day, A. S., Reynolds, J., Viboolmate, P. V., & Yoon, J. Y. (2023). Rapid and sensitive detection of miRNA via light scatter-aided emulsion-based isothermal amplification using a custom low-cost device. Biosensors and Bioelectronics, 237, 115444.Journal
Biosensors & bioelectronicsRights
© 2023 Elsevier B.V. All rights reserved.Collection Information
This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at repository@u.library.arizona.edu.Abstract
MicroRNAs are likely to be a next-generation clinical biomarker for many diseases. While gold-standard technologies, e.g., reverse transcription-quantitative polymerase chain reaction (RT-qPCR), exist for microRNA detection, there is a need for rapid and low-cost testing. Here, an emulsion loop-mediated isothermal amplification (eLAMP) assay was developed for miRNA that compartmentalizes a LAMP reaction and shortens the time-to-detection. The miRNA was a primer to facilitate the overall amplification rate of template DNA. Light scatter intensity decreased when the emulsion droplet got smaller during the ongoing amplification, which was utilized to moitor the amplification non-invasively. A custom low-cost device was designed and fabricated using a computer cooling fan, a Peltier heater, an LED, a photoresistor, and a temperature controller. It allowed more stable vortexing and accurate light scatter detection. Three miRNAs, miR-21, miR-16, and miR-192, were successfully detected using the custom device. Specifically, new template and primer sequences were developed for miR-16 and miR-192. Zeta potential measurements and microscopic observations confirmed emulsion size reduction and amplicon adsorption. The detection limit was 0.01 fM, corresponding to 2.4 copies per reaction, and the detection could be made in 5 min. Since the assays were rapid and both template and miRNA + template could eventually be amplified, we introduced the success rate (compared to the 95% confidence interval of the template result) as a new measure, which worked well with lower concentrations and inefficient amplifications. This assay brings us one step closer to allowing circulating miRNA biomarker detection to become commonplace in the clinical world.Note
24 month embargo; first published 10 June 2023EISSN
1873-4235PubMed ID
37329805Version
Final accepted manuscriptae974a485f413a2113503eed53cd6c53
10.1016/j.bios.2023.115444
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