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Developing cost-effective, consumer-accessible platforms for point-of-use environmental and clinical pathogen testing is a priority, to reduce reliance on laborious, time-consuming culturing approaches. Unfortunately, a system offering ultrasensitive detection capabilities in a form that requires little auxiliary equipment or training has remained elusive. Here, a colorimetric DNAzyme-crosslinked hydrogel sensor is presented. In the presence of a target pathogen, DNAzyme cleavage results in hydrogel dissolution, yielding the release of entrapped gold nanoparticles in a manner visible to the naked eye. Recognizing that Escherichia coli holds high relevance within both environmental and clinical environments, an E. coli-responsive DNAzyme is incorporated into this platform. Through the optimization of the hydrogel polymerization process and the discovery of bacteriophage-induced DNAzyme signal amplification, 10 1 CFU mL -1 E. coli is detected within real-world lake water samples. Subsequent pairing with an artificial intelligence model removed ambiguity in sensor readout, offering 96% true positive and 100% true negative accuracy. Finally, high sensor specificity and stability results supported clinical use, where 100% of urine samples collected from patients with E. coli urinary tract infections are accurately identified. No false positives are observed when testing healthy samples. Ultimately, this platform stands to significantly improve population health by substantially increasing pathogen testing accessibility., (© 2024 The Author(s). Advanced Materials published by Wiley‐VCH GmbH.)

Many aptamers have been generated by systematic evolution of ligands by exponential enrichment (SELEX) to recognize spike proteins of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2&ek), some of which have been engineered into dimeric and trimeric versions for enhanced affinity for diagnostic applications. However, no studies have been conducted to compare the utilities of monomeric, dimeric and trimeric aptamers in diagnostic assays with real clinical samples to answer the question of what levels of affinity an aptamer must have for accurate clinical diagnostics. Herein, we carried out a comparative study with two monomeric aptamers MSA1 and MSA5, one dimeric aptamer and two homotrimeric aptamers constructed with MSA1 and MSA5, with affinity varying by 1000-fold. Using a colorimetric sandwich assay to analyze 48 human saliva samples, we found that the trimeric aptamer assay (K d ≈10 pM) can identify the SARS-CoV-2 infection much more accurately than the dimeric aptamer assay (K d ≈100 pM) and monomeric aptamer assay (K d ≈10,000 pM). Based on the experimental data, we theoretically predict the levels of affinity an aptamer needs to possess to achieve 80-100 % sensitivity and 100 % specificity. The findings from this study highlight the need for deriving very high affinity aptamers to enable highly accurate detection of viral infection for future pandemics., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The detection of pathogens is critical for preventing and controlling health hazards across clinical, environmental, and food safety sectors. Functional nucleic acids (FNAs), such as aptamers and DNAzymes, have emerged as versatile molecular tools for pathogen detection due to their high specificity and affinity. This review focuses on the in vitro selection of FNAs for pathogens, with emphasis on the selection of aptamers for specific biomarkers and intact pathogens, including bacteria and viruses. Additionally, the selection of DNAzymes for bacterial detection is discussed. The integration of these FNAs into colorimetric biosensors has enabled the development of simple, cost-effective diagnostic platforms. Both non-catalytic and catalytic colorimetric biosensors are explored, including those based on gold nanoparticles, polydiacetylenes, protein enzymes, G-quadruplexes, and nanozymes. These biosensors offer visible detection through color changes, making them ideal for point-of-care diagnostics. The review concludes by highlighting current challenges and future perspectives for advancing FNA-based colorimetric biosensing technologies for pathogen detection., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

An on-going challenge with COVID-19, which has huge implications for future pandemics, is the rapid emergence of viral variants that makes diagnostic tools less accurate, calling for rapid identification of recognition elements for detecting new variants caused by mutations. We hypothesize that we can fight mutations of the viruses with mutations of existing recognition elements. We demonstrate this concept via rapidly evolving an existing DNA aptamer originally selected for the spike protein (S-protein) of wildtype SARS-CoV-2 to enhance the interaction with the same protein of the Omicron variants. The new aptamer, MBA5SA1, has acquired 22 mutations within its 40-nucleotide core sequence and improved its binding affinity for the S-proteins of diverse Omicron subvariants by >100-fold compared to its parental aptamer (improved from nanomolar to picomolar affinity). Deep sequencing analysis reveals dynamic competitions among several MBA5SA1 variants in response to increasing selection pressure imposed during in vitro selection, with MBA5SA1 being the final winner of the competition. Additionally, MBA5SA1 was implemented into an enzyme-linked aptamer binding assay (ELABA), which was applied for detecting Omicron variants in the saliva of infected patients. The assay produced a sensitivity of 86.5 % and a specificity of 100 %, which were established with 83 clinical samples., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Microglia play a crucial role in synaptic elimination by engulfing dystrophic neurons via triggering receptors expressed on myeloid cells 2 (TREM2). They are also involved in the clearance of beta-amyloid (Aβ) plaques in Alzheimer's disease (AD); nonetheless, the driving force behind TREM2-mediated phagocytosis of beta-amyloid (Aβ) plaques remains unknown. Here, using advanced 2D/3D/4D co-culture systems with loss-of-function mutations in TREM2 (a frameshift mutation engineered in exon 2) brain organoids/microglia/assembloids, it is identified that the clearance of Aβ via TREM2 is accelerated by externalized phosphatidylserine (ePtdSer) generated from dystrophic neurons surrounding the Aβ plaques. Moreover, it is investigated whether microglia from both sporadic (CRISPR-Cas9-based APOE4 lines) and familial (APP NL-G-F /MAPT double knock-in mice) AD models show reduced levels of TREM2 and lack of phagocytic activity toward ePtdSer-positive Aβ plaques. Herein new insight is provided into TREM2-dependent microglial phagocytosis of Aβ plaques in the context of the presence of ePtdSer during AD progression., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Continuous monitoring of clinically relevant biomarkers within the interstitial fluid (ISF) using microneedle (MN)-based assays, has the potential to transform healthcare. This study introduces the Wearable Aptalyzer, an integrated system fabricated by combining biocompatible hydrogel MN arrays for ISF extraction with an electrochemical aptamer-based biosensor for in situ monitoring of blood analytes. The use of aptamers enables continuous monitoring of a wide range of analytes, beyond what is possible with enzymatic monitoring. The Wearable Aptalyzer is used for real-time and multiplexed monitoring of glucose and lactate in ISF. Validation experiments using live mice and rat models of type 1 diabetes demonstrate strong correlation between the measurements collected from the Wearable Aptalyzer in ISF and those obtained from gold-standard techniques for blood glucose and lactate, for each analyte alone and in combination. The Wearable Aptalyzer effectively addresses the limitations inherent in enzymatic detection methods as well as solid MN biosensors and the need for reliable and multiplexed bioanalytical monitoring in vivo., (© 2024 The Author(s). Advanced Materials published by Wiley‐VCH GmbH.)

