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A Point-of-Care Nucleic Acid Quantification Method by Counting Light Spots Formed by LAMP Amplicons on a Paper Membrane

文献类型： 外文期刊

作者： Chen, Yanju 1 ; Zhu, Yuanyuan 1 ; Peng, Cheng 2 ; Wang, Xiaofu 2 ; Wu, Jian 1 ; Chen, Huan 3 ; Xu, Junfeng 2 ;

作者机构： 1.Zhejiang Univ, Coll Biosyst Engn & Food Sci, ZJU Hangzhou Global Sci & Technol Innovat Ctr, Hangzhou 310058, Peoples R China

2.Zhejiang Acad Agr Sci, Key Lab Traceabil Agr Genet Modified Organisms, Minist Agr & Rural Affairs, Hangzhou 310021, Peoples R China

3.Hangzhou Digital Micro Biotech Co Ltd, Hangzhou 311215, Peoples R China

关键词： nucleic acid quantification; digital detection; paper membrane; probe-based LAMP

期刊名称：BIOSENSORS-BASEL （ 影响因子：5.4； 五年影响因子：5.7 ）

ISSN：

年卷期： 2024 年 14 卷 3 期

页码：

收录情况： SCI

摘要： Nucleic acid quantification, allowing us to accurately know the copy number of target nucleic acids, is significant for diagnosis, food safety, agricultural production, and environmental protection. However, current digital quantification methods require expensive instruments or complicated microfluidic chips, making it difficult to popularize in the point-of-care detection. Paper is an inexpensive and readily available material. In this study, we propose a simple and cost-effective paper membrane-based digital loop-mediated isothermal amplification (LAMP) method for nucleic acid quantification. In the presence of DNA fluorescence dyes, the high background signals will cover up the amplicons-formed bright spots. To reduce the background fluorescence signals, a quencher-fluorophore duplex was introduced in LAMP primers to replace non-specific fluorescence dyes. After that, the amplicons-formed spots on the paper membrane can be observed; thus, the target DNA can be quantified by counting the spots. Take Vibrio parahaemolyticus DNA detection as an instance, a good linear relationship is obtained between the light spots and the copy numbers of DNA. The paper membrane-based digital LAMP detection can detect 100 copies target DNA per reaction within 30 min. Overall, the proposed nucleic acid quantification method has the advantages of a simple workflow, short sample-in and answer-out time, low cost, and high signal-to-noise, which is promising for application in resourced limited areas.

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