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In this work, a paper‐based electrochemical aptamer sensor was developed for the detection of aflatoxin B1 (AFB1) using a combination of MXene–Ti3C2Tx and nucleic acid aptamers. The prepared single‐layer or few‐layer MXene suspension is suction‐filtered onto MXene paper, which is cut to prepare MXene electrodes. To accomplish AFB1 specific detection, an amino‐labeled AFB1 aptamer is mounted on the surface of the carboxy‐functionalized MXene electrode. When AFB1 is present, it particularly binds to the aptamer to form a 3D structure, reducing the efficiency of electron transmission on the sensor surface. The difference in impedance signal change at the electrode/electrolyte interface is used to quantify AFB1. The results indicated that the detection range is 0.05–100 ng/mL, the detection limit is 0.04 ng/mL, and the recovery rate of AFB1 in corn samples is 97.8%–111.52% with the optimal detection conditions. The MXene paper‐based label‐free aptasensor is versatile and can detect different targets by simply swapping out the aptamers of different targets. The sensor also has a wide range of applications in food analysis and environmental testing. Practical applications: A paper‐based electrochemical aptamer sensor was developed to detect aflatoxin B1 using a combination of MXene–Ti3C2Tx and nucleic acid aptamers.The design is based on the preparation of MXene electrodes by pumping and filtering monolayer or multilayer MXene suspensions onto MXene paper and cutting.The MXene paper‐based label‐free aptamer sensor was designed to be versatile, allowing the detection of different targets by simply replacing the aptamer with one from a different targets. [ABSTRACT FROM AUTHOR]

There is growing interest in developing printable paper sensors to enable rapid testing of analytes for environmental, food safety, and clinical applications. A major challenge is to find suitable bioinks that are amenable to high-speed printing and remain functional after printing. We report on a simple and effective approach wherein an aqueous ink composed of megadalton-sized tandem repeating structure-switching DNA aptamers (concatemeric aptamers) is used to rapidly create patterned paper sensors on filter paper by inkjet printing. These concatemeric aptamer reporters remain immobilized at the point of printing through strong adsorption but retain sufficient segmental mobility to undergo structure switching and fluorescence signaling to provide both qualitative and quantitative detection of small molecules and protein targets. The convenience of inkjet printing allows for the patterning of internally referenced sensors with multiplexed detection, and provides a generic platform for on-demand printing of sensors even in remote locations. [ABSTRACT FROM AUTHOR]

Electrochemical aptamer‐based (E‐AB) sensors offer a powerful and general means for analyte detection in complex samples for various applications. Paper‐based E‐AB sensors could enable portable, low‐cost, and rapid detection of a broad range of targets, but it has proven challenging to fabricate suitable three‐electrode systems on paper. Here, we demonstrate a simple, economic, and environmentally friendly strategy for fabricating aptamer‐modified paper electrochemical devices (PEDs) via ambient vacuum filtration. The material, shape, size, and thickness of the three‐electrode PED system can be fully customized. We developed aptamer‐modified PEDs that enable sensitive and specific detection of small molecules in minimally processed biosamples. The sensitivity and stability of the PEDs are comparable to E‐AB sensors based on commercial gold electrodes. We believe our strategy can lead to the development of high performance PEDs for the on‐site detection of a variety of analytes. [ABSTRACT FROM AUTHOR]

Abstract: Micrometer‐sized functional nucleic acid (FNA) superstructures (denoted as 3D DNA) were examined as a unique class of biorecognition elements to produce highly functional bioactive paper surfaces. 3D DNA containing repeating sequences of either a DNA aptamer or DNAzyme was created from long‐chain products of rolling circle amplification followed by salt aging. The resulting 3D DNA retained its original spherical shape upon inkjet printing and adhered strongly to the paper surface via physisorption. 3D DNA paper sensors showed resistance to degradation by nucleases, suppressed nonspecific protein adsorption, and provided a much higher surface density of functional DNA relative to monomeric FNAs, making such species ideally suited for development of paper‐based biosensors. [ABSTRACT FROM AUTHOR]

Abstract: We report a paper‐based aptasensor platform that uses two reaction zones and a connecting bridge along with printed multifunctional bio/nano materials to achieve molecular recognition and signal amplification. Upon addition of analyte to the first zone, a fluorescently labelled DNA or RNA aptamer is desorbed from printed graphene oxide, rapidly producing an initial fluorescence signal. The released aptamer then flows to the second zone where it reacts with printed reagents to initiate rolling circle amplification, generating DNA amplicons containing a peroxidase‐mimicking DNAzyme, which produces a colorimetric readout that can be read in an equipment‐free manner or with a smartphone. The sensor was demonstrated using an RNA aptamer for adenosine triphosphate (a bacterial marker) and a DNA aptamer for glutamate dehydrogenase (Clostridium difficile marker) with excellent sensitivity and specificity. These targets could be detected in spiked serum or feacal samples, demonstrating the potential for testing clinical samples. [ABSTRACT FROM AUTHOR]

In this paper, a microfluidic paper-based analytical device is presented for sensitively detecting peptides based on the specific recognition of aptamer and the amplification strategy of hybridization chain reaction (HCR). The aptamer/initiation strand (AP/IS) was designed for the specific target recognition, and the presence of the target peptide could release the initiating sequence from the AP/IS. The released IS could trigger the HCR between two hairpin probes, which could complete the detection for the targets indirectly when the electrochemiluminescence (ECL) probe (Ru(phen)32+) is introduced into the double-strand DNA grooves,which could produce a significantly amplified ECL signal output. Mucin-1 was employed as a model peptide, and the detection limit of the proposed assay is calculated as 8.33 pMwith a range from 25 pM to 50 nM. The novel sensing platform can be widened for the detection of other kinds of peptides as well as potentials used for sensitively detecting other biomolecules, and even cells. [ABSTRACT FROM AUTHOR]