High-precision viral detection at point of need with clinical samples plays a pivotal role in the diagnosis of infectious diseases and the control of a global pandemic. However, the complexity of clinical samples that often contain very low viral concentrations makes it a huge challenge to develop simple diagnostic devices that do not require any sample processing and yet are capable of meeting performance metrics such as very high sensitivity and specificity. Herein we describe a new single-pot and single-step electrochemical method that uses real-time kinetic profiling of the interaction between a high-affinity aptamer and an antigen on a viral surface. This method generates many data points per sample, which when combined with machine learning, can deliver highly accurate test results in a short testing time. We demonstrate this concept using both SARS-CoV-2 and Influenza A viruses as model viruses with specifically engineered high-affinity aptamers. Utilizing this technique to diagnose COVID-19 with 37 real human saliva samples results in a sensitivity and specificity of both 100 % (27 true negatives and 10 true positives, with 0 false negative and 0 false positive), which showcases the superb diagnostic precision of this method., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Systematic evolution of ligands by exponential enrichment (SELEX) has been used to discover thousands of aptamers since its development in 1990. Aptamers are short single-stranded oligonucleotides capable of binding to targets with high specificity and selectivity through structural recognition. While aptamers offer advantages over other molecular recognition elements such as their ease of production, smaller size, extended shelf-life, and lower immunogenicity, they have yet to show significant success in real-world applications. By analyzing the importance of structured library designs, reviewing different SELEX methodologies, and the effects of chemical modifications, we provide a comprehensive overview on the production of aptamers for applications in drug delivery systems, therapeutics, diagnostics, and molecular imaging., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Despite extensive commercial and regulatory interventions, food spoilage and contamination continue to impose massive ramifications on human health and the global economy. Recognizing that such issues will be significantly eliminated by the accurate and timely monitoring of food quality markers, smart food sensors have garnered significant interest as platforms for both real-time, in-package food monitoring and on-site commercial testing. In both cases, the sensitivity, stability, and efficiency of the developed sensors are largely informed by underlying material design, driving focus toward the creation of advanced materials optimized for such applications. Herein, a comprehensive review of emerging intelligent materials and sensors developed in this space is provided, through the lens of three key food quality markers - biogenic amines, pH, and pathogenic microbes. Each sensing platform is presented with targeted consideration toward the contributions of the underlying metallic or polymeric substrate to the sensing mechanism and detection performance. Further, the real-world applicability of presented works is considered with respect to their capabilities, regulatory adherence, and commercial potential. Finally, a situational assessment of the current state of intelligent food monitoring technologies is provided, discussing material-centric strategies to address their existing limitations, regulatory concerns, and commercial considerations., (© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

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., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The early-stage placental barrier is characterized by a lack of fetal circulation and by a thick trophoblastic barrier, whereas the later-stage placenta consists of vascularized chorionic villi encased in a thin, differentiated trophoblast layer, ideal for nutrient transport. In this work, predictive models of early- and late-stage placental transport are created using blastocyst-derived placental stem cells (PSCs) by modulating PSC differentiation and model vascularization. PSC differentiation results in a thinner, fused trophoblast layer, as well as an increase in human chorionic gonadotropin secretion, barrier permeability, and secretion of certain inflammatory cytokines, which are consistent with in vivo findings. Further, gene expression confirms this shift toward a differentiated trophoblast subtype. Vascularization results in a molecule type- and size-dependent change in dextran and insulin permeability. These results demonstrate that trophoblast differentiation and vascularization have critical effects on placental barrier permeability and that this model can be used as a predictive measure to assess fetal toxicity of xenobiotic substances at different stages of pregnancy., (© 2023 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.)

