Systematic screening and optimization of single-stranded DNA aptamer specific for N-acetylneuraminic acid: A comparative study

Feng

et al. 2022

Sensors

Owing to the significant roles of adenosine triphosphate (ATP) in diverse biological processes, ATP level is used to research and evaluate the physiological processes of organisms. Aptamer-based biosensors have been widely reported to achieve this purpose, which are superior in their flexible biosensing mechanism, with a high sensitivity and good biocompatibility; however, the aptamers currently used for ATP detection have a poor ability to discriminate ATP from adenosine diphosphate (ADP) and adenosine monophosphate (AMP). Herein, an ATP-specific aptamer was screened and applied to construct a fluorescent aptasensor for ATP by using graphene oxide (GO) and strand displacement amplification (SDA). The fluorescence intensity of the sensor is linearly related to the concentration of ATP within 0.1 μM to 25 μM under optimal experimental conditions, and the detection limit is 33.85 nM. The biosensor exhibits a satisfactory specificity for ATP. Moreover, the experimental results indicate that the biosensor can be applied to determine the ATP in human serum. In conclusion, the screened aptamer and the biosensor have promising applications in the determination of the real energy charge level and ATP content in a complex biological system.

Section: Resultsmentioning

confidence: 99%

Fluorescent Aptasensor for Highly Specific Detection of ATP Using a Newly Screened Aptamer

Feng

et al. 2022

Sensors

“…MBs are among the representative beads used in SELEX that enable simple and rapid partitioning of target-bound and unbound aptamers through the use of a magnet [51,58,62,81,82]. The quenching of MBs against fluorescence [83,84] can also be used for aptamers.…”

Section: Bead-mediated Aptamer Selectionmentioning

confidence: 99%

“…The pre-enrichment of aptamers by additional PCR amplification is usually required before aptamer immobilization on the MBs. A recent study has compared these two strategies by screening aptamers for a low-molecular target: N-acetylneuraminic acid (Neu5Ac) (Figure 1A) [58]. In this targetimmobilized strategy, the MBs were first functionalized with epoxy and amino groups, which can bind to the hydroxyl and carboxyl group of Neu5Ac, respectively.…”

Section: Bead-mediated Aptamer Selectionmentioning

confidence: 99%

“…Aptamer discovery strategy using bead-based platforms: (A) N-acetylneuraminic acid (Neu5Ac)-specific aptamer selection using a magnetic bead platform. Reproduced with permission from[58]; (B) Riboflavin-specific aptamer selection using an agarose bead platform. Reproduced with permission from[64]; (C) His-tagged N-cadherin protein-specific aptamer selection using a nickel Sepharose bead platform.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Recent Advances in Micro/Nanomaterial-Based Aptamer Selection Strategies

Kim

Lee

et al. 2021

Molecules

Aptamers are artificial nucleic acid ligands that have been employed in various fundamental studies and applications, such as biological analyses, disease diagnostics, targeted therapeutics, and environmental pollutant detection. This review focuses on the recent advances in aptamer discovery strategies that have been used to detect various chemicals and biomolecules. Recent examples of the strategies discussed here are based on the classification of these micro/nanomaterial-mediated systematic evolution of ligands by exponential enrichment (SELEX) platforms into three categories: bead-mediated, carbon-based nanomaterial-mediated, and other nanoparticle-mediated strategies. In addition to describing the advantages and limitations of the aforementioned strategies, this review discusses potential strategies to develop high-performance aptamers.

“…The separation process using the magnetic field is simple, and fast, and can save target molecules. In addition, magnetic bead separation can be used for automated screening [ 26 , 27 ]. There are many types of surface modification for magnetic bead carriers, such as streptavidin-coated magnetic beads, amino-modified magnetic beads, carboxyl-modified magnetic beads, epoxy-activated magnetic beads, and tosyl-modified magnetic beads [ 28 ].…”

Section: Introductionmentioning

confidence: 99%

Screening of Single-Stranded DNA Aptamer Specific for Florfenicol and Application in Detection of Food Safety

et al. 2022

In this work, the single-stranded DNA (ssDNA) aptamers specific to florfenicol (FF) and having a high binding affinity were prepared using the magnetic bead-based systematic evolution of ligands by the exponential enrichment technique (MB-SELEX). After 10 rounds of the MB-SELEX screening, aptamers that can simultaneously recognize FF and its metabolite florfenicol amine (FFA) were obtained. The aptamer with the lowest dissociation constant (Kd) was truncated and optimized based on a secondary structure analysis. The optimal aptamer selected was Apt-14t, with a length of 43 nt, and its dissociation constant was 4.66 ± 0.75 nM, which was about 7 times higher than that of the full-length sequence. The potential binding sites and interactions with FF were demonstrated by molecular docking simulations. In addition, a colorimetric strategy for nanogold aptamers was constructed. The linear detection range of this method was 0.00128–500 ng/mL and the actual detection limit was 0.00128 ng/mL. Using this strategy to detect florfenicol in actual milk and eggs samples, the spiked recoveries were 88.9–123.1% and 84.0–112.2%, respectively, and the relative standard deviation was less than 5.6%, showing high accuracy.