We report a method to detect proteins via suppression of rolling circle amplification (RCA) by using an appropriate aptamer as the linear primer (denoted as an aptaprimer) to initiate RCA. In the absence of a protein target, the aptaprimer is free to initiate RCA, which can produce long DNA products that are detected via binding of a fluorescent intercalating dye. Introduction of a target causes the primer region within the aptamer to become unavailable for binding to the circular template, inhibiting RCA. Using SYBR Gold or QuantiFluor dyes as fluorescent probes to bind to the RCA reaction product, it is possible to produce a generic protein‐modulated RCA assay system that does not require fluorophore‐ or biotin‐modified DNA species, substantially reducing complexity and cost of reagents. Based on this modulation of RCA, we demonstrate the ability to produce both solution and paper‐based assays for rapid and quantitative detection of proteins including platelet derived growth factor and thrombin. [ABSTRACT FROM AUTHOR]

Protein biomarkers often exist as degradation fragments in biological samples, and affinity agents derived using a purified protein may not recognize them, limiting their value for clinical diagnosis. Herein, we present a method to overcome this issue, by selecting aptamers against a degraded form of the toxin B protein, which is a marker for diagnosing toxigenic Clostridium difficile infections. This approach has led to isolation of a DNA aptamer that recognizes degraded toxin B, fresh toxin B, and toxin B spiked into human stool samples. DNA aptamers selected using intact recombinant toxin B failed to recognize degraded toxin B, which is the form present in stored stool samples. Using this new aptamer, we produced a simple paper‐based analytical device for colorimetric detection of toxin B in stool samples, or in the NAP1 strain of Clostridium difficile. The combined aptamer‐selection and paper‐sensing strategy can expand the practical utility of DNA aptamers in clinical diagnosis. [ABSTRACT FROM AUTHOR]

A sensitive and rapid method for the detection of bisphenol A (BPA) using general pH meters or pH paper was developed. Aptamer‐functionalized Fe3O4 magnetic nanoparticles (MNPs) were synthesized to capture BPA in samples, and gold NPs (AuNPs) were modified with acetylcholinesterase and the complementary strand of BPA aptamer. The specific detection of BPA was realized using the combination of BPA‐responsive, aptamer‐cross‐linked MNPs‐AuNPs complex. As the acetylcholine was hydrolyzed to choline and acetic acid, the pH of the solution decreased. The resulting pH change could be measured in real‐time using a pH meter. The sensitivity of detection of this assay was 5 pg/mL with the range of 0.01 to 1000 ng/mL. The pH change could be determined with the naked eye from the color change of a pH indicator strip. This sensor technology is promising and reliable for rapid, sensitive, and on‐site monitoring of BPA in water samples. [ABSTRACT FROM AUTHOR]

Lipopolysaccharides (endotoxins), found on Gram‐negative bacteria, can trigger a severe immune response in humans leading to septic shock and in extreme cases, even death. Therefore, the detection and neutralization of lipopolysaccharides (LPS) is of utmost importance in the pharmaceutical and medical industries. The United States Food and Drug Administration (US FDA) recommended detection method for LPS, the Limulus amebocyte lysate (LAL) assay, is expensive, time consuming, complex, and is prone to interference from proteases. As an alternative, this paper proposes a rapid, label‐free fluorescence‐based assay using LPS‐specific aptamers and the SYBR Green DNA stain. The proposed method has a detection limit of 0.1 ng/ml, which is sufficient to detect the permissible levels of LPS in many pharmaceutical drugs and medical products. The fluorescence signal was found to be a linear function of the concentration of LPS in the range from 0.1 ng/ml to 105 ng/ml. [ABSTRACT FROM AUTHOR]

This paper is an overview of the recent advances in nanoporous anodic alumina sensing platforms based on the interaction of oligonucleotides and antibodies with bioelements (e.g., virus, bacteria, or biomarkers) that can cause or identify diseases. Nanoporous alumina is the common platform material for all these sensing devices that have an enormous potential for the next‐generation bioanalytical systems. Nanoporous alumina is a cost‐effective material with morphological flexibility that provides high sensitivity to the sensing systems due to its large surface‐to‐volume ratio. Immunosensors transduce the signal produced by the specific antibody–antigen interaction. They are widely used for clinical analysis due to the high selectivity and sensitivity of their immunoreaction, transduced by different methods (electrochemical, optical, piezoelectric, etc). Another important, outstanding and more recently type of biosensors are based on the oligonucleotic functionalization of nanoporous alumina. Oligonucleotides can bind to a wide variety of elements (small organic molecules, proteins, or cells) with high selectivity, specificity, and affinity, even higher to those of antibodies. The basic concepts and mechanisms of their sensing methods for health care biosensing are introduced. Their transduction methods and great potential applications will be evaluated and classified in detail, together with a discussion of possible future trends. [ABSTRACT FROM AUTHOR]

Aptamers as the recognition element of the stochastic nanopore sensors have been under intense investigation. This paper reviews recent research advances in aptamer‐based nanopore sensing techniques, including the classification and selection of nanopores (biological nanopores, solid‐state nanopores, and nanopipettes), different strategies of aptamer‐based nanopore sensing, and their values and outlook for applications in areas such as environmental analysis, precision diagnosis, pharmaceutical industry, and security. Furthermore, the single‐molecule nanopore sensors have been applied to reveal the aptamer‐target interactions, such as recognition orientation, binding sites, and conformational heterogeneity dissociation kinetics at the single‐molecule level. In this review, recent research efforts to develop aptamer‐based single‐molecule nanopore sensors with high selectivity and sensitivity are highlighted, and some perspectives are drawn. [ABSTRACT FROM AUTHOR]

Melamine is an illegal additive in milk products. Mel‐34 is an aptamer specific for melamine. In this paper, we studied the base‐pair function in the stem regions of Mel‐34 using label‐free thiazole orange (TO) staining assay. Then, we used isothermal titration calorimetry (ITC) to verify the binding activity, and the fluorescence resonance energy transfer (FRET) biosensor based molecule‐beacon was established to detect melamine in yogurt sample. We found one of the mutants (mut6) has four folds higher affinity than Mel‐34, which indicated its critical binding domain was G18‐C25 base pair. We detected melamine in 1 % yogurt sample using mut6 as recognition molecule, the linear range was below 20 μM and the limit of detection was 4.09 μM which is below the melamine threshold (1.0 mg/kg) in milk products. This work calls for carefully study the critical binding domain to discover higher affinity aptamer specific for small molecule using label‐free method. [ABSTRACT FROM AUTHOR]