Engineering functional nucleic acids that are active under unusual conditions will not only reveal their hidden abilities but also lay the groundwork for pursuing them for unique applications. Although many DNAzymes have been derived to catalyze diverse chemical reactions in aqueous solutions, no prior study has been set up to purposely derive DNAzymes that require an organic solvent to function. Herein, we utilized in vitro selection to isolate RNA-cleaving DNAzymes from a random-sequence DNA pool that were "compelled" to accept 35 % dimethyl sulfoxide (DMSO) as a cosolvent, via counter selection in a purely aqueous solution followed by positive selection in the same solution containing 35 % DMSO. This experiment led to the discovery of a new DNAzyme that requires 35 % DMSO for its catalytic activity and exhibits drastically reduced activity without DMSO. This DNAzyme also requires divalent metal ions for catalysis, and its activity is enhanced by monovalent ions. A minimized, more efficient DNAzyme was also derived. This work demonstrates that highly functional, organic solvent-dependent DNAzymes can be isolated from random-sequence DNA libraries via forced in vitro selection, thus expanding the capability and potential utility of catalytic DNA., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

With food production shifting away from traditional farm-to-table approaches to efficient multistep supply chains, the incidence of food contamination has increased. Consequently, pathogen testing via inefficient culture-based methods has increased, despite its lack of real-time capabilities and need for centralized facilities. While in situ pathogen detection would address these limitations and enable individual product monitoring, accurate detection within unprocessed, packaged food products without user manipulation has proven elusive. Herein, "Lab-in-a-Package" is presented, a platform capable of sampling, concentrating, and detecting target pathogens within closed food packaging, without intervention. This system consists of a newly designed packaging tray and reagent-infused membrane that can be paired universally with diverse pathogen sensors. The inclined food packaging tray maximizes fluid localization onto the sensing interface, while the membrane acts as a reagent-immobilizing matrix and an antifouling barrier for the sensor. The platform is substantiated using a newly discovered Salmonella-responsive nucleic acid probe, which enables hands-free detection of 10 3 colony forming units (CFU) g -1 target pathogen in a packaged whole chicken. The platform remains effective when contamination is introduced with toolsand surfaces, ensuring widespread efficacy. Its real-world use for in situ detection is simulated using a handheld fluorescence scanner with smartphone connectivity., (© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems are increasingly used in biosensor development. However, directly translating recognition events for non-nucleic acid targets by CRISPR into effective measurable signals represents an important ongoing challenge. Herein, it is hypothesized and confirmed that CRISPR RNAs (crRNAs) in a circular topology efficiently render Cas12a incapable of both site-specific double-stranded DNA cutting and nonspecific single-stranded DNA trans cleavage. Importantly, it is shown that nucleic acid enzymes (NAzymes) with RNA-cleaving activity can linearize the circular crRNAs, activating CRISPR-Cas12a functions. Using ligand-responsive ribozymes and DNAzymes as molecular recognition elements, it is demonstrated that target-triggered linearization of circular crRNAs offers great versatility for biosensing. This strategy is termed as "NAzyme-Activated CRISPR-Cas12a with Circular CRISPR RNA (NA3C)." Use of NA3C for clinical evaluation of urinary tract infections using an Escherichia coli-responsive RNA-cleaving DNAzyme to test 40 patient urine samples, providing a diagnostic sensitivity of 100% and specificity of 90%, is further demonstrated., (© 2023 Wiley-VCH GmbH.)

Alzheimer's disease (AD) is one of the main causes of dementia worldwide, whereby neuronal death or malfunction leads to cognitive impairment in the elderly population. AD is highly prevalent, with increased projections over the next few decades. Yet current diagnostic methods for AD occur only after the presentation of clinical symptoms. Evidence in the literature points to potential mechanisms of AD induction beginning before clinical symptoms start to present, such as the formation of amyloid beta (A β ) extracellular plaques and neurofibrillary tangles (NFTs). Biomarkers of AD, including A β 40 , and tau protein, amongst others, show promise for early AD diagnosis. Additional progress is made in the application of biosensing modalities to measure and detect significant changes in these AD biomarkers within patient samples, such as cerebral spinal fluid (CSF) and blood, serum, or plasma. Herein, a comprehensive review of the emerging nano-biomaterial approaches to develop biosensors for AD biomarkers' detection is provided. Advances, challenges, and potential of electrochemical, optical, and colorimetric biosensors, focusing on nanoparticle-based (metallic, magnetic, quantum dots) and nanostructure-based biomaterials are discussed. Finally, the criteria for incorporating these emerging nano-biomaterials in clinical settings are presented and assessed, as they hold great potential for enhancing early-onset AD diagnostics.β 42 , and tau protein, amongst others, show promise for early AD diagnosis. Additional progress is made in the application of biosensing modalities to measure and detect significant changes in these AD biomarkers within patient samples, such as cerebral spinal fluid (CSF) and blood, serum, or plasma. Herein, a comprehensive review of the emerging nano-biomaterial approaches to develop biosensors for AD biomarkers' detection is provided. Advances, challenges, and potential of electrochemical, optical, and colorimetric biosensors, focusing on nanoparticle-based (metallic, magnetic, quantum dots) and nanostructure-based biomaterials are discussed. Finally, the criteria for incorporating these emerging nano-biomaterials in clinical settings are presented and assessed, as they hold great potential for enhancing early-onset AD diagnostics., Competing Interests: Conflict of Interest The authors declare no conflict of interest.

