Noncoding small RNAs, such as microRNAs, are becoming the biomarkers of choice for multiple diseases in clinical diagnostics. A dysregulation of these microRNAs can be associated with many different diseases, such as cancer, dementia, and cardiovascular conditions. The key for effective treatment is an accurate initial diagnosis at an early stage, improving the patient's survival chances. In this work, the first clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a‐powered microfluidic, integrated electrochemical biosensor for the on‐site detection of microRNAs is introduced. Through this unique combination, the quantification of the potential tumor markers microRNA miR‐19b and miR‐20a is realized without any nucleic acid amplification. With a readout time of 9 min and an overall process time of less than 4 h, a limit of detection of 10 pm is achieved, using a measuring volume of less than 0.6 µL. Furthermore, the feasibility of the biosensor platform to detect miR‐19b in serum samples of children, suffering from brain cancer, is demonstrated. The validation of the obtained results with a standard quantitative real‐time polymerase chain reaction method shows the ability of the electrochemical CRISPR‐powered system to be a low‐cost, easily scalable, and target amplification‐free tool for nucleic acid based diagnostics.

https://onlinelibrary.wiley.com/doi/epdf/10.1002/adma.201905311

CRISPR/Cas13a‐Powered Electrochemical Microfluidic Biosensor for Nucleic Acid Amplification‐Free miRNA Diagnostics

Viruses are real menace to human safety that cause devastating viral disease. The high prevalence of these diseases is due to improper detecting tools. Therefore, there is a remarkable demand to identify viruses in a fast, selective and accurate way. Several biosensors have been designed and commercialized for detection of pathogenic viruses. However, they present many challenges. Nanotechnology overcomes these challenges and performs direct detection of molecular targets in real time. In this overview, studies concerning nanotechnology-based biosensors for pathogenic virus detection have been summarized, paying special attention to biosensors based on graphene oxide, silica, carbon nanotubes, gold, silver, zinc oxide and magnetic nanoparticles, which could pave the way to detect viral diseases and provide healthy life for infected patients.

Nanomaterial-based biosensors for detection of pathogenic virus

Development of biosensing platforms plays a key role in research settings for identification of biomarkers and in clinical applications for diagnostics. Biosensors based on nucleic acids have taken many forms, from simple duplex-based constructs to stimuli-responsive nucleic acid nanostructures. In this review, we look at various nucleic acid-based biosensors, the different read-out strategies employed, and their use in chemical and biological sensing. We also look at current developments in DNA nanotechnology-based biosensors and how rational design of such constructs leads to more efficient biosensing platforms.

Rationally Engineered Nucleic Acid Architectures for Biosensing Applications.

Aptamers evolved from live cells as effective molecular probes for cancer study

Polymeric DNA hydrogel: Design, synthesis and applications

Here we report a robust and sensitive DNA nanostructure-based electrochemical (E-nanoDNA) sensor that utilizes tetrahedral DNA nanostructures (TDNs) as an interfacial probe to detect biomolecules in a single-step procedure. In this study, we have firstly demonstrated that the use of TDNs can significantly suppress electrochemical background signals compared to traditional linear DNA probes upon introduction of base mismatches in the edges of TDNs. After further optimization of the two functional strands in the TDNs, quantitative, one-step detection of DNA can be achieved in the picomolar range in less than 10 min, and directly in complex media. Moreover, the baseline drift of this biosensor can be greatly decreased even after several hours in flowing whole blood in vitro, which suggests that the sensor holds potential to be employed in live animals. Furthermore, through replacing functional strands with aptamers or other DNA elements, this E-nanoDNA sensor can be easily used to probe various analytes, broadening the application range of the proposed sensor.