Recent advances in surface‐enhanced Raman spectroscopy (SERS) have resulted in multiplexing with unprecedented levels of sensitivity and selectivity in trace‐amount detection. However, quantification of multiple trace‐amount molecules with ng‐level accuracy has yet to be demonstrated due to nonuniform distribution of SERS enhancement and random adsorption of molecules at low concentrations. While Raman reporter‐free SERS is favorable for quantification in that the unique fingerprint spectra of molecules enable specific molecular identification, it has yet to be demonstrated due to poor reproducibility and insufficient SERS enhancement. Raman reporter‐free multiplex SERS with highly accurate quantification is successfully realized by versatile aptamer‐functionalized plasmonic Au nanogrids with uniform SERS enhancement. By cross‐point nano‐welding, monolithic Au nanogrids with excellent uniformity and high stability in aqueous media are produced. Raman reporter‐free multiplex detection and highly accurate quantification of concentration and composition is realized at picomolar levels. As a demonstration, Au nanogrids functionalized with bisphenol A‐specific aptamers successfully detect and quantify trace‐amounts of bisphenol A (8.49 ng) from thermal receipt paper. Moreover, principal component analysis is applied to multiplex SERS spectra to establish a ternary composition map, which can potentially serve as a practical reference for future Raman reporter‐free SERS. [ABSTRACT FROM AUTHOR]

Here we report on an ultra‐sensitive colorimetric sensing platform that takes advantage of both the strong amplification power of rolling circle amplification (RCA) and the high efficiency of a simple urease‐mediated litmus test. The presence of a target triggers the RCA reaction, and urease‐labelled DNA can hybridize to the biotinylated RCA products and be immobilized onto streptavidin‐coated magnetic beads. The urease‐laden beads are then used to hydrolyze urea, leading to an increase in pH that can be detected by a simple litmus test. We show this sensing platform can be easily integrated with aptamers for sensing diverse targets via the detection of human thrombin and platelet‐derived growth factor (PDGF) utilizing structure‐switching aptamers as well as SARS‐CoV‐2 in human saliva using a spike‐binding trimeric DNA aptamer. Furthermore, we demonstrate that this colorimetric sensing platform can be integrated into a simple paper‐based device for sensing applications. [ABSTRACT FROM AUTHOR]

Lead ion causes heavy metal pollution, which can cause serious damage to the ecological environment and human health. In this paper, a simple and sensitive photoelectrochemical method was designed for lead ion detection by using ZnO nanorods as the photoactive substrate and complementary strand aptamers as target recognition element. ZnO nanorods with favorable stability and photoelectric activity can provide a biocompatible microenvironment for aptamer loading. In the presence of lead ion, it would combine with the recognition aptamer S2 by competing with the aptamer S1, which led to changes in surface structure and charge density before and after the recognition of target, causing regular response of photocurrent. The developed method under optimized experimental conditions presented high sensitivity and good selectivity for lead ion detection with low detection limit as 3.5 × 10−11 mol/L. In addition, the proposed strategy was also further applied to the detection of lake water sample, which provided reliable experimental support and basis for further exploration and establishment of complex aptamer recognition and detection system. [ABSTRACT FROM AUTHOR]

In this paper, an electrochemical aptamer sensor was proposed for the highly sensitive detection of mercury ion (Hg2+). Carbon nanofiber (CNF) was prepared by electrospinning and high‐temperature carbonization, which was used for the loading of platinum nanoparticles (PtNPs) by the hydrothermal method. The Pt@CNF nanocomposite was modified on the surface of carbon ionic liquid electrode (CILE) to obtain Pt@CNF/CILE, which was further decorated by gold nanoparticles (AuNPs) through electrodeposition to get Au/Pt@CNF/CILE. Self‐assembling of the thiol‐based aptamer was further realized by the formation of Au‐S bond to get an electrochemical aptamer sensor (Aptamer/Au/Pt@CNF/CILE). Due to the specific binding of aptamer probe to Hg2+ with the formation of T‐Hg2+‐T structure, a highly sensitive quantitative detection of Hg2+ could be achieved by recording the changes of current signal after reacting with Hg2+ within the concentration range from 1.0 × 10−15 mol/L to 1.0 × 10−6 mol/L and the detection limit of 3.33 × 10−16 mol/L (3σ). Real water samples were successfully analyzed by this method. [ABSTRACT FROM AUTHOR]

BACKGROUND Residual antibiotics taken along with food consumed through the food chain are the main cause of the super‐bacteria and may damage organs such as liver and kidney. Therefore, monitoring residual antibiotic levels of products in the food chain is both important and a requirement. Maximum residual limits for kanamycin and neomycin are 150 ng mL−1 and 500 ng mL−1 respectively, which are challenging for most sensor platforms. In this paper, a novel method is presented for the determination of antibiotics residues in animal‐derived foods. RESULTS: Aptamer‐based kanamycin and neomycin biosensors based on the spectroscopic ellipsometer and the surface plasmon resonance‐enhanced total internal reflection ellipsometer methods as transducing element were developed. Detection limits of both sensor platforms were in the 0.1–1 nmol L−1 ranges, and the detection range was between the detection limit and 1000 nmol L−1. CONCLUSION: Both ellipsometry‐based aptasensors can be used as an alternative to the existing enzyme‐linked immunosorbent assay‐based method in terms of assay time (10 min), detection limit (0.22 ng mL−1 for neomycin and 0.048 ng mL−1 for kanamycin), and detection range. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Glycated albumin (GHSA) is a medium‐term glycaemic control marker of diabetes, which can be used as an alternative to or together with glycated haemoglobin (HbA1c). Currently available methods for the measurement of GHSA are limited in clinical practice because they involve slow and cumbersome processes of sample preparation, proteolytic digestion, and thermal incubation, and they suffer from limited analytical performance, and/or a lack of normalization to total albumin (HSA) levels. In this paper, we developed a simple electrochemical biosensor to measure GHSA values based on two DNA aptamers that specifically bind to GHSA and HSA. We used square wave voltammetry (SWV) to measure binding of the target proteins to their specific biotinylated aptamers, which had been immobilised on separate streptavidin (STR)‐modified screen‐printed carbon electrodes (SPCEs), in the presence of the redox mediator ferricyanide (Fe(CN)63−). This electrochemical aptasensing system had a detection limit of 3 ng/ml for GHSA and 0.2 μg/ml for HSA. The results exhibited high selectivity for GHSA over other molecules present in the blood. The developed sensor was able to measure the amount of GHSA in plasma samples. A statistically significant difference was observed in the elevated plasma GHSA levels in diabetic versus non‐diabetic patients. Moreover, the trends in these GHSA levels were consistent with those obtained using the HbA1c test. The sensing system reported herein could be applied as a point‐of‐care‐testing (POCT) device for measurements of clinically relevant GHSA values. [ABSTRACT FROM AUTHOR]