Fusobacterium nucleatum has been correlated to many poor human conditions including oral infections, adverse pregnancies and cancer, and thus molecular tools capable of detecting this human pathogen can be used to develop diagnostic tests for them. Using a new selection method targeting thermally stable proteins without a counter-selection step, we derived an fluorogenic RNA-cleaving DNAzyme, named RFD-FN1, that can be activated by a thermally stable protein target that is unique to F. nucleatum subspecies. High thermal stability of protein targets is a very desirable attribute for DNAzyme-based biosensing directly with biological samples because nucleases found inherently in these samples can be heat-inactivated. We further demonstrate that RFD-FN1 can function as a fluorescent sensor in both human saliva and human stool samples. The discovery of RFD-FN1 paired with a highly thermal stable protein target presents opportunities for developing simpler diagnostic tests for this important pathogen., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

There is a need for point-of-care bacterial sensing and identification technologies that are rapid and simple to operate. Technologies that do not rely on growth cultures, nucleic acid amplification, step-wise reagent addition, and complex sample processing are the key for meeting this need. Herein, multiple materials technologies are integrated for overcoming the obstacles in creating rapid and one-pot bacterial sensing platforms. Liquid-infused nanoelectrodes are developed for reducing nonspecific binding on the transducer surface; bacterium-specific RNA-cleaving DNAzymes are used for bacterial identification; and redox DNA barcodes embedded into DNAzymes are used for binding-induced electrochemical signal transduction. The resultant single-step and one-pot assay demonstrates a limit-of-detection of 10 2 CFU mL -1 , with high specificity in identifying Escherichia coli amongst other Gram positive and negative bacteria including Klebsiella pneumoniae, Staphylococcus aureus, and Bacillus subtilis. Additionally, this assay is evaluated for analyzing 31 clinically obtained urine samples, demonstrating a clinical sensitivity of 100% and specify of 100%. When challenging this assay with nine clinical blood cultures, E. coli-positive and E. coli-negative samples can be distinguished with a probability of p < 0.001., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Clostridium difficile frequently causes an infectious disease known as Clostridium difficile infection (CDI), and there is an urgent need for the development of more effective rapid diagnostic tests for CDI. Previously we have developed an RNA-cleaving fluorogenic DNAzyme (RFD) probe, named RFD-CD1, that is capable of detecting a specific strain of C. difficile but is too specific to recognize other pathogenic C. difficile strains. To overcome this issue, herein we report RFD-CD2, another RFD that is not only highly specific to C. difficile but also capable of recognizing diverse pathogenic C. difficile strains. Extensive sequence and structure characterization establishes a pseudoknot structure and a significantly minimized sequence for RFD-CD2. As a fluorescent sensor, RFD-CD2 can detect C. difficile at a concentration as low as 100 CFU/mL, thus making this DNAzyme an attractive molecular probe for rapid diagnosis of CDI caused by diverse strains of C. difficile., (© 2023 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

A new method for the detection of genomic RNA combines RNA cleavage by the 10-23 DNAzyme and use of the cleavage fragments as primers to initiate rolling circle amplification (RCA). 230 different 10-23 DNAzyme variants were screened to identify those that target accessible RNA sites within the highly structured RNA transcripts of SARS-CoV-2. A total of 28 DNAzymes were identified with >20 % cleavage, 5 with >40 % cleavage and one with >60 % in 10 min. The cleavage fragments from these reactions were then screened for coupling to an RCA reaction, leading to the identification of several cleavage fragments that could efficiently initiate RCA. Using a newly developed quasi-exponential RCA method with a detection limit of 500 aM of RNA, 14 RT-PCR positive and 15 RT-PCR negative patient saliva samples were evaluated for SARS-CoV-2 genomic RNA, achieving a clinical sensitivity of 86 % and specificity of 100 % for detection of the virus in <2.5 h., (© 2023 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

An Au-on-Au tip sensor is developed for the detection of Salmonella typhimurium (Salmonella), using a new synthetic nucleic acid probe (NAP) as a linker for the immobilization of a DNA-conjugated Au nanoparticle (AuNP) onto a DNA-attached thin Au layer inside a pipette tip. In the presence of Salmonella, RNase H2 from Salmonella (STH2) cleaves the NAP and the freed DNA-conjugated AuNP can be visually detected by a paper strip. This portable biosensor does not require any electronic, electrochemical or optical equipment. It delivers a detection limit of 3.2×10 3  CFU mL -1 for Salmonella in 1 h without cell-culturing or signal amplification and does not show cross-reactivity with several control bacteria. Further, the sensor reliably detects Salmonella spiked in food samples, such as ground beef and chicken, milk, and eggs. The sensor can be reused and is stable at ambient temperature, showing its potential as a point-of-need device for the prevention of food poisoning by Salmonella., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Engineered surfaces that repel pathogens are of great interest due to their role in mitigating the spread of infectious diseases. A robust, universal, and scalable omniphobic spray coating with excellent repellency against water, oil, and pathogens is presented. The coating is substrate-independent and relies on hierarchically structured polydimethylsiloxane (PDMS) microparticles, decorated with gold nanoparticles (AuNPs). Wettability studies reveal the relationship between surface texturing of micro- and/or nano-hierarchical structures and the omniphobicity of the coating. Studies of pathogen transfer with bacteria and viruses reveal that an uncoated contaminated glove transfers pathogens to >50 subsequent surfaces, while a coated glove picks up 10 4 (over 99.99%) less pathogens upon first contact and transfers zero pathogens after the second touch. The developed coating also provides excellent stability under harsh conditions. The remarkable anti-pathogen properties of this surface combined with its ease of implementation, substantiate its use for the prevention of surface-mediated transmission of pathogens., (© 2023 Wiley-VCH GmbH.)

Nucleic acids are remarkable molecules. In addition to Watson-Crick base pairing, the different structural motifs of these molecules can bind non-nucleic acid targets or catalyze chemical reactions. Additionally, nucleic acids are easily modified with different molecules or functional groups. These properties make nucleic acids, particularly DNA, ideally suited for use in electrochemical biosensors, both as biorecognition elements and redox reporter probes. In this Minireview, we will review the historical evolution of nucleic acids as probes in electrochemical biosensors. We will then review the specific examples of nucleic-acid-based biosensors that have been evaluated for clinical use in the areas of infectious disease, cancer, or cardiovascular health., (© 2022 Wiley-VCH GmbH.)

