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Systematic investigation of biomolecular interactions using combined frequency and motional resistance measurements
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Electronics. (Thin Films Group)
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Electronics. (Thin Films Group)
Attana AB, Stockholm.
Attana AB, Stockholm.
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2011 (English)In: Sensors and actuators. B, Chemical, ISSN 0925-4005, E-ISSN 1873-3077, Vol. 153, no 1, 135-144 p.Article in journal (Refereed) Published
Abstract [en]

The resonance frequency of acoustic biosensors is today used as a label-free technique for detecting mass changes on sensor surfaces. In combination with an appropriate continuous flow system it has earlier been used for affinity and kinetic rate determination. Here, we assess the potential of a modified acoustic biosensor, monitoring also the real-time dissipation through the resistance of the sensor, to obtain additional kinetic information related to the structure and conformation of the molecules on the surface. Actual interaction studies, including an attempt to determine avidity, are presented along with thorough verification of the experimental setup utilizing true viscous load exposure together with protein and DNA immobilizations. True viscous loads show a linear relationship between resistance and frequency as expected. However, in the interaction studies between antibodies and proteins, as well as in the immobilization of DNA and proteins, higher surface concentrations of interacting molecules led to a decrease (i.e. deviation from the linear trend) in the differential resistance to frequency ratio. This is interpreted as increased surface rigidity at higher surface concentrations of immobilized molecules. Consequently, studies that aim at obtaining biological binding information, such as avidity, from real-time resistance and dissipation data should be conducted at low surface concentrations. In addition, the differential resistance to frequency relationship was found to be highly dependent on the rigidity of the preceding layer(s) of immobilized molecules. This dependence can be utilized to obtain a higher signal-to-noise ratio for resistance measurement by using low surface densities of immobilized interaction partners.

Place, publisher, year, edition, pages
2011. Vol. 153, no 1, 135-144 p.
Keyword [en]
biosensor, interaction analysis, QCM, dissipation, motional resistance, kinetics
National Category
Analytical Chemistry Other Industrial Biotechnology
Research subject
Analytical Chemistry; Engineering Science with specialization in Microsystems Technology; Engineering Science with specialization in Electronics
URN: urn:nbn:se:uu:diva-107253DOI: 10.1016/j.snb.2010.10.019ISI: 000289019300020OAI: oai:DiVA.org:uu-107253DiVA: diva2:228405
Available from: 2009-07-30 Created: 2009-07-30 Last updated: 2016-04-19Bibliographically approved
In thesis
1. Development of Electroacoustic Sensors for Biomolecular Interaction Analysis
Open this publication in new window or tab >>Development of Electroacoustic Sensors for Biomolecular Interaction Analysis
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Biomolecular interaction analysis to determine the kinetics and affinity between interacting partners is important for the fundamental understanding of biology, as well as for the development of new pharmaceutical substances. A quartz crystal microbalance instrument suitable for kinetics and affinity analyses of interaction events was developed. The functionality of the sensor system was demonstrated by development of an assay for relative affinity determination of lectin-carbohydrate interactions.

Sensor surfaces allowing for effective immobilization of one interacting partner is a key functionality of a biosensor. Here, three different surfaces and immobilization methods were studied. First, optimized preparation conditions for sensor surfaces based on carboxyl-terminated self assembled monolayers were developed and were demonstrated to provide highly functional biosensor surfaces with low non-specific binding. Second, a method allowing for immobilization of very acidic biomolecules based on the use of an electric field was developed and evaluated. The electric field made it possible to immobilize the highly acidic C-peptide on a carboxylated surface. Third, a method for antibody immobilization on a carboxyl surface was optimized and the influence of immobilization pH on the immobilization level and antigen binding capacity was thoroughly assessed. The method showed high reproducibility for a set of antibodies and allowed for antibody immobilization also at low pH.

Three broadly different strategies to increase the sensitivity of electroacoustic sensors were explored. A QCM sensor with small resonator electrodes and reduced flow cell dimensions was demonstrated to improve the mass transport rate to the sensor surface. The use of polymers on QCM sensor surfaces to enhance the sensor response was shown to increase the response of an antibody-antigen model system more than ten-fold. Moreover, the application of high frequency thin film bulk acoustic resonators for biosensing was evaluated with respect to sensing range from the surface. The linear detection range of the thin film resonator was determined to be more than sufficient for biosensor applications involving, for instance, antibody-antigen interactions. Finally, a setup for combined frequency and resistance measurements was developed and was found to provide time resolved data suitable for kinetics determination.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 68 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 658
biosensor, protein interactions, kinetics, affinity, QCM, quartz crystal microbalance, piezoelectric resonators, dissipation, motional resistance
National Category
Other Industrial Biotechnology Analytical Chemistry
Research subject
Engineering Science with specialization in Microsystems Technology
urn:nbn:se:uu:diva-107211 (URN)978-91-554-7572-7 (ISBN)
Public defence
2009-09-11, Å80101, Ångströmlaboratoriet, Lägerhyddsvägen 1, Uppsala, 09:30 (English)
Available from: 2009-08-20 Created: 2009-07-29 Last updated: 2014-11-25Bibliographically approved

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