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Integrated Sensing Chip for Ultrasensitive Label-Free Detection of the Products of Loop-Mediated Isothermal Amplification

  • Subin Mao
    Subin Mao
    Electrical and Computer Engineering Department, Iowa State University, Ames, Iowa 50011, United States
    More by Subin Mao
  • Jinping Zhao
    Jinping Zhao
    Texas A&M AgriLife Research Center at Dallas, Texas A&M University System, Dallas, Texas 75252, United States
    More by Jinping Zhao
  • Xiaoke Ding
    Xiaoke Ding
    Electrical and Computer Engineering Department, Iowa State University, Ames, Iowa 50011, United States
    More by Xiaoke Ding
  • Van Anh Vuong
    Van Anh Vuong
    Texas A&M AgriLife Research Center at Dallas, Texas A&M University System, Dallas, Texas 75252, United States
  • Junqi Song*
    Junqi Song
    Texas A&M AgriLife Research Center at Dallas, Texas A&M University System, Dallas, Texas 75252, United States
    Department of Plant Pathology & Microbiology, Texas A&M University, College Station, Texas 77843, United States
    *Email: [email protected]
    More by Junqi Song
  • , and 
  • Long Que*
    Long Que
    Electrical and Computer Engineering Department, Iowa State University, Ames, Iowa 50011, United States
    *Email: [email protected]
    More by Long Que
Cite this: ACS Sens. 2023, 8, 6, 2255–2262
Publication Date (Web):June 5, 2023
https://doi.org/10.1021/acssensors.3c00227
Copyright © 2023 American Chemical Society

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    Abstract

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    Loop-mediated isothermal amplification (LAMP) is a nucleic acid amplification technique that has been widely used for the detection of pathogens in many organisms. Current LAMP-based sensors usually require the LAMP products to be labeled in order for them to be detected. Here, we present a novel label-free LAMP chip, which consists of a nanopore thin-film sensor embedded inside a LAMP reaction chamber. A fraction of LAMP primers is immobilized on the sensor surface, allowing the LAMP products to be synthesized and bound to the sensor surface via immobilized primers. After the LAMP reaction components are removed from the reaction chamber, the amplified LAMP products bound to the sensor surface give rise to significantly increased transducing signals, which can be measured by a portable optical spectrometer through an optical fiber probe. As a demonstration, we used the LAMP chip to detect the causal agent of late blight, Phytophthora infestans, which is one of the most devastating plant pathogens and poses a major threat to sustainable crop production worldwide. We show that this chip can detect as low as 1 fg/μL of P. infestans DNA in 30 min, which corresponds to an attomolar level of 1.6 × 10–6 attomole/μL and is at least 10 times more sensitive than the currently available methods. This label-free sensing technology holds great promise to open up a new avenue for ultrasensitive, highly specific, rapid, and cost-effective point-of-care diagnostics of plant, animal, human, and foodborne pathogens.

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    The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acssensors.3c00227.

    • Fabrication process flow of the sensor chips; setups of the measurement; LAMP assay results; detection limit of real-time PCR; real-time PCR calibration curve for P. infestans detection; LAMP assays of P. infestans-infected potato plants; and a table of the LAMP and PCR primer sequences for the P. infestans ITS gene (PDF)

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