LAMP-Based Point-of-Care Biosensors for Rapid Pathogen Detection

Das, Dhrubajyoti; Lin, Cheng‐Wen; Chuang, Huey-Ru

doi:10.3390/bios12121068

Cited by 18 publications

(12 citation statements)

References 189 publications

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“…Additionally, DNA melting curves can distinguish real amplicons from amplified primers [ 91 ], and they are easily collected after voltammetric LAMP with the same electroactive probes and on the same device [ 59 , 68 ]. LAMP could be combined with other assays for improved specificity [ 17 ], and it has also been shown that using multiple primer sets in a single LAMP reaction can improve both specificity and time-to-detection [ 92 ]. Methods that rely on sequence-specific DNA probes, such as the voltammetric detection reported by Liu et al (2020) ( Figure 2 B) [ 40 ], should also provide an additional layer of specificity.…”

Section: Discussionmentioning

confidence: 99%

“…Presently, the most common detection strategy is optical, and, because of the extremely high efficiency amplification, detection can often be by naked-eye colorimetric (e.g., color change of a pH indicator) or turbidimetric (precipitation of magnesium pyrophosphate) observations. Due to the lack of a requirement for detection equipment, this is very attractive for point-of-use applications [ 17 ]. However, these relatively simple methods typically lack sensitivity and the capacity for accurate quantification.…”

Section: Introductionmentioning

confidence: 99%

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Strategies for the Voltammetric Detection of Loop-Mediated Isothermal Amplification

Marangoni

Emadi

2023

Micromachines

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Loop-mediated isothermal amplification (LAMP) is rapidly developing into an important tool for the point-of-use detection of pathogens for both clinical and environmental samples, largely due to its sensitivity, rapidity, and adaptability to portable devices. Many methods are used to monitor LAMP, but not all are amenable to point-of-use applications. Common methods such as fluorescence often require bulky equipment, whereas colorimetric and turbidimetric methods can lack sensitivity. Electrochemical biosensors are becoming increasingly important for these applications due to their potential for low cost, high sensitivity, and capacity for miniaturization into integrated devices. This review provides an overview of the use of voltammetric sensors for monitoring LAMP, with a specific focus on how electroactive species are used to interface between the biochemical products of the LAMP reaction and the voltammetric sensor. Various strategies for the voltammetric detection of DNA amplicons as well as pyrophosphate and protons released during LAMP are presented, ranging from direct DNA binding by electroactive species to the creative use of pyrophosphate-detecting aptamers and pH-sensitive oligonucleotide structures. Hurdles for adapting these devices to point-of-use applications are also discussed.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Strategies for the Voltammetric Detection of Loop-Mediated Isothermal Amplification

Marangoni

Emadi

2023

Micromachines

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“…7 A key advantage of LAMP is its simplicity, as the singletemperature incubation requirement enables on-site testing in the field without the reliance on costly machinery. 7,8 Consequently, RT-LAMP assays have been developed to rapidly identify CSFV. [9][10][11][12] However, these previously developed assays necessitated a laborious and timeconsuming RNA extraction step.…”

mentioning

confidence: 99%

“…Additionally, the inclusion of a color‐based indicator in the reaction mixture allows for visual interpretation of LAMP results without the need for specialized equipment 7 . A key advantage of LAMP is its simplicity, as the single‐temperature incubation requirement enables on‐site testing in the field without the reliance on costly machinery 7,8 . Consequently, RT‐LAMP assays have been developed to rapidly identify CSFV 9–12 .…”

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confidence: 99%

Enhancing classical swine fever virus identification: the advantages of Field‐LAMP testing

Tran,

et al. 2023

Aust Veterinary J

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Classical swine fever virus (CSFV) identification has witnessed significant advancements with the development of rapid reverse‐transcription loop‐mediated isothermal amplification (RT‐LAMP) assays. However, conventional RT‐LAMP assays for CSFV diagnosis are hindered by a laborious RNA extraction step. Moreover, the need for thermal incubators and expensive micropipettes has limited their application in field settings. Addressing these challenges, our study presents a groundbreaking solution—an electro‐free and point‐of‐care (POC) tool known as the field‐LAMP assay—for the rapid clinical detection of CSFV. By eliminating the RNA extraction requirement, advancing the colorimetric read‐out and lyophilized reaction reagents, our field‐LAMP assay streamlines the diagnostic process, saving valuable time and effort. This novel approach also overcomes the dependency on electric‐dependent thermal incubators and expensive micropipettes, making it practical and accessible for use in the field. The successful development of the field‐LAMP assay marks a significant milestone in CSFV detection. This electro‐free and POC tool offers several advantages, including its ability to deliver rapid results without compromising accuracy, facilitating prompt response and containment measures.

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“…Loop-mediated isothermal amplification (LAMP) has become a powerful alternative to polymerase chain reaction (PCR) for pathogen detection in clinical specimens , and food matrices. , Owing to the ability of Bst DNA polymerase to display a high strand displacement activity at a constant temperature (60–65 °C), isothermal LAMP has the potential to revolutionize molecular biology using simpler instrumentation, as well as faster and more efficient assays than PCR, even for crude samples . Consequently, LAMP has been adopted as a low-technology molecular analysis tool for resource-limiting areas and point-of-care (POC) applications . Achieving efficient detection at the POC, though, depends largely on the method selected for amplicon detection.…”

mentioning

confidence: 99%

Naked-Eye Detection of LAMP-Produced Nucleic Acids in Saliva Using Chitosan-Capped AuNPs in a Single-Tube Assay

Grammatikos,

Svoliantopoulos,

Gizeli

2023

Anal. Chem.

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Loop-mediated isothermal amplification (LAMP) is a low-technology molecular assay that is highly adaptable to point-of-care (POC) applications. However, achieving sensitive naked-eye detection of the amplified target in a crude sample is challenging. Herein, we report a simple yet highly efficient and sensitive methodology for the colorimetric visualization of a single target copy in saliva using chitosan-capped gold nanoparticles (Chit-AuNPs) synthesized via a green chemistry approach. The presence or absence of free Chit in the Chit-AuNPs solution was shown to affect LAMP colorimetric detection oppositely: the observed stabilization in the negative samples and aggregation in the positive samples in the presence of free Chit were reversed in the case of neat Chit-AuNPs. The mechanism of the two assays was investigated and attributed to electrostatic and depletion effects exerted between the Chit-AuNPs, free Chit, and the solution components. The developed contamination-free, one-tube assay successfully amplified and detected down to 1–5 cfu of Salmonella and 10 copies of SARS-CoV-2 per reaction (25 μL) used, respectively, as model DNA and RNA targets in the presence of 20% saliva, making the method suitable for POC applications. Compared to the commonly used pH-sensitive dyes, Chit-AuNPs are shown to have an enhanced sensitivity toward naked-eye colorimetric observation owing to the direct detection of DNA amplicons. Thus, this is a simple, highly sensitive, fast, and versatile naked-eye detection methodology that could be coupled to any LAMP or RT-LAMP assay, avoiding the need of using complicated sample pretreatments and/or AuNPs long and laborious functionalization processes.

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LAMP-Based Point-of-Care Biosensors for Rapid Pathogen Detection

Cited by 18 publications

References 189 publications

Strategies for the Voltammetric Detection of Loop-Mediated Isothermal Amplification

Strategies for the Voltammetric Detection of Loop-Mediated Isothermal Amplification

Enhancing classical swine fever virus identification: the advantages of Field‐LAMP testing

Naked-Eye Detection of LAMP-Produced Nucleic Acids in Saliva Using Chitosan-Capped AuNPs in a Single-Tube Assay

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