The SARS-CoV-2 virus and COVID-19 pandemic continue to demand effective diagnostic and therapeutic solutions. Finding these solutions requires highly functional molecular recognition elements. Nucleic acid aptamers represent a possible solution. Characterized by their high affinity and specificity, aptamers can be rapidly identified from random-sequence nucleic acid libraries. Over the past two years, many labs around the world have rushed to create diverse aptamers that target two important structural proteins of SARS-CoV-2: the spike (S) protein and nucleocapsid (N) protein. These have led to the identification of many aptamers that show real promise for the development of diagnostic tests and therapeutic agents for SARS-CoV-2. Herein we review all these developments, with a special focus on the development of diverse aptasensors for detecting SARS-CoV-2. These include electrochemical and optical sensors, lateral flow devices, and aptamer-linked immobilized sorbent assays., Competing Interests: The authors declare no conflict of interest., (© 2022 Wiley‐VCH GmbH.)

The cover feature image shows nucleic acid aptamers armed and ready for our battle against the monstrous SARS-CoV-2 virus. Often thought of as "chemical antibodies", these molecular recognition elements are equipped with several unique benefits and have thus been a popular research subject worldwide. Many aptamers for recognizing the spike and nucleocapsid proteins of SARS-CoV-2 have been developed and examined as diagnostic and therapeutic weaponry for the war against COVID-19 and future pandemics. More information can be found in the Review by J. D. Brennan, Y. Li, and co-workers., (© 2022 Wiley‐VCH GmbH.)

Pen-side testing of farm animals for infectious diseases is critical for preventing transmission in herds and providing timely intervention. However, most existing pathogen tests have to be conducted in centralized labs with sample-to-result times of 2-4 days. Herein we introduce a test that uses a dual-electrode electrochemical chip (DEE-Chip) and a barcode-releasing electroactive aptamer for rapid on-farm detection of porcine epidemic diarrhea viruses (PEDv). The sensor exploits inter-electrode spacing reduction and active field mediated transport to accelerate barcode movement from electroactive aptamers to the detection electrode, thus expediting assay operation. The test yielded a clinically relevant limit-of-detection of 6 nM (0.37 μg mL -1 ) in saliva-spiked PEDv samples. Clinical evaluation of this biosensor with 12 porcine saliva samples demonstrated a diagnostic sensitivity of 83 % and specificity of 100 % with a concordance value of 92 % at an analysis time of one hour., (© 2022 Wiley-VCH GmbH.)

Recent multi-omics analyses paved the way for a comprehensive understanding of pathological processes. However, only few studies have explored Alzheimer's disease (AD) despite the possibility of biological subtypes within these patients. For this study, unsupervised classification of four datasets (genetics, miRNA transcriptomics, proteomics, and blood-based biomarkers) using Multi-Omics Factor Analysis+ (MOFA+), along with systems-biological approaches following various downstream analyses are performed. New subgroups within 170 patients with cerebral amyloid pathology (Aβ+) are revealed and the features of them are identified based on the top-rated targets constructing multi-omics factors of both whole (M-TPAD) and immune-focused models (M-IPAD). The authors explored the characteristics of subtypes and possible key-drivers for AD pathogenesis. Further in-depth studies showed that these subtypes are associated with longitudinal brain changes and autophagy pathways are main contributors. The significance of autophagy or clustering tendency is validated in peripheral blood mononuclear cells (PBMCs; n = 120 including 30 Aβ- and 90 Aβ+), induced pluripotent stem cell-derived human brain organoids/microglia (n = 12 including 5 Aβ-, 5 Aβ+, and CRISPR-Cas9 apolipoprotein isogenic lines), and human brain transcriptome (n = 78). Collectively, this study provides a strategy for precision medicine therapy and drug development for AD using integrative multi-omics analysis and network modelling., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Aptamers that can recognize the spike (S) protein of SARS-CoV-2 with high affinity and specificity are useful molecules towards the development of diagnostics and therapeutics to fight COVID-19. However, this S protein is constantly mutating, producing variants of concern (VoCs) that can significantly weaken the binding by aptamers initially engineered to recognize the S protein of the wildtype virus or a specific VoC. One strategy to overcome this problem is to develop universal aptamers that are insensitive to all or most of the naturally emerging mutations in the protein. We have recently demonstrated this concept by subjecting a pool of S protein-binding DNA aptamers for one-round parallel-SELEX experiments targeting 5 different S protein variants for binding-based sequence enrichment, followed by bioinformatic analysis of the enriched pools. This effort has led to the identification of a universal aptamer that recognizes 8 different variants of the spike protein with equally excellent affinity., (© 2022 Wiley-VCH GmbH.)

The surface fouling of biomedical devices has been an ongoing issue in healthcare. Bacterial and blood adhesion in particular, severely impede the performance of such tools, leading to poor patient outcomes. Various structural and chemical modifications have been shown to reduce fouling, but all existing strategies lack the combination of physical, chemical, and economic traits necessary for widespread use. Herein, a lubricant infused, hierarchically micro- and nanostructured polydimethylsiloxane surface is presented. The surface is easy to produce and exhibits the high flexibility and optical transparency necessary for incorporation into various biomedical tools. Tests involving two clinically relevant, priority pathogens show up to a 98.5% reduction in the biofilm formation of methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. With blood, the surface reduces staining by 95% and suppresses thrombin generation to background levels. Furthermore, the surface shows applicability within applications such as catheters, extracorporeal circuits, and microfluidic devices, through its effectiveness in dynamic conditions. The perfusion of bacterial media shows up to 96.5% reduction in bacterial adhesion. Similarly, a 95.8% reduction in fibrin networks is observed following whole blood perfusion. This substrate stands to hold high applicability within biomedical systems as a means to prevent fouling, thus improving performance., (© 2022 Wiley-VCH GmbH.)