Raman spectroscopy is of course a well-established analytical technique due to its ability to provide highly specific Raman fingerprints for purposes of identifying molecular components in simple and complex samples. Coherent Raman spectroscopies can offer significant advantages over spontaneous Raman observations for some applications, as has been well established since the commercial accessibility of high-powered lasers in the visible. Central to this work is a machine learning-assisted low-frequency Raman based methodology, informed by DFT results, that can characterize the twist angle of these thin layered samples. [Extracted from the article]

DNA nanostructures exhibit versatile geometries and possess sophisticated capabilities not found in other nanomaterials. They serve as customizable nanoplatforms for orchestrating the spatial arrangement of molecular components, such as biomolecules, antibodies, or synthetic nanomaterials. This is achieved by incorporating oligonucleotides into the design of the nanostructure. In the realm of drug delivery to cancer cells, there is a growing interest in active targeting assays to enhance efficacy and selectivity. The active targeting approach involves a "key‐lock" mechanism where the carrier, through its ligand, recognizes specific receptors on tumor cells, facilitating the release of drugs. Various DNA nanostructures, including DNA origami, Tetrahedral, nanoflower, cruciform, nanostar, nanocentipede, and nanococklebur, can traverse the lipid layer of the cell membrane, allowing cargo delivery to the nucleus. Aptamers, easily formed in vitro, are recognized for their targeted delivery capabilities due to their high selectivity for specific targets and low immunogenicity. This review provides a comprehensive overview of recent advancements in the formation and modification of aptamer‐modified DNA nanostructures within drug delivery systems. [ABSTRACT FROM AUTHOR]

Rücktitelbild: In Vitro Selection of a DNA Aptamer Targeting Degraded Protein Fragments for Biosensing (Angew. Keywords: aptamers; biosensors; in vitro selection; paper-based analytical devices; protein degradation EN aptamers biosensors in vitro selection paper-based analytical devices protein degradation 8042 8042 1 05/06/20 20200511 NES 200511 B Ein DNA-Aptamer b , das auf natürlich abgebaute Proteinfragmente abzielt, wurde von M. Liu, J. D. Brennan, Y. Li und Mitarbeitern selektiert (siehe die Zuschrift auf S. 7780). Aptamers, biosensors, in vitro selection, paper-based analytical devices, protein degradation. [Extracted from the article]

Sudan dye is generally used as a coloring agent in textile and cosmetic industries. However, Sudan dye exhibits potentials to cause cancer, toxicity, and allergy because of its metabolites including amines and naphthol related substances. Despite banning Sudan dye, the Sudan dye is used by many unscrupulous food manufacturers for enhancing food appearance. This paper introduces a new aptamer based on Sudan III detection methods. An in vitro selection and amplification method named SELEX (Systematic Evolution of Ligands by Exponential Enrichment) is employed to screen out Sudan III specific aptamer. The desired aptamer is obtained after 12 rounds of screening and cloning. To establish aptamer affinity and specificity for Sudan III, ELONA (enzyme‐linked oligonucleotide assay) and dot blot analyses are performed. ELONA assay indicates that 100 nM is the optimum concentration of the aptamer for Sudan III detection, while the minimum detection limit of Sudan III is 4 ng/well. The aptamer exhibits stable secondary structure with the stem‐loop and G‐quadruplex in the main structure. The selected single ssDNA aptamer C07 demonstrates binding affinity and the highest specificity to Sudan III (C07: KD = 22.37 ± 3.943 nmol L−1). Based on the results, a good correlation is observed between aptamer C07 and Sudan III and it is concluded that the new Sudan III specific aptamer has potential to be applied for rapid and accurate detection of the Sudan III in food products. Practical Applications: Examination of the chili sauce sample in a simulation experiment indicates that the aptamer C07 can be applied to rapid detection in foods. The nucleic acid aptamers‐based food detection method has the advantages of high sensitivity, specificity, easy operation, short detection time, and low cost. It can realize the rapid detection of illegal additives in foods, which has a great significance in the field of food safety research. The SELEX method is used to select a new Sudan III specific aptamer. ELONA and Dot blot assays further suggest that aptamer C07 can be used for selective detection of Sudan III. Therefore, the detection in chili sauce samples offers feasibility for more complex samples. [ABSTRACT FROM AUTHOR]

Hemin/G‐quadruplex (hGQ) DNAzymes are horseradish peroxidase‐mimicking catalysts capable of the oxidation of a variety of substrates. We now implement aptamer‐functionalized hGQ DNAzymes, also known as nucleoapzymes, to achieve increased bioconjugation of N‐methyl luminol to tyrosine‐containing residues and peptides. We found that the presence of a tyrosinamide‐binding aptamer leads to a 12‐fold increase in the catalytic rate constant (kcat), and the saturation kinetics curves that were obtained provide evidence for the involvement of the substrate binding site in the reaction. The application of the best performing nucleoapzymes for the modification of Tyr‐containing peptides reveals that (i) the aptamer also recognizes the ligand structure when this is embedded in a larger peptide structure, and (ii) distant residues in the peptide substrate can influence the conversion. As such, we show that nucleoapzymes display enzyme‐like features and provide an additional tool in the toolbox of bioconjugation chemistry. [ABSTRACT FROM AUTHOR]

We report a generalizable strategy for biosensing that takes advantage of the resistance of DNA aptamers against nuclease digestion when bound with their targets, coupled with toehold mediated strand displacement (TMSD) and rolling circle amplification (RCA). A DNA aptamer containing a toehold extension at its 5′‐end protects it from 3′‐exonuclease digestion by phi29 DNA polymerase (phi29 DP) in a concentration‐dependent manner. The protected aptamer can participate in RCA in the presence of a circular template that is designed to free the aptamer from its target via TMSD. The absence of the target leads to aptamer digestion, and thus no RCA product is produced, resulting in a turn‐on sensor. Using two different DNA aptamers, we demonstrate rapid and quantitative real‐time fluorescence detection of two human proteins: platelet‐derived growth factor (PDGF) and thrombin. Sensitive detection of PDGF was also achieved in human serum and human plasma, demonstrating the selectivity of the assay. [ABSTRACT FROM AUTHOR]

