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Antibodies Covalently Immobilized on Actin Filaments for Fast Myosin Driven Analyte Transport
Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.ORCID iD: 0000-0001-6878-3142
Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.ORCID iD: 0000-0003-2819-3046
Lund University.
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2012 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 10, e46298Article in journal (Refereed) Published
Abstract [en]

Biosensors would benefit from further miniaturization, increased detection rate and independence from external pumps and other bulky equipment. Whereas transportation systems built around molecular motors and cytoskeletal filaments hold significant promise in the latter regard, recent proof-of-principle devices based on the microtubule-kinesin motor system have not matched the speed of existing methods. An attractive solution to overcome this limitation would be the use of myosin driven propulsion of actin filaments which offers motility one order of magnitude faster than the kinesin-microtubule system. Here, we realized a necessary requirement for the use of the actomyosin system in biosensing devices, namely covalent attachment of antibodies to actin filaments using heterobifunctional cross-linkers. We also demonstrated consistent and rapid myosin II driven transport where velocity and the fraction of motile actin filaments was negligibly affected by the presence of antibody-antigen complexes at rather high density (>20 mu m(-1)). The results, however, also demonstrated that it was challenging to consistently achieve high density of functional antibodies along the actin filament, and optimization of the covalent coupling procedure to increase labeling density should be a major focus for future work. Despite the remaining challenges, the reported advances are important steps towards considerably faster nanoseparation than shown for previous molecular motor based devices, and enhanced miniaturization because of high bending flexibility of actin filaments.

Place, publisher, year, edition, pages
2012. Vol. 7, no 10, e46298
National Category
Biochemistry and Molecular Biology
Research subject
Natural Science, Biomedical Sciences
Identifiers
URN: urn:nbn:se:lnu:diva-22698DOI: 10.1371/journal.pone.0046298ISI: 000309454000032Scopus ID: 2-s2.0-84867081685OAI: oai:DiVA.org:lnu-22698DiVA: diva2:574434
Available from: 2012-12-05 Created: 2012-12-05 Last updated: 2017-12-07Bibliographically 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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Kumar, Sarojten Siethoff, LassePersson, MalinMånsson, Alf

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