Invited for the cover of this issue are John Brennan, Yingfu Li, and co-workers at McMaster University. The image depicts MSA52 as a universal DNA aptamer that recognizes spike proteins of diverse SARS-CoV-2 variants of concern. Read the full text of the article at 10.1002/chem.202200078., (© 2022 Wiley-VCH GmbH.)

We report on a unique DNA aptamer, denoted MSA52, that displays universally high affinity for the spike proteins of wildtype SARS-CoV-2 as well as the Alpha, Beta, Gamma, Epsilon, Kappa, Delta and Omicron variants. Using an aptamer pool produced from round 13 of selection against the S1 domain of the wildtype spike protein, we carried out one-round SELEX experiments using five different trimeric spike proteins from variants, followed by high-throughput sequencing and sequence alignment analysis of aptamers that formed complexes with all proteins. A previously unidentified aptamer, MSA52, showed K d values ranging from 2 to 10 nM for all variant spike proteins, and also bound similarly to variants not present in the reselection experiments. This aptamer also recognized pseudotyped lentiviruses (PL) expressing eight different spike proteins of SARS-CoV-2 with K d values between 20 and 50 pM, and was integrated into a simple colorimetric assay for detection of multiple PL variants. This discovery provides evidence that aptamers can be generated with high affinity to multiple variants of a single protein, including emerging variants, making it well-suited for molecular recognition of rapidly evolving targets such as those found in SARS-CoV-2., (© 2022 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

Detection of pathogenic bacteria in complex biological matrices remains a major challenge. Herein, we report the selection and optimization of a new DNAzyme for Staphylococcus aureus (SA) and the use of the DNAzyme to develop a simple lateral flow device (LFD) for detection of SA in nasal mucus. The DNAzyme was generated by in vitro selection using a crude extra/intracellular mixture derived from SA, which could be used directly for simple solution or paper-based fluorescence assays for SA. The DNAzyme was further modified to produce a DNA cleavage fragment that acted as a bridging element to bind DNA-modified gold nanoparticles to the test line of a LFD, producing a simple colorimetric dipstick test. The LFD was evaluated with nasal mucus samples spiked with SA, and demonstrated that SA detection was possible in minutes with minimal sample processing., (© 2021 Wiley-VCH GmbH.)

There is a constant drive for affordable point-of-care testing (POCT) technologies for the detection of infectious human diseases. Herein, we report a simple platform for DNA detection that takes advantage of four techniques: commercially available pregnancy test strips (PTS), amplicon generation via loop-mediated isothermal amplification (LAMP), toehold-mediated strand displacement, and noncovalent immobilization of DNA on paper surface with DNA nanoflowers. This simple, separation-free platform is highly specific, as demonstrated with the detection of rtL180M, a single-nucleotide polymorphism observed in hepatitis B virus (HBV) associated with antiviral drug resistance. It is very sensitive, capable of detecting the targeted mutation at 2 copies μL -1 . It is able to correctly identify the unmutated and rtL180M genome types of HBV in clinical samples. Given its wide adaptability, we expect this platform can be easily modified for the detection of genetic variations associated with various pathogens and human diseases., (© 2021 Wiley-VCH GmbH.)

We report a simple and rapid saliva-based SARS-CoV-2 antigen test that utilizes a newly developed dimeric DNA aptamer, denoted as DSA1N5, that specifically recognizes the spike proteins of the wildtype virus and its Alpha and Delta variants with dissociation constants of 120, 290 and 480 pM, respectively, and binds pseudotyped lentiviruses expressing the wildtype and alpha trimeric spike proteins with affinity constants of 2.1 pM and 2.3 pM, respectively. To develop a highly sensitive test, DSA1N5 was immobilized onto gold electrodes to produce an electrochemical impedance sensor, which was capable of detecting 1000 viral particles per mL in 1:1 diluted saliva in under 10 min without any further sample processing. Evaluation of 36 positive and 37 negative patient saliva samples produced a clinical sensitivity of 80.5 % and specificity of 100 % and the sensor could detect the wildtype virus as well as the Alpha and Delta variants in the patient samples, which is the first reported rapid test that can detect any emerging variant of SARS-CoV-2., (© 2021 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

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., (© 2021 Wiley-VCH GmbH.)

Legionella pneumophila is a deadly bacterial pathogen that has caused numerous Legionnaires' disease outbreaks, where cooling towers were the most common source of exposure. Bacterial culturing is used for L. pneumophila detection, but this method takes approximately 10 days to complete. In this work, an RNA-cleaving fluorogenic DNAzyme, named LP1, was isolated. Extensive characterization revealed that LP1 is reactive with multiple infectious isolates of L. pneumophila but inactive with 25 other common bacterial species. LP1 is likely activated by a protein target, capable of generating a detectable signal in the presence of as few as 10 colony-forming units of L. pneumophila, and able to maintain its activity in cooling tower water from diverse sources. Given that similar DNAzymes have been incorporated into many sensitive assays for bacterial detection, LP1 holds the potential for the development of biosensors for monitoring the contamination of L. pneumophila in exposure sources., (© 2020 Wiley-VCH GmbH.)

