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Label-free sensing of adenosine based on force variations induced by molecular recognition.

Li J, Li Q, Ciacchi LC, Wei G - Biosensors (Basel) (2015)

Bottom Line: The sensitive force response to molecular recognition provided an adenosine detection limit in the range of 0.1 to 1 nM.The addition of guanosine, cytidine, and uridine had no significant interference with the sensing of adenosine, indicating a strong selectivity of this sensor architecture.In addition, operational parameters that may affect the sensor, such as loading rate and solution ionic strength, were investigated.

View Article: PubMed Central - PubMed

Affiliation: Hybrid Materials Interfaces Group, Faculty of Production Engineering, University of Bremen, Am Fallturm 1, 28359 Bremen, Germany. jinfeng@uni-bremen.de.

ABSTRACT
We demonstrate a simple force-based label-free strategy for the highly sensitive sensing of adenosine. An adenosine ssDNA aptamer was bound onto an atomic force microscopy (AFM) probe by covalent modification, and the molecular-interface adsorption force between the aptamer and a flat graphite surface was measured by single-molecule force spectroscopy (SMFS). In the presence of adenosine, the molecular recognition between adenosine and the aptamer resulted in the formation of a folded, hairpin-like DNA structure and hence caused a variation of the adsorption force at the graphite/water interface. The sensitive force response to molecular recognition provided an adenosine detection limit in the range of 0.1 to 1 nM. The addition of guanosine, cytidine, and uridine had no significant interference with the sensing of adenosine, indicating a strong selectivity of this sensor architecture. In addition, operational parameters that may affect the sensor, such as loading rate and solution ionic strength, were investigated.

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SMFS experiments and statistical analysis: (a,b) Typical FD curve by peeling aptamer from graphite surface (a) before and (b) after adding 1 μM adenosine; (c,d) Distributions of the desorption forces and tip-sample separations corresponding to the FD curves of (a); (e,f) same, for the FD curves of (b). After the adding of adenosine, the mean desorption force by peeling aptamer from graphite surface increased from 117.8 ± 29.5 to 164.3 ± 25.4 pN, and the mean tip-sample separation decreased from about 36.1 ± 2.0 to 25.2 ± 3.4 nm.
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biosensors-05-00085-f004: SMFS experiments and statistical analysis: (a,b) Typical FD curve by peeling aptamer from graphite surface (a) before and (b) after adding 1 μM adenosine; (c,d) Distributions of the desorption forces and tip-sample separations corresponding to the FD curves of (a); (e,f) same, for the FD curves of (b). After the adding of adenosine, the mean desorption force by peeling aptamer from graphite surface increased from 117.8 ± 29.5 to 164.3 ± 25.4 pN, and the mean tip-sample separation decreased from about 36.1 ± 2.0 to 25.2 ± 3.4 nm.

Mentions: SMFS measurements were then performed between the aptamer-tethered AFM probe and the graphite surface in binding buffer and adenosine solution (1 μM in binding buffer), respectively. Before the SMFS measurement in the presence of adenosine, the AFM probe was kept submerged in the solution (a few hundred μm above the graphite surface) for 1 h to allow enough time for the molecular recognition between aptamer and adenosine to take place. Typical FD curves obtained for both systems under a loading rate of 3.29 × 105 pN·s−1 are presented in Figure 4a,b, respectively. The approaching traces of both FD curves display a small jump-to-contact force at about 5 nm separation, probably due to the initial contact between the hanging ssDNA strand and the graphite surface [19]. On retraction, an initial large adhesive force is observed in both FD curves, which is associated to the nonspecific adhesive junction between the monolayer coating of the AFM probe and the hydrophobic surface [19,20,21]. After the initial pull-off, there is a large drop in force, but a roughly constant force persists with increasing tip-sample separation in both curves. This stable force plateau is interpreted as the progressive desorption of a single ssDNA strand from the graphite surface [19,22,25]. Importantly, for long enough PEG linkers and DNA aptamers, the non-specific interaction between probe and surface will not affect the value of the plateau force. In the absence of adenosine, in many cases the length of the plateau is up to the contour length of the fully extended 32-m ssDNA (approximately 18 nm). In some of the curves, the force decreases in characteristic discrete steps (not shown here), which we interpret as resulting from the successive detachment of a small number of ssDNA strands.


