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The synthesis of Co2+/CS/Lum/AuNFs and the label-free sensing of HCVcp is reported.

Herein, a label-free fluorescent signal amplification system based on a catalyzed hairpin assembly (CHA) is reported. In this system, two hairpin probes, H1 and H2, were well-designed in which G-quadruplex sequences were integrated into H2. The CHA reaction was triggered by target/trigger DNA and G-quadruplex sequences were released, which can bind the fluorescent amyloid dye thioflavin T (ThT) to provide fluorescence signals. At the same time, target/trigger DNA was released from the product of the CHA reaction (H1–H2), which continued to initiate the next CHA cycle, and the signal was eventually amplified. This signal amplification approach has been successfully used to develop a label-free fluorescent sensing platform for sensitive detection of human immunodeficiency virus (HIV) DNA and Pb2+.

Circulating tumor DNA (ctDNA), carrying tumor-specific sequence mutations, is a promising biomarker for classification, diagnosis and prognosis of cancers. However, there is still a great challenge in discriminating single-base difference between ctDNA and its coexisting analogue (normal circulating DNA, ncDNA) at a serum sample. A locked nucleic acid (LNA) probe combined with α-HL nanopore sensor was designed, which achieved a high signal-to-background ratio (SBR) of ∼8.34 × 103, as well as a significant discrimination capability (∼12.3 times) of single-base difference. The accurate discrimination strategy is label-free, convenient, selective and sensitive, which has great potential in the early diagnosis of diseases and biomedical research fields.

A rapid and colorimetric biosensor for Pb2+ detection has been constructed on the basis of Pb2+-dependent GR-5 DNAzyme and the self-replicating catalyzed hairpin assembly (SRCHA) reaction. In the presence of Pb2+, the phosphodiester bond of the internal RNA base (rA) in GR-5 DNAzyme was cleaved by the enzyme strand and trigger DNA was released. Then the SRCHA reaction was initiated and the H1–H2 complex equipped with single-stranded DNA (ssDNA) sticky ends on both sides was formed. The sequences of the newly formed sticky ends on one side of H1–H2 were the same as that of trigger DNA and then additional CHA reaction cycles were initiated. Meanwhile, the intact G-quadruplex was reunited on the other side of H1–H2 for colorimetric signal readout. By taking advantage of the self-replication of trigger DNA, the response signal in this sensing system was significantly amplified and Pb2+ can be detected in a linear range from 20 to 100 nM with a detection limit of 2.6 nM within 20 min. In addition, this method can be applied to the reliable monitoring of spiked Pb2+ in environmental water and serum samples with satisfying results, providing a potential method for on-site and real-time detection of Pb2+ for environmental protection and related disease prevention.

The simultaneous discrimination of let-7 microRNAs (miRNAs) would greatly facilitate the early diagnosis and prognosis monitoring of diseases. In this work, a molecular beacon DNA probe was designed to be able to flip out its mismatched cytosine base within the α-hemolysin (α-HL) latch and generate completely separated blocking currents to identify the single-base difference. As a result, the characteristic blocking current of fully matched MB/let-7a and single-base mismatched MB/let-7f was 84.30 ± 0.92 and 87.05 ± 0.86% (confidence level P 95%), respectively. Let-7 miRNA family let-7a and let-7f were completely simultaneously discriminated, which could be attributed to the following strengths. (1) The statistic distribution of blocking current is extremely concentrated with a small relative standard deviation (RSD) of less than 1% and a narrow distribution range. (2) Complete separation is achieved with a high separation resolution of 1.54. (3) The cytosine base flipping out within the α-HL latch provides a universal labeling-free strategy to simultaneously discriminate the single-base mismatch. Overall, the target let-7f sequences were detected with a linear range from 0.001 to 10 pM in human serum samples containing 200 nM let-7a. Great potential has been demonstrated for precise detection, early diagnosis, and prognosis monitoring of diseases related to single-base difference.

Current tools for dNTP analysis mainly rely on expensive fluorescent labeling, mass spectrometry or electrochemistry. Single-molecule assay by protein nanopores with an internal diameter of ca. 1-3.6 nm provides a useful tool for dNTP sensing. However, the most commonly used protein nanopores require additional modifications to enable dNTP detection. In this study, the PaMscS channel (mechanosensitive channel of small conductance from Pseudomonas aeruginosa) embedded in the bilayer lipid membrane (BLM) of E. coli polar lipid extract was applied as a nanopore for single molecular sensing. Two mutants of PaMscS nanopores on the side portal region (PaMscS W130A and PaMscS K180R) were selected for direct dNTP or pyrophosphoric acid (PPi) detection without aptamer or protein modification. Notably, the PaMscS mutant pore can be adjusted by regulation of osmolarity differences, which is crucial for the optimal detection of specific molecules. In addition, we established a PaMscS-based diagnosis method for the rapid sensing of disease-associated nucleic acids by monitoring the consumption of dNTPs, with 86% specificity and 100% sensitivity among 22 clinical samples. This protein nanopore, without aptamer or modification, paves a new way for dNTPs, PPi direct sensing and nucleic acid detection with low cost but high versatility.

