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Comprehensive characterization of molecular interactions based on nanomechanics.

Ghatkesar MK, Lang HP, Gerber C, Hegner M, Braun T - PLoS ONE (2008)

Bottom Line: Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro.This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology.The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: National Center of Competence for Research in Nanoscience, Institute of Physics, University of Basel, Basel, Switzerland.

ABSTRACT
Molecular interaction is a key concept in our understanding of the biological mechanisms of life. Two physical properties change when one molecular partner binds to another. Firstly, the masses combine and secondly, the structure of at least one binding partner is altered, mechanically transducing the binding into subsequent biological reactions. Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro. This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology. To prove the concept we measured lipid vesicle (approximately 748*10(6) Da) adsorption on the sensor interface followed by subsequent binding of the bee venom peptide melittin (2840 Da) to the vesicles. The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

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Schematic of the homemade measurement set-up for combined mode measurements.For more information see also Braun et al. 2007 [15]. An array of silicon cantilevers was mounted onto a piezo element. A sinusoidal excitation signal generated from a network analyzer swept the requested frequency range vibrating the cantilevers (1). The laser beam deflection detection technique was used to monitor the response of individual cantilevers (2). A frequency analyzer (3) compared input- and output signals and continuously recorded amplitude and phase spectra (4) as well as static deflection (bending) of the cantilevers (5). NOSETools software was used to analyze the spectra and extract the mass adsorption on the cantilever (postprocessing, 6). [5], [15], [21], [35] A scanning electron microscopy image of a cantilever array is shown in the supplemental Figure S1.
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pone-0003610-g001: Schematic of the homemade measurement set-up for combined mode measurements.For more information see also Braun et al. 2007 [15]. An array of silicon cantilevers was mounted onto a piezo element. A sinusoidal excitation signal generated from a network analyzer swept the requested frequency range vibrating the cantilevers (1). The laser beam deflection detection technique was used to monitor the response of individual cantilevers (2). A frequency analyzer (3) compared input- and output signals and continuously recorded amplitude and phase spectra (4) as well as static deflection (bending) of the cantilevers (5). NOSETools software was used to analyze the spectra and extract the mass adsorption on the cantilever (postprocessing, 6). [5], [15], [21], [35] A scanning electron microscopy image of a cantilever array is shown in the supplemental Figure S1.

Mentions: This work focuses on the development and testing of an instrument that measures the integral nanomechanics of molecular interactions [1]–[3]. This device relies upon the unique ability of thin cantilevers [4] to detect both the mass of the adsorbed molecules and nanomechanical changes on the cantilever interface, e. g. structural rearrangements. The mass is measured via the resonance frequency of the cantilever (dynamic mode) [5], [6]. Structural changes are detected by static bending of cantilevers (static mode) as demonstrated recently [7]–[9]. Here nanomechanical interaction changes generate a surface-stress difference between the asymmetrically functionalized cantilever interfaces forcing the beam to bend [10]–[14]. Technical details of the method are presented in Figure 1. In summary, the micro-fabricated cantilever arrays (supplemental Figure S1) are actuated for a given frequency range and the response is recorded as amplitude and phase spectra for the individual cantilever sensors. These spectra are post-processed and various physical properties of the system can be extracted, such as the adsorbed mass (dynamic mode) and the static cantilever bending (static mode) [15].


Comprehensive characterization of molecular interactions based on nanomechanics.

Ghatkesar MK, Lang HP, Gerber C, Hegner M, Braun T - PLoS ONE (2008)

Schematic of the homemade measurement set-up for combined mode measurements.For more information see also Braun et al. 2007 [15]. An array of silicon cantilevers was mounted onto a piezo element. A sinusoidal excitation signal generated from a network analyzer swept the requested frequency range vibrating the cantilevers (1). The laser beam deflection detection technique was used to monitor the response of individual cantilevers (2). A frequency analyzer (3) compared input- and output signals and continuously recorded amplitude and phase spectra (4) as well as static deflection (bending) of the cantilevers (5). NOSETools software was used to analyze the spectra and extract the mass adsorption on the cantilever (postprocessing, 6). [5], [15], [21], [35] A scanning electron microscopy image of a cantilever array is shown in the supplemental Figure S1.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2572191&req=5

pone-0003610-g001: Schematic of the homemade measurement set-up for combined mode measurements.For more information see also Braun et al. 2007 [15]. An array of silicon cantilevers was mounted onto a piezo element. A sinusoidal excitation signal generated from a network analyzer swept the requested frequency range vibrating the cantilevers (1). The laser beam deflection detection technique was used to monitor the response of individual cantilevers (2). A frequency analyzer (3) compared input- and output signals and continuously recorded amplitude and phase spectra (4) as well as static deflection (bending) of the cantilevers (5). NOSETools software was used to analyze the spectra and extract the mass adsorption on the cantilever (postprocessing, 6). [5], [15], [21], [35] A scanning electron microscopy image of a cantilever array is shown in the supplemental Figure S1.
Mentions: This work focuses on the development and testing of an instrument that measures the integral nanomechanics of molecular interactions [1]–[3]. This device relies upon the unique ability of thin cantilevers [4] to detect both the mass of the adsorbed molecules and nanomechanical changes on the cantilever interface, e. g. structural rearrangements. The mass is measured via the resonance frequency of the cantilever (dynamic mode) [5], [6]. Structural changes are detected by static bending of cantilevers (static mode) as demonstrated recently [7]–[9]. Here nanomechanical interaction changes generate a surface-stress difference between the asymmetrically functionalized cantilever interfaces forcing the beam to bend [10]–[14]. Technical details of the method are presented in Figure 1. In summary, the micro-fabricated cantilever arrays (supplemental Figure S1) are actuated for a given frequency range and the response is recorded as amplitude and phase spectra for the individual cantilever sensors. These spectra are post-processed and various physical properties of the system can be extracted, such as the adsorbed mass (dynamic mode) and the static cantilever bending (static mode) [15].

Bottom Line: Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro.This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology.The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: National Center of Competence for Research in Nanoscience, Institute of Physics, University of Basel, Basel, Switzerland.

ABSTRACT
Molecular interaction is a key concept in our understanding of the biological mechanisms of life. Two physical properties change when one molecular partner binds to another. Firstly, the masses combine and secondly, the structure of at least one binding partner is altered, mechanically transducing the binding into subsequent biological reactions. Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro. This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology. To prove the concept we measured lipid vesicle (approximately 748*10(6) Da) adsorption on the sensor interface followed by subsequent binding of the bee venom peptide melittin (2840 Da) to the vesicles. The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

Show MeSH
Related in: MedlinePlus