DNA computation is considered a fascinating alternative to silicon‐based computers; it has evoked substantial attention and made rapid advances. Besides realizing versatile functions, implementing spatiotemporal control of logic operations, especially at the cellular level, is also of great significance to the development of DNA computation. However, developing simple and efficient methods to restrict DNA logic gates performing in live cells is still a challenge. In this work, a series of DNA logic gates was designed by taking full advantage of the diversity and programmability of the G‐quadruplex (G4) structure. More importantly, by further using the high affinity and specific endocytosis of cells to aptamer G4, an INHIBIT logic gate has been realized whose operational site is precisely restricted to specific live cells. The design strategy might have great potential in the field of molecular computation and smart bio‐applications. [ABSTRACT FROM AUTHOR]

DNA‐encoded chemical libraries are typically screened against purified protein targets. Recently, cell‐based selections with encoded chemical libraries have been described, commonly revealing suboptimal performance due to insufficient recovery of binding molecules. We used carbonic anhydrase IX (CAIX)‐expressing tumor cells as a model system to optimize selection procedures with code‐specific quantitative polymerase chain reaction (qPCR) as selection readout. Salt concentration and performing PCR on cell suspension had the biggest impact on selection performance, leading to 15‐fold enrichment factors for high‐affinity monovalent CAIX binders (acetazolamide; KD=8.7 nM). Surprisingly, the homobivalent display of acetazolamide at the extremities of both complementary DNA strands led to a substantial improvement of both ligand recovery and enrichment factors (above 100‐fold). The optimized procedures were used for selections with a DNA‐encoded chemical library comprising 1 million members against tumor cell lines expressing CAIX, leading to a preferential recovery of known and new ligands against this validated tumor‐associated target. This work may facilitate future affinity selections on cells against target proteins which might be difficult to express otherwise. [ABSTRACT FROM AUTHOR]

Lung cancer is the first malignant tumor with morbidity and mortality in the world. To identify a novel lung cancer biomarker facilitating early detection lung cancer, we had obtained a nucleic acid aptamer (Apt‐5) that specifically recognized lung cancer serum. An affinity purification experiment by using magnetic nanoparticles was used to pull down the target proteins, protein CLEC3B was subsequently identified as the target of the aptamer Apt‐5 by MS analysis. The aptamer Apt‐5 displayed high affinity to the CLEC3B protein with a dissociation constant (KD) of 23.9±5.5 nM by SPR. The specificity and selectivity of the CLEC3B protein with the aptamer Apt‐5 were verified by EMSA. It was further confirmed by ELISA experiments that the average concentration of CLEC3B in lung cancer serum was higher than that in healthy human serum (3.272±0.1732 ng/mL vs. 2.104±0.1174 ng/mL, P<0.001.). Moreover, the sensitivity and specificity of ELISA assay in this study are 90 % and 80 %, respectively. So the protein CLEC3B might serve as a potential lung cancer biomarker for early diagnosis and therapy. [ABSTRACT FROM AUTHOR]

Pathogenic and corrosive bacteria pose a significant risk to human health or economic well‐being. The specific, sensitive, and on‐site detection of these bacteria is thus of paramount significance but remains challenging. Taking inspiration from immunoassays with primary and secondary antibodies, we describe here a rational design of microbial sensor (MS) under a dual‐specificity recognition strategy using Pseudomonas aeruginosa (P. aeruginosa) as the detection model. In the MS, engineered aptamers are served as the primary recognition element, while polydopamine‐N‐acetyl‐D‐galactosamine (PDA‐Gal NAc) nanoparticles are employed as the secondary recognition element, which will also generate and amplify changes in the output voltage signal. To achieve self‐powering capability, the MS is constructed based on a triboelectric nanogenerator (TENG) with the specific aptamers immobilized on the TENG electrode surface. The as‐prepared MS‐TENG system exhibits good stability in output performance under external forces, and high specificity toward P. aeruginosa, with no cross‐reactivity observed. A linear relationship (R2 = 0.995) between the output voltage and P. aeruginosa concentration is established, with a limit of detection estimated at around 8.7 × 103 CFU mL−1. The utilization of PDA‐Gal NAc nanoparticles is found to play an important role in enhancing the specific and reliability of detection, and the underlying mechanisms are further clarified by computational simulations. In addition, the MS‐TENG integrates a wireless communication module, enabling real‐time monitoring of bacterial concentration on mobile devices. This work introduces a pioneering approach to designing self‐powered smart microbial sensors with high specificity, using a double recognition strategy applicable to various bacteria beyond P. aeruginosa. [ABSTRACT FROM AUTHOR]

Periodontal disease is one of the most common forms of inflammation. It is currently diagnosed by observing symptoms such as gingival bleeding and attachment loss. However, the detection of biomarkers that precede such symptoms would allow earlier diagnosis and prevention. Aptamers are short oligonucleotides or peptides that fold into three‐dimensional conformations conferring the ability to bind molecular targets with high affinity and specificity. Here we report the selection of aptamers that bind specifically to the bacterium Tannerella forsythia, a pathogen frequently associated with periodontal disease. Two aptamers with the highest affinity were examined in more detail, revealing that their binding is probably dependent on mirolysin, a surface‐associated protease secreted by the T. forsythia type‐9 secretion system. The aptamers showed minimal cross‐reactivity to other periodontopathogens and are therefore promising leads for the development of new tools to study the composition of the periodontitis‐associated dysbiotic bacteriome as well as inexpensive new diagnostic assays. [ABSTRACT FROM AUTHOR]