A rapid room-temperature DNA nanowires preparation strategy on the basis of self-replicating catalyzed hairpin assembly (SRCHA) was reported. In this system, three hairpin probes (P1, P2, and P3) were well-designed and partially hybridize to each other, and two split trigger DNA sequences were integrated into P1 and P3, respectively. When the SRCHA was initiated by the trigger DNA, a series of DNA assembly steps based on the toehold-mediated DNA strand displacement were activated, and the Y shaped DNA (P1-P2-P3) was formed. In that case, the two split trigger DNA sequences will come into close-enough proximity to form the trigger DNA replicas, which can initiate the additional SRCHA reaction cycles for DNA nanowire preparation, and eventually a rapid room-temperature DNA nanowires preparation strategy without need of fuel strands was successfully developed. Furthermore, the prepared DNA nanowires have been used to develop a rapid and signal amplified sensing platform for sensitive adenosine triphosphate (ATP) detection.

Convenient and time-saving one-step strategies for detecting ultralow concentrations of protein biomarkers play key roles in rapid disease diagnosis. In this study, we report a one-step detection method based on a nanofibrous sensing platform via the combination of proximity-induced DNA strand displacement (PiDSD), catalytic hairpin assembly (CHA) amplification and thioflavin T (ThT) binding. The interface behaviors on the nanofibrous membrane were studied to promote interface reaction kinetics and thermodynamics. Thrombin was used as a model biomarker, and the nanofibrous sensing platform achieved a limit of detection as low as 1.0 pM, a wide linear range of 50 pM to 5 nM, excellent specificity and good long-term stability. Compared with previous one-step thrombin detection methods, our one-step detection method is label-free, convenient and much more sensitive; it has potential applications for protein detection in point-to-care testing (POCT) and early diagnosis.

Dental caries is one of the most prevalent human oral diseases. The pH of the oral environment is lower than the critical pH point, which causes the demineralization of the hard tissues of teeth and finally leads to dental caries. Therefore, it is important to have an accurate, sensitive detecting assay of the critical pH. An electrospun fibrous membrane was reported to modulate immobilized bromocresol green (BCG), a common pH indicator, by shifting the acid dissociation constant (pKa) from 4.6 to 5.3. Based on spectroscopic and colorimetric characterizations, it was demonstrated that the BCG–polystyrene/polyvinylpyrrolidone (BCG–PS/PVP) electrospun fibrous membrane achieved a sensitive and fast response; moreover, its use exhibits a convenient sensing strategy for visualizing the critical pH point of 5.5 that induces tooth demineralization. Compared with a common pH meter or pH test papers, the BCG–PS/PVP electrospun fibrous membrane could distinguish pH 5.5 from its adjacent pH value with higher accuracy and much more convenience. We could also evaluate the inhibitory effect of the anti-caries agents on bacterial growth and acid production by the platform. The BCG–PS/PVP electrospun fibrous membrane is expected to have potential applications in the monitoring and prevention of early caries.

A novel and rapid room-temperature DNA amplification strategy to simultaneously amplify and detect target HIV DNA was developed by coupling catalyzed hairpin assembly (CHA) with a “non-sequence dependent” nicking endonuclease. In this system, a Nt.BsmAI nicking endonuclease recognition sequence and HIV target DNA sequence were integrated into hairpin probes H1 and H3, respectively. The HIV target DNA was repeatedly used to trigger a CHA reaction, forming numerous H1–H2 duplexes. Then the newly formed sticky ends of H1–H2 caused the hairpin structure of H3 to open. As a result, the double-stranded nicking enzyme recognition site was reunited, which was recognized and cleaved by the nicking enzyme. Finally, the target DNA replica with a sequence matching that of the HIV target DNA was obtained. The above reactions, namely, the CHA reaction, hairpin opening and enzyme cleavage, can be continuously repeated, leading to rapid room-temperature DNA amplification. In addition, this proposed rapid DNA amplification strategy enables picomolar detection of HIV DNA target within 10 min, providing a potential clinical application at point-of-care.

A bicyclic pillar[5]arene derivative fused with a bipyridine side ring, a so-called molecular universal joint (MUJ), was synthesized, and the pair of enantiomers was resolved by high-performance liquid chromatography enantioresolution. The electrochemiluminescent detection based on the ruthenium complex of the enantiopure MUJ showed excellent chiral discrimination toward certain amino acids.