Label-free sensing of adenosine based on force variations induced by molecular recognition.

Li J, Li Q, Ciacchi LC, Wei G - Biosensors (Basel) (2015)

SMFS experiments and statistical analysis: (a,b) Typical FD curve by peeling aptamer from graphite surface (a) before and (b) after adding 1 μM adenosine; (c,d) Distributions of the desorption forces and tip-sample separations corresponding to the FD curves of (a); (e,f) same, for the FD curves of (b). After the adding of adenosine, the mean desorption force by peeling aptamer from graphite surface increased from 117.8 ± 29.5 to 164.3 ± 25.4 pN, and the mean tip-sample separation decreased from about 36.1 ± 2.0 to 25.2 ± 3.4 nm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384084&req=5

biosensors-05-00085-f004: SMFS experiments and statistical analysis: (a,b) Typical FD curve by peeling aptamer from graphite surface (a) before and (b) after adding 1 μM adenosine; (c,d) Distributions of the desorption forces and tip-sample separations corresponding to the FD curves of (a); (e,f) same, for the FD curves of (b). After the adding of adenosine, the mean desorption force by peeling aptamer from graphite surface increased from 117.8 ± 29.5 to 164.3 ± 25.4 pN, and the mean tip-sample separation decreased from about 36.1 ± 2.0 to 25.2 ± 3.4 nm.
Mentions: SMFS measurements were then performed between the aptamer-tethered AFM probe and the graphite surface in binding buffer and adenosine solution (1 μM in binding buffer), respectively. Before the SMFS measurement in the presence of adenosine, the AFM probe was kept submerged in the solution (a few hundred μm above the graphite surface) for 1 h to allow enough time for the molecular recognition between aptamer and adenosine to take place. Typical FD curves obtained for both systems under a loading rate of 3.29 × 105 pN·s−1 are presented in Figure 4a,b, respectively. The approaching traces of both FD curves display a small jump-to-contact force at about 5 nm separation, probably due to the initial contact between the hanging ssDNA strand and the graphite surface [19]. On retraction, an initial large adhesive force is observed in both FD curves, which is associated to the nonspecific adhesive junction between the monolayer coating of the AFM probe and the hydrophobic surface [19,20,21]. After the initial pull-off, there is a large drop in force, but a roughly constant force persists with increasing tip-sample separation in both curves. This stable force plateau is interpreted as the progressive desorption of a single ssDNA strand from the graphite surface [19,22,25]. Importantly, for long enough PEG linkers and DNA aptamers, the non-specific interaction between probe and surface will not affect the value of the plateau force. In the absence of adenosine, in many cases the length of the plateau is up to the contour length of the fully extended 32-m ssDNA (approximately 18 nm). In some of the curves, the force decreases in characteristic discrete steps (not shown here), which we interpret as resulting from the successive detachment of a small number of ssDNA strands.

Bottom Line: The sensitive force response to molecular recognition provided an adenosine detection limit in the range of 0.1 to 1 nM.The addition of guanosine, cytidine, and uridine had no significant interference with the sensing of adenosine, indicating a strong selectivity of this sensor architecture.In addition, operational parameters that may affect the sensor, such as loading rate and solution ionic strength, were investigated.

View Article: PubMed Central - PubMed

Affiliation: Hybrid Materials Interfaces Group, Faculty of Production Engineering, University of Bremen, Am Fallturm 1, 28359 Bremen, Germany. jinfeng@uni-bremen.de.

ABSTRACT
We demonstrate a simple force-based label-free strategy for the highly sensitive sensing of adenosine. An adenosine ssDNA aptamer was bound onto an atomic force microscopy (AFM) probe by covalent modification, and the molecular-interface adsorption force between the aptamer and a flat graphite surface was measured by single-molecule force spectroscopy (SMFS). In the presence of adenosine, the molecular recognition between adenosine and the aptamer resulted in the formation of a folded, hairpin-like DNA structure and hence caused a variation of the adsorption force at the graphite/water interface. The sensitive force response to molecular recognition provided an adenosine detection limit in the range of 0.1 to 1 nM. The addition of guanosine, cytidine, and uridine had no significant interference with the sensing of adenosine, indicating a strong selectivity of this sensor architecture. In addition, operational parameters that may affect the sensor, such as loading rate and solution ionic strength, were investigated.

Show MeSH
Related in: MedlinePlus