φ29 DNA polymerase (Polφ29) is capable of synthesizing long-chain single-stranded (ss) DNA molecules by copying the sequence of a small ss circular DNA template (ssCDT) in a process known as rolling circle amplification (RCA). The use of a ssCDT in RCA, however, comes with a key drawback: the rate of DNA synthesis is significantly reduced. We hypothesize that this issue can be overcome using a very long linear ssDNA template with a repeating sequence. To test this idea, we engineered a DNA assembly, which we denote "micrometer-sized DNA track" (μDT). This μDT, with an average length of ≈13.5 μm, is made of a long chain DNA with a primer-binding domain at its 3' end and ≈1000 repeating sequence units at its 5' end, each carrying a DNA anchor. We find that Polφ29 copies μDT at a speed ≈5-time faster than it does a related ssCDT. We use this to design a simple all-in-one printed paper device for rapid and sensitive detection of microRNA let-7. This paper sensor is capable of detecting 1 pM let-7a in 10 minutes., (© 2020 Wiley-VCH GmbH.)

Scope: Alzheimer's disease is a neurodegenerative condition marked by the formation and aggregation of amyloid-β (Aβ) peptides. There exists, to this day, no cure or effective prevention for the disease; however, there is evidence that a healthy diet and certain food products can slow down first occurrence and progression. To investigate if food ingredients can interact with peptide aggregates, synthetic membranes that contained aggregates consisting of cross-β sheets of the membrane active fragment A β 25 -- 35 are prepared., Methods and Results: The impact of resveratrol, found in grapes, caffeine, the main active ingredient in coffee, β-carotene, found in orange fruits and vegetables, and epigallocatechin gallate (EGCG), a component of green tea, on the size and volume fraction of Aβ aggregates is studied using optical and fluorescence microscopy, X-ray diffraction, UV-vis spectroscopy, and molecular dynamics simulations. All compounds are membrane active and spontaneously partitioned in the synthetic brain membranes. While resveratrol and caffeine lead to membrane thickening and reduced membrane fluidity, β-carotene and EGCG preserve or increase fluidity., Conclusion: Resveratrol and caffeine do not reduce the volume fraction of peptide aggregates while β-carotene significantly reduces plaque size. Interestingly, EGCG dissolves peptide aggregates and significantly decreases the corresponding cross-β and β-sheet signals., (© 2020 Wiley-VCH GmbH.)

Herein, we report on the design of a programmable DNA ribbon using long-chain DNA molecules with a user-defined repetitive padlock sequence. The DNA ribbon can be further combined with gold nanoparticles (AuNPs) to create a composite nanomaterial that contains an AuNP core and a high-density DNA crown carrying a cancer-cell-targeting DNA aptamer, a fluorescent tag for location tracking, and a cell-killing drug. This composite material can be efficiently internalized by cancer cells and its cellular location can be tracked by fluorescence imaging. The system offers several attractive characteristics, including simple design, tunable DNA crown, high drug-loading capacity, selective cell targeting, and pH-sensitive drug release. These features make such a material a promising therapeutic agent., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Purpose: Antimicrobial resistance (AMR), especially multidrug resistance, is one of the most serious global threats facing public health. The authors proof-of-concept study assessing the suitability of shotgun proteomics as an additional approach to whole-genome sequencing (WGS) for detecting AMR determinants., Experimental Design: Previously published shotgun proteomics and WGS data on four isolates of Campylobacter jejuni are used to perform AMR detection by searching the Comprehensive Antibiotic Resistance Database, and their detection ability relative to genomics screening and traditional phenotypic testing measured by minimum inhibitory concentration is assessed., Results: Both genomic and proteomic approaches identify the wild-type and variant molecular determinants responsible for resistance to tetracycline and ciprofloxacin, in agreement with phenotypic testing. In contrast, the genomic method identifies the presence of the β-lactamase gene, bla OXA - 61 , in three isolates. However, its corresponding protein product is detected in only a single isolate, consistent with results obtained from phenotypic testing., (© 2019 Her Majesty the Queen in Right of Canada. Proteomics - Clinical Applications published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We report on a programmable all-DNA biosensing system that centers on the use of a 4-way junction (4WJ) to transduce a DNAzyme reaction into an amplified signal output. A target acts as a primary input to activate an RNA-cleaving DNAzyme, which then cleaves an RNA-containing DNA substrate that is designed to be a component of a 4WJ. The formation of the 4WJ controls the release of a DNA output that becomes an input to initiate catalytic hairpin assembly (CHA), which produces a second DNA output that controls assembly of a split G-quadruplex as a fluorescence signal generator. The 4WJ can be configured to produce either a turn-off or turn-on switch to control the degree of CHA, allowing target concentration to be determined in a quantitative manner. We demonstrate this approach by creating a sensor for E. coli that could detect as low as 50 E. coli cells mL -1 within 85 min and offers an amplified bacterial detection method that does not require a protein enzyme., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

DNA detection is usually conducted under nondenaturing conditions to favor the formation of Watson-Crick base-paring interactions. However, although such a setting is excellent for distinguishing a single-nucleotide polymorphism (SNP) within short DNA sequences (15-25 nucleotides), it does not offer a good solution to SNP detection within much longer sequences. Here we report on a new detection method capable of detecting SNP in a DNA sequence containing 35-90 nucleotides. This is achieved through incorporating into the recognition DNA sequence a previously discovered DNA molecule that forms a stable G-quadruplex in the presence of 7 molar urea, a known condition for denaturing DNA structures. The systems are configured to produce both colorimetric and fluorescent signals upon target binding., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Invited for the cover of this issue is the group of Yingfu Li at McMaster University. The image depicts a molecular switch for ultra-specific detection of DNA utilizing a guanine-quadruplex (up-right structure) resistant to denaturation by urea (ball-and-stick structures). Read the full text of the article at 10.1002/chem.201903536., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

