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Rapid Newcastle Disease Virus Detection based on Loop-Mediated Isothermal Amplification and Optomagnetic Readout
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Physics.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Nanotechnology and Functional Materials.
Technical University of Denmark.
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2016 (English)In: ACS Sensors, ISSN 2379-3694, Vol. 1, no 10, p. 1228-1234Article in journal (Refereed) Published
Abstract [en]

Rapid and sensitive diagnostic methods based on isothermal amplification are ideal substitutes for PCR in out-of-lab settings. However, there are bottlenecks in terms of establishing low-cost and user-friendly readout methods for isothermal amplification schemes. Combining the high amplification efficiency of loop-mediated isothermal amplification (LAMP) with an optomagnetic' nanoparticle-based readout system, we demonstrate ultrasensitive and rapid detection of Newcastle disease virus RNA. Biotinylated amplicons of LAMP and reverse transcription LAMP (RT-LAMP) bind to streptavidin-coated magnetic nanoparticles (MNPs) resulting in a dramatical increase in the hydrodynamic size of the MNPs. This increase was measured by an optomagnetic readout system and provided quantitative information on the amount of LAMP target sequence. Our assay resulted in a limit of detection of 10 aM of target sequence with a total assay time of 30 min. The assay has also been tested on clinical samples (vaccine and tissue specimens) with a performance comparable to real-time RT-PCR By changing the LAMP primers, this strategy can serve as a general method for the detection of other DNA/RNA targets with high specificity and sensitivity.

Place, publisher, year, edition, pages
2016. Vol. 1, no 10, p. 1228-1234
Keywords [en]
out-of-lab diagnostics, Newcastle disease virus, loop-mediated isothermal amplification, optomagnetic bioassay, magnetic nanoparticles
National Category
Medical Laboratory and Measurements Technologies Physical Sciences
Research subject
Engineering Science with specialization in Solid State Physics
Identifiers
URN: urn:nbn:se:uu:diva-305174DOI: 10.1021/acssensors.6b00379ISI: 000386747600012OAI: oai:DiVA.org:uu-305174DiVA, id: diva2:1034522
Funder
Swedish Research Council Formas, 221-2012-444Swedish Research Council Formas, 221-2014-574Swedish Research Council Formas, 2011-1692Available from: 2016-10-12 Created: 2016-10-12 Last updated: 2018-03-13Bibliographically approved
In thesis
1. Magnetic Nanoparticle Based Biosensors for Pathogen Detection and Cancer Diagnostics
Open this publication in new window or tab >>Magnetic Nanoparticle Based Biosensors for Pathogen Detection and Cancer Diagnostics
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes several magnetic nanoparticle (MNP)-based biosensing strategies which take advantage of different magnetic sensors, molecular tools and nanotechnologies. Proposed biosensors can be classified into three groups, i.e., immunoassay-based, molecular amplification-based, and nanoparticle assembly-based. The principal motivation is to develop and optimize biosensors for out-of-lab and point-of-care testing.

Immunoassay-based biosensors described in this thesis employ antibodies as the bio-recognition element for the detection of bacteria cells/fragments or proteins. Two typical immunoassay formats, i.e., direct and competitive format, are studied and compared for bacteria detection. In addition, in the protein biomarker detection, MNP chains are formed in the presence of target analytes as well as in the external rotating magnetic field. The high shape/magnetic anisotropy of the chains provides better optomagnetic performance.

Two different molecular amplification methods, i.e., rolling circle amplification (RCA) and loop-mediated isothermal amplification (LAMP), are described under the topic of molecular amplification-based biosensors. In RCA-based biosensors, DNA probe modified MNPs bind to the amplicons after amplification. In LAMP-based biosensors, MNPs are either modified with primers that keep growing during the amplification, or are co-precipitated with the by-product (Mg2P2O7) of the amplification.

The design of the nanoparticle assembly-based biosensors described in this thesis is based on duplex-specific nuclease (DSN)-assisted target recycling and core-satellite magnetic superstructures. In the presence of target microRNA, DSN cuts the DNA scaffold of the core-satellite assembly, releasing MNP satellites that can be quantified by the sensor.

Different kinds of target analytes, i.e., pathogens or cancer biomarkers, are detected at the aiming for rapid, low-cost and user-friendly diagnostics.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. p. 55
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1647
Keywords
Magnetic biosensors, magnetic nanoparticles, homogeneous assays, volumetric sensing
National Category
Other Engineering and Technologies not elsewhere specified
Research subject
Engineering Science with specialization in Solid State Physics
Identifiers
urn:nbn:se:uu:diva-346014 (URN)978-91-513-0278-2 (ISBN)
Public defence
2018-05-04, Häggsalen, Ångströmlaboratoriet, Lägerhyddsv. 1, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2018-04-13 Created: 2018-03-13 Last updated: 2018-04-24

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Tian, BoZardán Gómez de la Torre, TeresaSvedlindh, PeterStrömberg, Mattias

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