Droplet digital molecular beacon-LAMP assay via pico-injection for ultrasensitive detection of pathogens.

Ma, Zhiyuan; Ma, Mengshao; Cao, Xiaobao; Jiang, Yuyang; Gao, Dan

Ma, Zhiyuan; Ma, Mengshao; Cao, Xiaobao; Jiang, Yuyang; Gao, Dan.

Afiliación

Ma Z; The State Key Laboratory of Chemical Oncogenomics, Shenzhen International Graduate School and Open FIESTA, Shenzhen International Graduate School, Tsinghua University, Shenzhen, 518055, People's Republic of China.
Ma M; Key Laboratory of Metabolomics at Shenzhen, Graduate School at Shenzhen, Tsinghua University, Shenzhen, 518055, China.
Cao X; The State Key Laboratory of Chemical Oncogenomics, Shenzhen International Graduate School and Open FIESTA, Shenzhen International Graduate School, Tsinghua University, Shenzhen, 518055, People's Republic of China.
Jiang Y; Key Laboratory of Metabolomics at Shenzhen, Graduate School at Shenzhen, Tsinghua University, Shenzhen, 518055, China.
Gao D; Guangzhou Laboratory, Guangdong Province, 510320, China. xcao@ethz.ch.

Mikrochim Acta ; 191(7): 430, 2024 06 29.

Article en En | MEDLINE | ID: mdl-38949666

ABSTRACT

ABSTRACT

A pico-injection-aided digital droplet detection platform is presented that integrates loop-mediated isothermal amplification (LAMP) with molecular beacons (MBs) for the ultrasensitive and quantitative identification of pathogens, leveraging the sequence-specific detection capabilities of MBs. The microfluidic device contained three distinct functional units including droplet generation, pico-injection, and droplet counting. Utilizing a pico-injector, MBs are introduced into each droplet to specifically identify LAMP amplification products, thereby overcoming issues related to temperature incompatibility. Our methodology has been validated through the quantitative detection of Escherichia coli, achieving a detection limit as low as 9 copies/µL in a model plasmid containing the malB gene and 3 CFU/µL in a spiked milk sample. The total analysis time was less than 1.5 h. The sensitivity and robustness of this platform further demonstrated the potential for rapid pathogen detection and diagnosis, particularly when integrated with cutting-edge microfluidic technologies.

Asunto(s)

Escherichia coli; Límite de Detección; Leche; Técnicas de Amplificación de Ácido Nucleico; Técnicas de Amplificación de Ácido Nucleico/métodos; Escherichia coli/aislamiento & purificación; Escherichia coli/genética; Leche/microbiología; Animales; Técnicas de Diagnóstico Molecular/métodos; Técnicas Analíticas Microfluídicas/instrumentación; Técnicas Analíticas Microfluídicas/métodos; ADN Bacteriano/análisis; ADN Bacteriano/genética

Palabras clave

Droplet digital loop-mediated isothermal amplification; Droplet microfluidics; Molecular beacon; Nucleic acid quantification; Pathogen detection

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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Técnicas de Amplificación de Ácido Nucleico / Leche / Escherichia coli / Límite de Detección Límite: Animals Idioma: En Revista: Mikrochim Acta Año: 2024 Tipo del documento: Article Pais de publicación: Austria

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