New surface coatings that enhance hemocompatibility and biofunctionality of synthetic vascular grafts such as expanded poly(tetrafluoroethylene) (ePTFE) and poly(ethylene terephthalate) (PET) are urgently needed. Lubricant-infused surfaces prevent nontargeted adhesion and enhance the biocompatibility of blood-contacting surfaces. However, limited success has been made in incorporating biofunctionality onto these surfaces and generating biofunctional lubricant-infused coatings that both prevent nonspecific adhesion and enhance targeted binding of biomolecules remains a challenge. Here, a new generation of fluorosilanized lubricant-infused PET surfaces with built-in biofunctional nanoprobes is reported. These surfaces are synthesized by starting with a self-assembled monolayer of fluorosilane that is partially etched using plasma modification technique, thereby creating a hydroxyl-terminated fluorosilanized PET surface. Simultaneously, silanized nanoprobes are produced by amino-silanizing anti-CD34 antibody in solution and directly coupling the anti-CD34-aminosilane nanoprobes onto the hydroxyl terminated, fluorosilanized PET surface. The PET surfaces are then lubricated, creating fluorosilanized biofunctional lubricant-infused PET substrates. Compared with unmodified PET surfaces, the designed biofunctional lubricant-infused PET surfaces significantly attenuate thrombin generation and blood clot formation and promote targeted binding of endothelial cells from human whole blood., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Phosphatidylinositol (PI) lipids have a predominance of a single molecular species present through the organism. In healthy mammals this molecular species is 1-stearoyl-2-arachidonoyl (18:0/20:4) PI. Although the importance of PI lipids for cell physiology has long been appreciated, less is known about the biological role of enriching PI lipids with 18:0/20:4 acyl chains. In conditions with dysfunctional lipid metabolism, the predominance of 18:0/20:4 acyl chains is lost. Recently, molecular mechanisms underpinning the enrichment or alteration of these acyl chains in PI lipids have begun to emerge. In the majority of the cases a common feature is the presence of enzymes bearing substrate acyl chain specificity. However, in cancer cells, it has been shown that one (not the only) of the mechanisms responsible for the loss in this acyl chain enrichment is mutation on the transcription factor p53 gene, which is one of the most highly mutated genes in cancers. There is a compelling need for a global picture of the specificity of the acyl chain composition of PIs. This can be possible once high-resolution spatio-temporal information is gathered in a cellular context; which can ultimately lead to potential novel targets to combat conditions with altered PI acyl chain profiles., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The reliable detection of pathogenic bacteria in complex biological samples using simple assays or devices remains a major challenge. Herein, we report a simple colorimetric paper device capable of providing specific and sensitive detection of Helicobacter pylori (H. pylori), a pathogen strongly linked to gastric carcinoma, gastric ulcers, and duodenal ulcers, in stool samples. The sensor molecule, an RNA-cleaving DNAzyme obtained through in vitro selection, is activated by a protein biomarker from H. pylori. The colorimetric paper sensor, designed on the basis of the RNA-cleaving property of the DNAzyme, is capable of sensitive detection of H. pylori in human stool samples with minimal sample processing and provides results in minutes. It remains fully functional under storage at ambient temperature for at least 130 days. This work lays a foundation for developing DNAzyme-enabled paper-based point-of-care diagnostic devices for monitoring pathogens in complex samples., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We report on the first effort to select DNA aptamers from a circular DNA library, which resulted in the discovery of two high-affinity circular DNA aptamers that recognize the glutamate dehydrogenase (GDH) from Clostridium difficile, an established antigen for diagnosing Clostridium difficile infection (CDI). One aptamer binds effectively in both the circular and linear forms, the other is functional only in the circular configuration. Interestingly, these two aptamers recognize different epitopes on GDH, demonstrating the advantage of selecting aptamers from circular DNA libraries. A sensitive diagnostic test was developed to take advantage of the high stability of circular DNA aptamers in biological samples and their compatibility with rolling circle amplification. This test is capable of identifying patients with active CDI using stool samples. This work represents a significant step forward towards demonstrating the practical utility of DNA aptamers in clinical diagnosis., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The aggregation of amyloid beta (Aβ) proteins in senile plaques is a critical event during the development of Alzheimer's disease, and the postmortem detection of Aβ-rich proteinaceous deposits through fluorescent staining remains one of the most robust diagnostic tools. In animal models, fluorescence imaging can be employed to follow the progression of the disease, and among the different imaging methods, two-photon microscopy (TPM) has emerged as one of the most powerful. To date, several near-infrared-emissive two-photon dyes with a high affinity for Aβ fibrils have been developed, but there has often been a tradeoff between excellent two-photon cross-sections and large fluorescence signal-to-background ratios. In the current work, we introduced a twisted intramolecular charge state (TICT)-based de-excitation pathway, which results in a remarkable fluorescence increase of around 167-fold in the presence of Aβ fibrils, while maintaining an excellent two-photon cross section, thereby enabling high-contrast ex vivo and in vivo TPM imaging. Overall, the results suggest that adopting TICT de-excitation in two-photon fluorophores may represent a general method to overcome the tradeoff between probe brightness and signal-to-background ratio., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)