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Evaluation of the Sulfo-Succinimidyl-4-(N-Maleimidomethyl) Cyclohexane-1-Carboxylate coupling chemistry for attachment of oligonucleotides to magnetic nanobeads
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Nanotechnology and Functional Materials. (Nanoteknologi och funktionella material)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Physics.
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2011 (English)In: Journal of Nanoscience and Nanotechnology, ISSN 1533-4880, Vol. 11, no 10, 8532-8537 p.Article in journal (Refereed) Published
Abstract [en]

The sulfo-SMCC (Succinimidyl-4-(N-maleimidomethyl) cyclohexane-1-carboxylate) coupling chemistry was evaluated for immobilization of oligonucleotides onto 130 nm sized magnetic nanobeads aimed for bio-detection in a magnetic readout assay. The chemistry was found to produce a high surface coverage of similar to 93 +/- 10 oligonucleotides per bead whereas stability tests showed that about 50% of the oligonucleotides detached from the bead surfaces after eight weeks of storage in a buffer solution. It was shown that bead aggregation prior to magnetic readout could be suppressed by incubating the samples at 70 degrees C for 30 min. The same temperature was also shown to be the most favorable for hybridization between the oligonucleotide functionalized beads and rolling circle amplified DNA molecules. This should simplify the heating procedure in a biosensor in which hybridization and magnetic readout is performed in the same compartment.

Place, publisher, year, edition, pages
American Scientific Publishers, 2011. Vol. 11, no 10, 8532-8537 p.
Keyword [en]
DNA Detection, Oligonucleotide Coupling Chemistry, Sulfo-SMCC, Magnetic Nanobeads, Biosensor
National Category
Nano Technology
Research subject
Engineering Science with specialization in Nanotechnology and Functional Materials
URN: urn:nbn:se:uu:diva-161323DOI: 10.1166/jnn.2011.5667ISI: 000298363900014OAI: oai:DiVA.org:uu-161323DiVA: diva2:455740
Available from: 2011-11-11 Created: 2011-11-11 Last updated: 2012-03-29Bibliographically approved
In thesis
1. Detection of Biomolecules Using Volume-Amplified Magnetic Nanobeads
Open this publication in new window or tab >>Detection of Biomolecules Using Volume-Amplified Magnetic Nanobeads
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes a new approach to biomolecular analysis, called the volume-amplified magnetic nanobead detection assay (VAM-DNA). It is a sensitive, specific magnetic bioassay that offers a potential platform for the development of low-cost, easy-to-use diagnostic devices. The VAM-NDA consists of three basic steps: biomolecular target recognition, enzymatic amplification of the probe-target complex using the rolling circle amplification (RCA) technique, and addition of target complementary probe-tagged magnetic nanobeads which exhibit Brownian relaxation behavior. Target detection is demonstrated by measuring the frequency-dependent complex magnetization of the magnetic beads. The binding of the RCA products (target DNA-sequence coils) to the bead surface causes a dramatic increase in the bead size, corresponding essentially to the size of the DNA coil (typically around one micrometer). This causes a decrease in the Brownian relaxation frequency, since it is inversely proportional to the hydrodynamic size of the beads. The concentration of the DNA coils is monitored by measuring the decrease in amplitude of the Brownian relaxation peaks of free beads.

The parameters oligonucleotide surface coverage, bead concentration, bead size and RCA times were investigated in this thesis to characterize features of the assay. It was found that all of these parameters affect the outcome and efficiency of the assay.

The possibility of implementing the assay on a portable, highly sensitive AC susceptometer platform was also investigated. The performance of the assay under these circumstances was compared with that using a superconducting quantum interference device (SQUID); the sensitivity of the assay was similar for both platforms. It is concluded that, the VAM-NDA opens up the possibility to perform biomolecular detection in point-of-care and outpatient settings on portable platforms similar to the one tested in this thesis.

Finally, the VAM-NDA was used to detect Escherichia coli bacteria and the spores of Bacillus globigii, the non-pathogenic simulant of Bacillus anthracis. A limit of detection of at least 50 bacteria or spores was achieved. This shows that the assay has great potential for sensitive detection of biomolecules in both environmental and biomedical applications.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2012. 65 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 905
Magnetic biosensor, magnetic nanobeads, Brownian relaxation, padlock probe, rolling circle amplification, DNA detection, protein detection
National Category
Nano Technology
Research subject
Engineering Science with specialization in Nanotechnology and Functional Materials
urn:nbn:se:uu:diva-169431 (URN)978-91-554-8288-6 (ISBN)
Public defence
2012-04-13, Å 2005, Ångströmlaboratoriet, Lägerhyddsvägen 1, Uppsala, 09:30 (English)
Available from: 2012-03-23 Created: 2012-02-29 Last updated: 2012-03-29Bibliographically approved

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Zardán Gómez de la Torre, TeresaHerthnek, DavidSvedlindh, PeterNilsson, MatsStrömme, Maria
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