Accelerated reaction by loop-mediated isothermal amplification using loop primers

Nagamine, Kentaro; Hase, Tetsu; Notomi, Tsugunori

doi:10.1006/mcpr.2002.0415

Cited by 1,699 publications

(1,305 citation statements)

References 15 publications

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“…1a), and after approximately 35 minutes in a normal water bath that maintains temperature at 62- The T. evansi type B LAMP test in study has been developed with the focus on reading colour change or using a LFD format to limit post DNA manipulation, therefore it is crucial that the assay amplifies the correct target. Theoretically, LAMP is highly specific since amplification of the target DNA is achieved through the use of six primers targeting eight sections of the desired DNA (Notomi et al, 2000;Nagamine et al, 2002). However, the strategy is still prone to false positives that can result from amplicon handling and formation of primer dimers (Njiru et al, 2008b).…”

Section: Discussionmentioning

confidence: 99%

Loop-mediated Isothermal Amplification (LAMP) test for detection of Trypanosoma evansi strain B

Njiru

Ouma

Enyaru

et al. 2010

Experimental Parasitology

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Please cite this article as: Njiru, Z.K., Ouma, J.O., Enyaru, J.C., Dargantes, A.P., Loop-mediated Isothermal Amplification (LAMP) test for detection of Trypanosoma evansi strain B, Experimental Parasitology (2010), doi: 10.1016/j.exppara. 2010.01.017 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. ACCEPTED MANUSCRIPT

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Section: Discussionmentioning

confidence: 99%

Loop-mediated Isothermal Amplification (LAMP) test for detection of Trypanosoma evansi strain B

Njiru

Ouma

Enyaru

et al. 2010

Experimental Parasitology

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“…These methods simplify molecular diagnostics by eliminating thermocycling and reducing equipment costs and power requirements, without sacrificing sensitivity or specificity. In particular, loop-mediated isothermal amplification (LAMP) 20,21 has a high tolerance for common components present in clinical samples that inhibit PCR and other reactions. 22−25 This robust tolerance toward inhibitors further streamlines assays, because it simplifies sample preparation and complexity/expense of equipment.…”

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confidence: 99%

“Paper Machine” for Molecular Diagnostics

2015

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Clinical tests based on primer-initiated amplification of specific nucleic acid sequences achieve high levels of sensitivity and specificity. Despite these desirable characteristics, these tests have not reached their full potential because their complexity and expense limit their usefulness to centralized laboratories. This paper describes a device that integrates sample preparation and loop-mediated isothermal amplification (LAMP) with end point detection using a hand-held UV source and camera phone. The prototype device integrates paper microfluidics (to enable fluid handling) and a multilayer structure, or a "paper machine", that allows a central patterned paper strip to slide in and out of fluidic path and thus allows introduction of sample, wash buffers, amplification master mix, and detection reagents with minimal pipetting, in a hand-held, disposable device intended for point-of-care use in resource-limited environments. This device creates a dynamic seal that prevents evaporation during incubation at 65°C for 1 h. This interval is sufficient to allow a LAMP reaction for the Escherichia coli malB gene to proceed with an analytical sensitivity of 1 doublestranded DNA target copy. Starting with human plasma spiked with whole, live E. coli cells, this paper demonstrates full integration of sample preparation with LAMP amplification and end point detection with a limit of detection of 5 cells. Further, it shows that the method used to prepare sample enables concentration of DNA from sample volumes commonly available from fingerstick blood draw.

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“…10 The reportedly high specificity of the LAMP reaction is based on the recognition of target sequences by 2 pairs of primers, whereas the sensitivity is driven by 1 additional pair of loop primers for acceleration of the reaction. 9 Special requirements for LAMP primer design, however, hamper the development of diagnostic assays when conserved regions in the target gene are scarce. The extremely high variability of the AIV hemagglutinin (HA) gene as a target for HA subtype-specific diagnostics poses a particularly difficult case not only for the design of primers for conventional PCR, but even more for appropriate LAMP primer sets.…”

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confidence: 99%

Evaluation of Two Commercial Loop-Mediated Isothermal Amplification Assays for Detection of Avian Influenza H5 and H7 Hemagglutinin Genes

Postel

Letzel

Frischmann³

et al. 2010

J VET Diagn Invest

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Abstract. Real-time reverse transcription loop-mediated isothermal amplification (real-time RT-LAMP) holds substantial potential as a highly sensitive, specific, and easy-to-perform molecular technique for pathogen detection in clinical samples. In the current study, the analytical and diagnostic performance of 2 commercial realtime RT-LAMP kits, Avian Flu H5 and Avian Flu H7, in detecting Avian influenza virus (AIV) infections were evaluated and compared with validated real-time reverse transcription polymerase chain reaction (RT-PCR) assays using RNA from reference virus isolates of subtypes H5 (n 5 24) and H7 (n 5 25) and of phylogenetically related subtypes (n 5 20). When real-time RT-LAMP was carried out according to the recommendations of the manufacturer, 3 out of 24 H5 isolates and 8 out of 25 H7 reference strains were not detected. Prolonging the amplification phase resulted in detection of all H5 isolates but also in false positive detection of 2 non-H5 isolates. Real-time RT-LAMP specific to H7 failed to detect 2 H7 isolates after prolonged amplification. According to the examination of RNA log dilutions, the sensitivity of the real-time RT-LAMP assays, for a number of historic but also recent strains, was considerably lower compared with subtype-specific real-time RT-PCR assays. Application of the real-time RT-LAMP assays for analysis of diagnostic samples from wild birds confirmed their lower sensitivity. Commercial real-time RT-LAMP as tested in this study with a broad range of AIV H5 and H7 strains of phylogenetically diverse yet recent origin, holds some promise for routine veterinary diagnostic purposes, although real-time RT-LAMP was markedly more vulnerable to a reduction of detection limits because of strainspecific sequence variation than subtype-specific real-time RT-PCR.

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Accelerated reaction by loop-mediated isothermal amplification using loop primers

Cited by 1,699 publications

References 15 publications

Loop-mediated Isothermal Amplification (LAMP) test for detection of Trypanosoma evansi strain B

Loop-mediated Isothermal Amplification (LAMP) test for detection of Trypanosoma evansi strain B

“Paper Machine” for Molecular Diagnostics

Evaluation of Two Commercial Loop-Mediated Isothermal Amplification Assays for Detection of Avian Influenza H5 and H7 Hemagglutinin Genes

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