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Magnetic capture from blood rescues molecular motor function in diagnostic nanodevices
Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.ORCID iD: 0000-0001-6878-3142
Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.ORCID iD: 0000-0003-2819-3046
Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
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2013 (English)In: Journal of Nanobiotechnology, ISSN 1477-3155, E-ISSN 1477-3155, Vol. 11, 14Article in journal (Refereed) Published
Abstract [en]

Background: Introduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs. Molecular motor driven nanodevices have unique potentials in this regard due to unprecedented level of miniaturization and independence of external pumps. However motor function has been found to be inhibited by body fluids. Results: We report here that a unique procedure, combining separation steps that rely on antibody-antigen interactions, magnetic forces applied to magnetic nanoparticles (MPs) and the specificity of the actomyosin bond, can circumvent the deleterious effects of body fluids (e.g. blood serum). The procedure encompasses the following steps: (i) capture of analyte molecules from serum by MP-antibody conjugates, (ii) pelleting of MP-antibody-analyte complexes, using a magnetic field, followed by exchange of serum for optimized biological buffer, (iii) mixing of MP-antibody-analyte complexes with actin filaments conjugated with same polyclonal antibodies as the magnetic nanoparticles. This causes complex formation: MP-antibody-analyte-antibody-actin, and magnetic separation is used to enrich the complexes. Finally (iv) the complexes are introduced into a nanodevice for specific binding via actin filaments to surface adsorbed molecular motors (heavy meromyosin). The number of actin filaments bound to the motors in the latter step was significantly increased above the control value if protein analyte (50-60 nM) was present in serum (in step i) suggesting appreciable formation and enrichment of the MP-antibody-analyte-antibody-actin complexes. Furthermore, addition of ATP demonstrated maintained heavy meromyosin driven propulsion of actin filaments showing that the serum induced inhibition was alleviated. Detailed analysis of the procedure i-iv, using fluorescence microscopy and spectroscopy identified main targets for future optimization. Conclusion: The results demonstrate a promising approach for capturing analytes from serum for subsequent motor driven separation/detection. Indeed, the observed increase in actin filament number, in itself, signals the presence of analyte at clinically relevant nM concentration without the need for further motor driven concentration. Our analysis suggests that exchange of polyclonal for monoclonal antibodies would be a critical improvement, opening for a first clinically useful molecular motor driven lab-on-a-chip device.

Place, publisher, year, edition, pages
2013. Vol. 11, 14
Keyword [en]
Magnetic nanoparticle, Biomolecular motor, Myosin, Nanoseparation, Lab-on-a-chip, Bioconjugation
National Category
Medical Biotechnology
Research subject
Natural Science, Biomedical Sciences
Identifiers
URN: urn:nbn:se:lnu:diva-27565DOI: 10.1186/1477-3155-11-14ISI: 000319316500001OAI: oai:DiVA.org:lnu-27565DiVA: diva2:637252
Available from: 2013-07-17 Created: 2013-07-17 Last updated: 2017-02-16Bibliographically approved
In thesis
1. Towards Myosin Powered Lab-on-a-Chip Devices
Open this publication in new window or tab >>Towards Myosin Powered Lab-on-a-Chip Devices
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Mot utvecklandet av myosindrivna laboratorier på chip
Abstract [en]

Myosins are protein motors that use chemical energy in the form of adenosinetriphosphate to produce force and motion. These molecular motors might be usedto power transportation in Lab-on-a-chip devices where a series of laboratory tasks(e.g. separation, concentration and detection) are performed in one sequence on asmall chip. Because of the small size, lab-on-a-chip devices are predicted to befaster and more sensitive than conventional systems. Further potential advantagesinclude cost efficiency and the possibility to perform many analyzes in parallel.Substituting microfluidics with myosin based transport would allow furtherminiaturization and make lab-on-a-chip devices more readily portable by reducingthe need for external power supplies. However, there are also limitations thathamper the development of such devices. Here we investigate several aspects of amyosin powered lab-on-a-chip device and present ways to overcome criticallimitations. First we demonstrate covalent attachment of antibodies to actinfilament shuttles with retained ability of the filaments to be propelled by myosinfragments, previously believed to be difficult. Secondly we develop a separationmethod to overcome the deleterious effects of body fluids on the actomyosinsystem. Thirdly, we explore the possibility to concentrate actin shuttles on ananostructured surface and achieve >20 times concentration in <1 min. Monte-Carlo simulations of the concentration process suggest further room forimprovement. Fourth, we develop novel techniques for fast and automaticdetection of fluorescence at certain check points which improves S/N ratio >20times. Finally, we take the first steps towards the development of threedimensional,nanowire-based transport systems, important both for lab-on-a-chipapplications and fundamental studies. Our results demonstrate the potential of amyosin based lab-on-a-chip device and lay the foundation for furtherdevelopments. Thus, we anticipate that this work will influence future studiestowards a complete diagnostic lab-on-a-chip work-up based on molecular motors.In addition, the work might also have implications for the development of futurebiocomputation and drug screening devices as well as novel biophysical studies ofthe actomyosin system.

Place, publisher, year, edition, pages
Växjö: Linnaeus University Press, 2013. 200 p.
Series
Linnaeus University Dissertations, 144/2013
Keyword
myosin, actin, molecular motors, lab-on-a-chip, nanobiotechnology, bionanotechnology, diagnostics, nanowires, nanowire arrays
National Category
Biochemistry and Molecular Biology
Research subject
Natural Science, Biomedical Sciences
Identifiers
urn:nbn:se:lnu:diva-28384 (URN)978-91-87427-45-9 (ISBN)
Public defence
2013-09-13, N2007, Västergård, Smålandsgatan 26E, Kalmar, 09:00 (English)
Opponent
Supervisors
Funder
EU, FP7, Seventh Framework Programme, 228971
Available from: 2013-09-10 Created: 2013-08-22 Last updated: 2016-05-03Bibliographically approved

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