This article provides a comprehensive review of biosensing with DNAzymes, providing an overview of different sensing applications while highlighting major progress and seminal contributions to the field of portable biosensor devices and point-of-care diagnostics. Specifically, the field of functional nucleic acids is introduced, with a specific focus on DNAzymes. The incorporation of DNAzymes into bioassays is then described, followed by a detailed overview of recent advances in the development of in vivo sensing platforms and portable sensors incorporating DNAzymes for molecular recognition. Finally, a critical perspective on the field, and a summary of where DNAzyme-based devices may make the biggest impact are provided.

Biosensing with DNAzymes

With rapid development of nuclear industry, the overexploitation of uranium has brought a series of radioactive pollution problems to water, soil, atmosphere and ecosystem. Uranium and its compounds are highly carcinogenic and biologically toxic, posing a deadly threat to human health. Therefore, how to design a highly selective and sensitive sensor to detect uranyl ions in the environment and organisms has become an urgent task. This review summarizes relevant literature on the detection of uranyl ions by different sensors over the years. The detection methods can be roughly divided into three categories: optical sensors, electrochemical sensors and other sensors; further classification has also been carried out according to their detection mechanism. The detection limits and performances of different sensors are discussed to intuitively understand different capability for these methods in detecting uranyl ions. We also discuss the trend and future direction of uranium to provide a valuable reference for further developing better uranyl sensors.

Sensors for determination of uranium: A review

DNA switches are ideally suited for numerous nanotechnological applications, and increasing efforts are being directed toward their engineering. In this review, we discuss how to engineer these switches starting from the selection of a specific DNA-based recognition element, to its adaptation and optimisation into a switch, with applications ranging from sensing to drug delivery, smart materials, molecular transporters, logic gates and others. We provide many examples showcasing their high programmability and recent advances towards their real life applications. We conclude with a short perspective on this exciting emerging field.

Programmable DNA switches and their applications

Since their discovery almost three decades ago, DNAzymes have been used extensively in biosensing. Depending on the type of DNAzyme being used, these functional oligonucleotides can act as molecular recognition elements within biosensors, offering high specificity to their target analyte, or as reporters capable of transducing a detectable signal. Several parameters need to be considered when designing a DNAzyme-based biosensor. In particular, given that many of these biosensors immobilize DNAzymes onto a sensing surface, selecting an appropriate immobilization strategy is vital. Suboptimal immobilization can result in both DNAzyme detachment and poor accessibility toward the target, leading to low sensing accuracy and sensitivity. Various approaches have been employed for DNAzyme immobilization within biosensors, ranging from amine and thiol-based covalent attachment to non-covalent strategies involving biotin-streptavidin interactions, DNA hybridization, electrostatic interactions, and physical entrapment. While the properties of each strategy inform its applicability within a proposed sensor, the selection of an appropriate strategy is largely dependent on the desired application. This is especially true given the diverse use of DNAzyme-based biosensors for the detection of pathogens, metal ions, and clinical biomarkers. In an effort to make the development of such sensors easier to navigate, this paper provides a comprehensive review of existing immobilization strategies, with a focus on their respective advantages, drawbacks, and optimal conditions for use. Next, common applications of existing DNAzyme-based biosensors are discussed. Last, emerging and future trends in the development of DNAzyme-based biosensors are discussed, and gaps in existing research worthy of exploration are identified.

DNAzyme-Based Biosensors: Immobilization Strategies, Applications, and Future Prospective.

A novel nest hybridization chain reaction based electrochemical assay for sensitive detection of circulating tumor DNA.

A novel fluorescent and colorimetric strategy with enzyme-free dual amplification was developed for ultra-sensitively detection of uranyl in river waters samples. The enzyme-strand recycling and hairpin catalytic assembly (HCA) reaction were used as dual enzyme-free amplification strategies. Three hairpins labeled with fluorophore can absorb on the surface of gold nanoparticles (AuNPs) to prevent them from aggregation in salt solution and quench the fluorescent signal simultaneously. In the presence of UO22+, the HCA reaction between hairpins was induced to form rigid DNA triangles, resulting in the aggregation of AuNPs with a red to blue color change and a “turn-on” fluorescent signal. The limit of detection was as low as 0.1 pM. Importantly, this strategy provided colorimetric and fluorescent dual signals for lower false-positives and higher anti-interference ability. It was successfully used for detection of uranyl in real waters samples.

Ultra-sensitive fluorescent and colorimetric detection of UO22+ based on dual enzyme-free amplification strategies

A simple, one-step and amplified Hg2+ detection strategy based on DNAzyme motor was described here Fluorophore labeled DNA probes and long arm DNA are immobilized on the AuNPs by Au-S bond The fluorescent signal of DNA probe is significantly quenched by AuNPs In the presence of Hg2+, thymine-Hg2+-thymine (T-Hg2+-T) interaction can causes the duplex structure of split E-DNA and then form an entire E-DNA Then the E-DNA can bind with fluorophore labeled DNA to form Mg2+-dependent DNAzyme structure With the assistance of Mg2+, the fluorophore labeled DNA can be circularly cleaved by DNAzyme and then leave from the AuNPs surface The recycling cleavage and binding processes caused processive moving of E-DNA on AuNPs surface and significant fluorescent signal recovery A good linear relationship was obtained between fluorescence intensity and Hg2+ concentration in the range from 01 nM to 5 nM The limit of detection was calculated to be 30 pM This strategy was successfully used for detection of Hg2+ in Chinese herbs (Atractylodes macrocephala Koidz), showing a promising practical application future

Simple, one-step and amplified Hg2+ detection strategy based on DNAzyme motor

Herein, a fast and visible colorimetric method for bisphenol A (BPA) detection was developed using hemin-functionalized reduced graphene oxide (H–rGO) composites and aptamer. The aptamer can be stably adsorbed on the surface of H–rGO, preventing H–rGO from salt-induced aggregation. However, in the presence of BPA, the aptamer can bind with BPA; this results in the aggregation of H–rGO. Consequently, the supernatant contains a little H–rGO and shows light blue color after the chromogenic reaction. The color difference can be used for the quantitative detection of BPA. The designed aptasensor displayed a linear response for BPA in the range from 5 nM to 100 nM with a detection limit of 2 nM. This colorimetric aptasensing platform offered great advantages, including label-free conditions and visible result readout platform, for quick screening of BPA for on-site analysis and in-house diagnosis.

A visual detection of bisphenol A based on peroxidase-like activity of hemin–graphene composites and aptamer

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Haixia Zhong

Papers

Ultra-sensitive fluorescent and colorimetric detection of UO22+ based on dual enzyme-free amplification strategies

Simple, one-step and amplified Hg2+ detection strategy based on DNAzyme motor

A visual detection of bisphenol A based on peroxidase-like activity of hemin–graphene composites and aptamer

Fluorophore-based host-guest assembly complexes for imaging and therapy.