Extracellular vesicles and particles (EVPs) are recognized as ideal liquid biopsy tools for cancer detection, and membrane proteins are commonly used EVP biomarkers. However, bulk analysis of EVP membrane protein biomarkers typically fails to meet the clinical requirement for diagnostic accuracy. We investigated the correlation between the membrane protein expression level, the binding kinetics to aptamers and the sizes of EVPs with interferometric plasmonic microscopy (iPM), and demonstrated the implementation of the correlative signature to determine cancer types. Using EVPs collected from both cell model and clinical plasma samples with liver, lung, breast, or prostate cancer, we found that the selective set of membrane protein expression levels of five protein markers and their binding kinetics were highly heterogeneous across various sizes of EVPs, resulting in the low overall accuracy (<50%) in cancer classification with bulk analysis of all populations. By grouping the EVPs into three subpopulations according to their sizes, the overall accuracy could be increased to about 70%. We further grouped the EVPs into subpopulations with a 10 nm interval in sizes and analysed the correlation between the membrane proteins and sizes with a machine learning algorithm. The results show that the overall accuracy to discriminate cancer types could be improved to 85%. Therefore, this work highlights the significance of size‐dependent subtyping of EVPs and suggests that the correlation between the selective set of membrane proteins and sizes of EVP can serve as a signature for clinical cancer diagnosis. [ABSTRACT FROM AUTHOR]

In this paper, a novel strategy of electrochemical amplified detection of thrombin based on G-quadruplex-linked supersandwich structure was described. In the presence of K+ and hemin, the original hairpin DNA sequence activated an autonomous cross-opening process to build up hemin/G-quadruplex structure and can hybridize to form supersandwich structure containing multiple signal labels. With the addition of thrombin, it conjugated with its aptamer, leading to a remarkably descended signal. The supersandwich-amplified electrochemical sensor system was highly sensitive in the concentration range from 10−6 to 10−10 M with a detection limit of 10 pM and also demonstrated excellent selectivity. The amplifying supersandwich structure with multiple labels can be implemented as a versatile sensing platform for analyzing other DNA in the presence of the appropriate probe. [ABSTRACT FROM AUTHOR]

This paper presents the first example of a sensitive, selective, and stable phosphorescent sensor based on phosphorescence energy transfer (PET) for thrombin that functions through thrombin-aptamer recognition events. In this work, an efficient PET donor-acceptor pair using Mn-doped ZnS quantum dots labeled with thrombin-binding aptamers (TBA QDs) as donors, and carbon nanodots (CNDs) as acceptors has been constructed. Due to the π-π stacking interaction between aptamer and CNDs, the energy donor and acceptor are taken into close proximity, leading to the phosphorescence quenching of donors, TBA QDs. A maximum phosphorescence quenching efficiency as high as 95.9 % is acquired. With the introduction of thrombin to the 'off state' of the TBA-QDs-CNDs system, the phosphorescence is 'turned on' due to the formation of quadruplex-thrombin complexes, which releases the energy acceptor CNDs from the energy donors. Based on the restored phosphorescence, an aptamer-based turn-on thrombin biosensor has been demonstrated by using the phosphorescence as a signal transduction method. The sensor displays a linear range of 0-40 n M for thrombin, with a detection limit as low as 0.013 n M in pure buffers. The proposed aptasensor has also been used to monitor thrombin in complex biological fluids, including serum and plasma, with satisfactory recovery ranging from 96.8 to 104.3 %. This is the first time that Mn-doped ZnS quantum dots and CNDs have been employed as a donor-acceptor pair to construct PET-based biosensors, which combines both the photophysical merits of phosphorescence QDs and the superquenching ability of CNDs and thus affords excellent analytical performance. We believe this proposed method could pave the way to a new design of biosensors using PET systems. [ABSTRACT FROM AUTHOR]

Optical fiber biochemical sensing technology, as a cutting‐edge, multidisciplinary, and universal sensing technology, is gradually becoming the research focus in the field of biochemical sensing. They enjoy distinctive characteristics such as high cost‐effectiveness, flexibility, resistance to electromagnetic interference, and small size and can be used to detect biochemical analytes at ultra‐low concentrations with ultra‐low sample consumption, even when these biochemical analytes are in hard‐to‐reach environments. An aptamer is a DNA or RNA oligonucleotide fragment with a high affinity for the target selected from random sequences. Due to their advantages of simple synthesis, low cost, good stability, good specificity, and strong affinity, they are gradually becoming ideal recognition elements and may replace antibodies as next‐generation target receptors. Therefore, aptamers, as functional materials, are integrated into optical fiber sensors, which are called optical fiber aptasensors, providing new opportunities for the construction of high‐performance and intelligent biochemical sensors. This article aims to review the recent research progress of fiber‐optic aptamer biochemical sensors, with a focus on their sensing mechanisms and applications. Finally, based on the existing deficiencies of fiber‐optic aptamer biochemical sensors and practical requirements, their future development trends are briefly discussed. [ABSTRACT FROM AUTHOR]

Background: Aflatoxin B1 (AFB1) has been considered as a prevailing natural carcinogenic mycotoxin in various food and feedstuff products, posing serious endangers to human health. The quantitative and selective detection of AFB1 was significantly essential for food safety control. Results: We developed a label‐free fluorescent aptasensor using a target‐binding aptamer probe as the rolling circle amplification (RCA) primer for AFB1 detection. In the presence of AFB1, the probe preferentially combined with AFB1 rather than binding to the circular template, inhibiting the initiation of RCA process. The detection limit of the proposed method was as low as 6.6 nM, and the linear dynamic response range was four orders of magnitude wide. Besides, it revealed excellent specificity for AFB1 and high‐quality detection performance in food samples. Conclusion: The presented detection method was simple, rapid, and economical, which indicated a promising application in food safety monitoring. [ABSTRACT FROM AUTHOR]

Cancer is among the leading causes of death and an important barrier to improving life expectancy globally. Prostate cancer is the second most common cancer in men worldwide. The detection of biomarkers in body fluids is the key topic for the diagnosis and prognosis of prostate cancer. Despite advances in prostate cancer detection methods, therapeutic agents and new biomarkers, prostate cancer remains a serious challenge. Prostate‐specific antigen (PSA) is widely recognized as an important biomarker for the diagnosis of prostate cancer although researchers have also investigated the use of alternative biomarkers. Design and development of novel biosensors for prostate cancer detection has become a hot research area with advances in nanotechnology aiding biosensor development. This article reviews recent achievements and progress that nanomaterials and nanotechnology have made in biomarkers based biosensors for prostate cancer detection and covers: i) PSA‐targeted biosensors (immunosensors, aptamer‐based, peptide‐based and nanopore‐based biosensors), ii) sarcosine oxidase‐targeted biosensors, iii) other biomarkers based biosensors (prostate cancer antigen 3 (PCA3), vascular endothelial growth factor (VEGF) and prostate‐specific membrane antigen (PSMA)) including dual biomarkers based biosensors (PSA‐VEGF, PSA‐PSMA, PSA‐PCA3 and PSA‐sarcosine). The aim of this review is to provide insights into how nanomaterials in combination with various biomarkers are now aiding biosensor development in prostate cancer diagnostics. [ABSTRACT FROM AUTHOR]