/pdf/design-of-dna-nanostructure-based-interfacial-probes-for-the-247wt6mluk.pdf

Design of DNA nanostructure-based interfacial probes for the electrochemical detection of nucleic acids directly in whole blood

Laborious and costly detection of miRNAs has brought challenges to its practical applications, especially for home health care, rigorous military medicine, and the third world. In this work, we present a pH-responsive miRNA amplification method, which allows the detection of miRNA just using a pH test paper. The operation is easy and no other costly instrument is involved, making the method very friendly. In our strategy, a highly efficient isothermal amplification of miRNA is achieved using an improved netlike rolling circle amplification (NRCA) technique. Large amounts of H+ can be produced as a byproduct during the amplification to induce significant changes of pH, which can be monitored directly using a pH test paper or pH-sensitive indicators. The degree of color changes depends on the amount of miRNA, making it possible for quantitative analysis. As an example, the method is successfully applied to quantify a miRNA (miR-21) in cancer cells. The results agree well with that from the prevalent qRT-PCR...

Detection of microRNA: A Point-of-Care Testing Method Based on a pH-Responsive and Highly Efficient Isothermal Amplification

Colorimetric detection of hepatitis B virus (HBV) DNA based on DNA-templated copper nanoclusters.

Nanozymes have received great attention owing to the advantages of easy preparation and low cost. Unlike natural enzymes that readily adapt to physiological environments, artificial nanozymes are apt to passivate in complex clinical samples (e.g., serum), which may damage the catalytic capability and consequently limit the application in biomedical analysis. To conquer this problem, in this study, we fabricated novel nanozyme@DNA hydrogel architecture by incorporating nanozymes into a pure DNA hydrogel. Gold nanoparticles (AuNPs) were adopted as a model nanozyme. Results indicate that AuNPs incorporated in the DNA hydrogel retain their catalytic capability in serum as they are protected by the hydrogel, whereas AuNPs alone totally lose the catalytic capability in serum. The detection of hydrogen peroxide and glucose in serum based on the catalysis of the AuNPs@DNA hydrogel was achieved. The detection limit of each reaches 1.7 and 38 μM, respectively, which is equal to the value obtained using natural enzymes. Besides the mechanisms, some other advantages, such as recyclability and availability, have also been explored. This nanozyme@DNA hydrogel architecture may have a great potential for the utilization of nanozymes as well as the application of nanozymes for biomedical analysis in complex physiological samples.

Fabrication of nanozyme@DNA hydrogel and its application in biomedical analysis

Hydrogels are of great interest in the field of biosensing for their good biocompatibility, plasticity, and capability of providing 3D scaffolds. Nevertheless, the application of hydrogels has not been linked with broad surface biosensing systems yet. To overcome the limitations, here for the first time, surface-immobilized pure DNA hydrogels were synthesized using a surficial primer-induced strategy and adopted for biosensing applications. The DNA hydrogel 3D scaffold is successfully constructed on a transparent ITO electrode, which facilitates both colourimetric and electrochemical measurements. Results show that the hydrogel is able to wrap enzymes solidly and exhibits favourable stability under different conditions. Owing to the free diffusion of the micromolecular targets throughout the hydrogel, while isolating the enzymes from the macromolecular interferences outside the hydrogel, the direct colourimetric and electrochemical detection of hydrogen peroxide and bilirubin in serum is achieved. The detection limit of hydrogen peroxide in serum is 22 nM by colourimetric analysis and 13 nM by electrochemical measurement. The detection limit of bilirubin is 32 nM, a favourable limit that could be used in jaundice diagnosis. In addition, the enzyme@hydrogel can be easily regenerated and the catalytic activity is retained for a few cycles, thus allowing the recycling of the hydrogel-based biosensing system. The successful integration of DNA hydrogels with surface biosensing systems will greatly expand the applications of hydrogels for diagnostic and environmental monitoring purposes.

/pdf/surface-immobilized-and-self-shaped-dna-hydrogels-and-their-3sy19h84jy.pdf

Xiaoxia Mao

Papers

Design of DNA nanostructure-based interfacial probes for the electrochemical detection of nucleic acids directly in whole blood

Detection of microRNA: A Point-of-Care Testing Method Based on a pH-Responsive and Highly Efficient Isothermal Amplification

Colorimetric detection of hepatitis B virus (HBV) DNA based on DNA-templated copper nanoclusters.

Fabrication of nanozyme@DNA hydrogel and its application in biomedical analysis

Surface-immobilized and self-shaped DNA hydrogels and their application in biosensing.