Label-Free Sensing of Adenosine Based on Force Variations Induced by Molecular Recognition

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.

Structural Disruption of an Adenosine-Binding DNA Aptamer on Graphene: Implications for Aptasensor Design

“…This phenomenon may, for example, result in the different behavior reported for the interaction of ssDNA and dsDNA with solid surfaces. With a highly flexible backbone and lack of defined secondary structure, ssDNA is thought to bind strongly to graphene and carbon nanotube (CNT) substrates. ,,,− ,, Accordingly, the consensus prediction from molecular dynamics (MD) simulations is that ssDNA adsorbs in a flat conformation with a majority of the nucleobases in direct contact with the substrate, although interbase interactions have been found to be important in some cases. − In contrast, dsDNA (and regions of ssDNA possessing well-defined secondary structure) tend to interact more weakly with CNTs and graphene, with the interfacial molecular structure relatively less affected by the adsorption process. ,,,,− Also, the molecular-level structure of the interaction of dsDNA/dsRNA with graphene interfaces appears less clear than in the case of ssDNA, with previously proposed surface-binding modes of dsDNA including an upright orientation via surface contact with the 3′ and 5′ terminal bases, or in a parallel orientation. ,,,, Overall, the interaction of oligonucleotides with graphitic substrates therefore depends on the balance between nucleotide base pairing, nucleotide π–π interactions, backbone flexibility, and nucleotide–substrate π–π interactions. In many instances, DNA aptamers may contain both regions of dsDNA and ssDNA, and therefore the molecular-scale details of the aptamer–graphene interaction may depend strongly on the nature of the aptamer.…”

“…However, we note here that these experiments were conducted for the graphene oxide substrate, which might yield very different outcomes than those generated for a graphene substrate. Furthermore, recent experimental efforts based on atomic force microscopy (AFM), namely, single molecule force spectroscopy (SMFS), investigated the interaction of this aptamer with the aqueous graphite interface . These SMFS experiments showed that by pulling the surface-adsorbed aptamer from aqueous graphite interface ,,, this produced a stable force plateau in the measured force–distance curves, which can be interpreted as representing the successive, base-by-base desorption of the strand from the surface.…”

“…Considering the aromatic character of both the nucleobases and graphitic substrates, it is not surprising that previous studies indicate a strongly favorable interaction between the two. − However, despite the strong favorable interaction with graphene at the nucleobase level, the greater macromolecular structure of an oligonucleotide may play a pivotal role in governing how the aptamer interacts with the aqueous graphitic interface. This phenomenon may, for example, result in the different behavior reported for the interaction of ssDNA and dsDNA with solid surfaces.…”

Biosensing, Characterization of Biosensors, and Improved Drug Delivery Approaches Using Atomic Force Microscopy: A Review

“…The detection method demonstrated in this study showed immense potential to develop more drug sensors. In another recent study, AFM force-based sensor was developed (Li J et al, 2015) for the detection of adenosine with higher sensitivity and probe the molecular recognition events between DNA aptamer and adenosine. This label-free detection procedure provides an easier and less timeconsuming way of identification of other similar chemical molecules.…”

Single-molecule force spectroscopy: A facile technique for studying the interactions between biomolecules and materials interfaces

“…Firstly, SMFS technique can be utilized to fabricate highly sensitive sensor platforms for detecting DNA, RNA, protein, enzyme, drug molecules, and metallic ions. Readers are suggested to study our previous review on the SMFS-based bioimaging and biosensing [25,92,93]. Here several recent studies on SMFS-based biosensing are introduced.…”