Electrochemical aptasensors can detect cancer biomarkers such as mucin 1 (MUC1) to provide point-of-care diagnosis that is low-cost, specific and sensitive. Herein, a DNA hairpin containing MUC1 aptamer was thiolated, conjugated with methylene blue (MB) redox tag, and immobilized on a gold electrode by self-assembly. The fabrication process was characterized by scanning electron microscopy, X-ray spectroscopy analysis and electrochemistry techniques. The results evidenced a stable and sensitivity sensor presenting wide linear detection range (0.65-110 ng/mL). Therefore, it was able to precisely detect MUC1 production patterns in normal (RWPE-1) and prostate cancer cells (LNCaP and PC3). The biosensor has ability to detect MUC1 in complex samples being an efficient and useful platform for cancer diagnosing in early stages and for physiological applications such as cancer treatment monitoring. [ABSTRACT FROM AUTHOR]

Comprehensive Summary: Sensitive and specific imaging of tumor‐associated biomarkers offers an effective way for cancer diagnosis. However, most of the reported imaging probes are limited to the targeting of a single biomarker, which may impair imaging precision. Here, we report an aptamer‐based, AND‐gate DNA nanodevice that relies on the sequential processing of two tumor‐associated proteins to produce a specific fluorescent signal for precise tumor imaging. The DNA nanodevice integrates an AS1411 aptamer to bind cell surface nucleolin for tumor targeting and a DNA sensor to respond to intracellular apurinic/apyrimidinic endonuclease 1 for activatable imaging. We evaluated the performance of the system in both living cells and a tumor‐bearing mouse model, and demonstrated its capability in tumor cell‐specific imaging. We anticipate that this strategy will accelerate the design of multivariate protein‐stimulated probes for tumor diagnosis. [ABSTRACT FROM AUTHOR]

Virus recognition has been driven to the forefront of molecular recognition research due to the COVID‐19 pandemic. Development of highly sensitive recognition elements, both natural and synthetic is critical to facing such a global issue. However, as viruses mutate, it is possible for their recognition to wane through changes in the target substrate, which can lead to detection avoidance and increased false negatives. Likewise, the ability to detect specific variants is of great interest for clinical analysis of all viruses. Here, a hybrid aptamer‐molecularly imprinted polymer (aptaMIP), that maintains selective recognition for the spike protein template across various mutations, while improving performance over individual aptamer or MIP components (which themselves demonstrate excellent performance). The aptaMIP exhibits an equilibrium dissociation constant of 1.61 nM toward its template which matches or exceeds published examples of imprinting of the spike protein. The work here demonstrates that "fixing" the aptamer within a polymeric scaffold increases its capability to selectivity recognize its original target and points toward a methodology that will allow variant selective molecular recognition with exceptional affinity. [ABSTRACT FROM AUTHOR]

Comprehensive Summary: Nucleic acids are the hereditary information storage medium of life. Due to its high programmability and good biocompatibility, the applications of nucleic acids are not limited to their natural function. However, the efficiency of nucleic acids is often hampered by inherent limitations in numerous practical applications, including limited access to complex functional patterns due to only four building blocks, facile degradation by nucleases, rapid renal clearance, poor pharmacokinetic properties, and so on. To end this, the researchers developed unnatural base pairs (UBPs) and numerous artificial analogues of nucleosides and oligonucleotides in recent decades. The developed UBPs and nucleoside base analogues together constitute artificial nucleobase compilation, which promotes the development of biomedical sciences. Here, we describe the development of artificial nucleobase compilation and summarized its applications in precise molecular medicine, including PCR‐based diagnostics, aptamer‐based diagnostics, nucleobase analogue drugs construction, ASO modification, aptamer‐based therapeutics and biomaterials construction. This review provides an overview of current opportunities and challenges of artificial nucleobase‐related precise molecular medicine. [ABSTRACT FROM AUTHOR]

Cortisol is a steroid hormone naturally produced by the adrenal glands. It participates in and controls several processes in the body and is considered an important physiological biomarker. Due to its very low concentrations in body fluids, its detection requires high sensitivity and specificity. Here, we present a simple electrochemical biosensor based on reduced graphene oxide (rGO) modified with gold nanoparticles (AuNPs) for the immobilization of the cortisol‐specific aptamer (Ap) (MCH‐Ap‐AurGO/GCE). Important analytical parameters for identifying the target analyte were optimized, such as conditions and amount of immobilized Ap, the influence of the concentration and nature of the supporting electrolyte, pH of the medium, and incubation time. The optimized conditions for the aptasensor were: concentration of Ap 1.0 × 10−6 mol L−1, support electrolyte Tris/HCl 50 mmol L−1, MgCl2 10 mmol L−1, and NaCl 10 mmol L−1, at pH 5.0 and incubation time of 15 min. A linear response range was obtained from 1 × 10−18 up to 1 × 10−11 mol L−1 of cortisol with a detection limit (LOD) of 1.0 × 10−18 mol L−1. A curve adjusted for operational purposes in the saliva sample was fitted for the concentration range between 0.5 × 10−14 and 1 × 10−11 mol L−1, with a linear regression equation ΔRtc/Rtc1 = 2.70 + 0.17 × log([Cortisol]). The aptasensor demonstrated a great potential for detecting cortisol in a simple, fast, and highly sensitive way, opening its path for application in real samples, which present levels below the concentration in which cortisol is commonly found in body fluids. [ABSTRACT FROM AUTHOR]

In this study, we exploit a recently developed fragment diabatization‐based excitonic model, FrDEx, to simulate the electronic circular dichroism (ECD) spectra of three guanine‐rich DNA sequences arranged in guanine quadruple helices with different topologies: thrombin binding aptamer (antiparallel), c‐Myc promoter (parallel), and human telomeric sequence (3+1 hybrid). Starting from time‐dependent density functional theory (TD‐DFT) calculations with the M052X functional, we apply our protocol to parameterize the FrDEX Hamiltonian, which accounts for electron density overlap and includes both the coupling with charge transfer transitions and the effect of the surrounding bases on the local excitation of each chromophore. The TD‐DFT/M052X spectral shapes are in good agreement with the experimental ones, the main source of discrepancy being related to the intrinsic error on the computed transition energies of guanine monomer. FrDEx spectra are fairly close to the reference TD‐DFT ones, allowing a significant advance with respect to a more standard excitonic Hamiltonian. We also show that the ECD spectra are sensitive to the inclusion of the inner K + cation in the calculation. [ABSTRACT FROM AUTHOR]

Paraquat is a widely used herbicide for controlling weeds and grasses in agriculture, and its contaminated residues in agricultural areas are of increasing concern. This work reports the development of the sensitive and easy‐to‐use colorimetric aptasensor for screening paraquat residues in agricultural soil. The short DNA fragments derived from the original aptamer were analyzed for their capability to interact with paraquat by molecular dynamic simulation. The paraquat‐aptasensor was developed using the selected DNA fragment and gold nanoparticles. Its limit of detection (LOD) for paraquat is 2.76 nM, which is more sensitive than the aptasensor with long‐length aptamer (LOD = 12.98 nM). The developed aptasensor shows the selectivity to paraquat, but not to other tested herbicides; ametryn, atrazine, difenzoquat, 2,4‐D‐dimethyl ammonium, and glufosinate. The recovery rates of paraquat detection in the spiked soil samples were in a range of 99.5%–105.1%, with relative standard deviation values of <4%. The developed aptasensor was used to screen for paraquat residues in agricultural soils, and three out of 23 soil samples were tested positive for paraquat, which was confirmed by a high‐performance liquid chromatography analysis. These results suggested the potential application of the developed aptasensor to detect paraquat residues in agricultural sites. [ABSTRACT FROM AUTHOR]

L‐lactate is a key metabolite indicative of physiological states, glycolysis pathways, and various diseases such as sepsis, heart attack, lactate acidosis, and cancer. Detection of lactate has been relying on a few enzymes that need additional oxidants. In this work, DNA aptamers for L‐lactate were obtained using a library‐immobilization selection method and the highest affinity aptamer reached a Kd of 0.43 mM as determined using isothermal titration calorimetry. The aptamers showed up to 50‐fold selectivity for L‐lactate over D‐lactate and had little responses to other closely related analogs such as pyruvate or 3‐hydroxybutyrate. A fluorescent biosensor based on the strand displacement method showed a limit of detection of 0.55 mM L‐lactate, and the sensor worked in 90 % serum. Simultaneous detection of L‐lactate and D‐glucose in the same solution was achieved. This work has broadened the scope of aptamers to simple metabolites and provided a useful probe for continuous and multiplexed monitoring. [ABSTRACT FROM AUTHOR]

In this study, a simple electrochemical sensing platform with the employment of a bivalent binding aptamer‐cDNA probe (BBA‐cDNA) structure is constructed for the detection of aflatoxin B1 (AFB1) as a mycotoxin. The BBA‐cDNA structure is composed of two strands of aptamer (Apts) and their complementary strand (CS). Using a simple but accurate design, the presented measurement approach showed enhanced sensitivity and selectivity for AFB1 detection with a LOD of 0.1 ng/mL. The approach presented in this study can be applied to the development of biosensors for the measurement of various toxins by substituting the proper aptamers and complementary strands. [ABSTRACT FROM AUTHOR]

Comprehensive Summary: The applicability of G‐quadruplexes (G4s) as antiviral targets, therapeutic agents and diagnostic tools for coronavirus disease 2019 (COVID‐19) is currently being evaluated, which has drawn the extensive attention of the scientific community. During the COVID‐19 pandemic, research in this field is rapidly accumulating. In this review, we summarize the latest achievements and breakthroughs in the use of G4s as antiviral targets, therapeutic agents and diagnostic tools for COVID‐19, particularly using G4 ligands. Finally, strength and weakness regarding G4s in anti‐SARS‐CoV‐2 field are highlighted for prospective future projects. [ABSTRACT FROM AUTHOR]

Cell‐specific aptamers offer a powerful tool to study membrane receptors at the single‐molecule level. Most target receptors of aptamers are highly expressed on the cell surface, but difficult to analyze in situ because of dense distribution and fast velocity. Therefore, we herein propose a random sampling‐based analysis strategy termed ligand dilution analysis (LDA) for easily implemented aptamer‐based receptor study. Receptor density on the cell surface can be calculated based on a regression model. By using a synergistic ligand dilution design, colocalization and differentiation of aptamer and monoclonal antibody (mAb) binding on a single receptor can be realized. Once this is accomplished, precise binding site and detailed aptamer‐receptor binding mode can be further determined using molecular docking and molecular dynamics simulation. The ligand dilution strategy also sets the stage for an aptamer‐based dynamics analysis of two‐ and three‐dimensional motion and fluctuation of highly expressed receptors on the live cell membrane. [ABSTRACT FROM AUTHOR]

Electrochemical biosensors based on structure‐switching aptamers offer many advantages because they can operate directly in complex samples and offer the potential to integrate with miniaturized electronics. Unfortunately, these biosensors often suffer from cross‐reactivity problems when measuring a target in samples containing other chemically similar molecules, such as precursors or metabolites. While some progress has been made in selecting highly specific aptamers, the discovery of these reagents remains slow and costly. In this work, a novel strategy is demonstrated to distinguish molecules with miniscule difference in chemical composition (such as a single hydroxyl group)—with cross reactive aptamer probes—by tuning the charge state of the surface on which the aptamer probes are immobilized. As an exemplar, it is shown that the strategy can distinguish between DOX and many structurally similar analytes, including its primary metabolite doxorubicinol (DOXol). Then the ability to accurately quantify mixtures of these two molecules based on their differential response to sensors with different surface‐charge properties is demonstrated. It is believed that this methodology is general and can be extended to a broad range of applications. [ABSTRACT FROM AUTHOR]