In this work, a novel, rapid and enzyme-free colorimetric biosensor for the detection of nucleic acids has been developed based on entropy-driven (EDC) circuit and DNAzyme-mediated autocatalytic reactions. On sensing the target DNA, the EDC reaction could be initiated and the intact Mg2+-dependent DNAzyme was formed in the reaction product; then, a "mimic target" DNA was generated during the cleavage process of DNAzyme, which in turn catalyzed the EDC reaction corresponding to an autocatalytic process. Meanwhile, numerous G-quadruplex structures were released and further interacted with hemin to form peroxidase-mimicking DNAzyme, inducing a remarkably amplified colorimetric signal. This autocatalytic EDC (AEDC) sensing system exhibited a linear relationship in the range from 20 pM to 10 nM with a detection limit of 10.2 pM. More importantly, benefitting from the Mg2+-dependent DNAzyme-mediated autocatalytic reaction, the detection time (20 min) was significantly reduced compared to that for the reported EDC strategies. In addition, this sensing system has been applied for the detection of target DNA in human serum samples, indicating that it is promising for the on-site and real-time detection of nucleic acids in biomedical research and disease diagnosis.

With the aim of detecting low frequency of drug resistant mutation T790M against wild-type sequences, we reported a two-dimensional signal analysis strategy by combining a three locked nucleic acids (LNAs)-modified probe (LP15-3t) and an α-HL nanopore. The specific hybridization of the LP15-3t probe with the T790M generated unique long two-level signals, including characteristic blocking current and characteristic dwell time. Due to the significant dwell time difference (114.2-fold) and the blocking current difference ranging from 81% to 96%, this two-dimensional signal analysis strategy can simultaneously distinguish T790M sequences with a sensitivity of 0.0001% against wild-type sequences. The LOD of T790M was 0.1 pM. This high discrimination capability would have great potential in the detection of rare mutation sequences and the early monitoring of clinical outcome of NSCLC patients with TKI drug resistance.

Circulating tumor cells (CTCs) have been utilized in the diagnosis and prognosis of tumor. However, the CTC concentration is extremely low to be detected in peripheral blood. Many existing methods suffer from either expensive labeling or complex operation. In this study, we constructed a label- and enzyme-free and sensitive method to detect the breast cancer CTCs. First of all, a probe containing a breast cancer cell-specific aptamer and a complementary single-stranded DNA (trigger DNA P1) were designed. When the target cells are present, the aptamer binds to the CTCs and releases P1 which triggers the strand displacement amplification. This process generates three-way junction structure DNA, the specific translocation signals of which are identified by nanopore assay. The detection limit of tumor cells is 5 in the current experimental setup and can be further reduced. Furthermore, the method is demonstrated in a clinical sample test with high recovery rate and accuracy. Our results suggest that this method could be applied to early diagnosis of metastatic recurrence and prognosis determination.

A facile method to quickly construct polyaniline nanorods (PANI-NR) on acidized titanium (Ti) foils was reported. In this strategy, the acidized Ti foils (Ti-ac) can act as nucleation sites for controllable growth of PANI-NR. Attributed to the synergistic effect of the PANI-NR, doped CNTs and Ti-ac, the PANI-NR/CNT/Ti-ac composite exhibited decreased redox potential and improved electron transport capability. PANI-NR/CNT/Ti-ac was used as an excellent platform for electrochemically sensing ascorbic acid (AA). At a low AA concentration range from 0.4 μM to 125 μM (R2 = 0.998), a high sensitivity of 281 μA mM−1 cm−2 was obtained. The detection limit of AA was 80 nM at a signal-to-noise ratio of 3, which was better than that of reported PANI microtubes (280 nM), PANI nanoparticles (8.3 μM) and PANI film/PAA/MWCNTs (250 nM). A strategy for fabricating the PANI-NR/CNT composite on Ti-ac was provided and the improved electrochemical performance suggests promising application of PANI-NR/CNT/Ti-ac as a sensing platform.

Polyaniline (PANI) microtubes are becoming greatly significant electrochemical materials owing to their large geometric surface area, high conductivity and ideal electrocatalytic activity. In this work, using glucose oxidase (GOx) as a model redox protein, a direct electron transfer strategy based on PANI microtubes was developed for fabricating sensitive biosensors. There is a strong electrostatic interaction between the positively charged PANI microtubes and negatively charged GOx, which promotes the immobilization of GOx on the PANI microtube surface. The immobilized GOx displayed a pair of well-defined quasi-reversible redox peaks with a potential of −0.39 V (vs. SCE) and an ideal electron transfer rate constant (ks) of 3.0 s−1 in PBS solution (0.1 M, pH = 5.5) on the PANI microtubes instead of a bare glass carbon electrode (GCE). The amperometric response of the GOx/PANI microtube modified electrode was linearly proportional to the concentration of glucose in the range of 4.0 μM to 0.80 mM. The glucose detection limit was 0.8 μM at a signal-to-noise ratio of 3, which was better than those reported for the GOx/PANI film (1 mM) and GOx/PANI nanowire (50 μM). The advantages might be attributed to the PANI microtubes' large geometric surface for carrying enzyme (GOx) and efficient electrocatalytic activity to facilitate the direct electron transfer of GOx, as well as an efficient GOx biocatalyst reaction on the microtube surface. A promising application of PANI microtube-based biosensors was offered.

An easy and convenient method for the synthesis of polyaniline (PANI) microtubes was developed using polystyrene (PS) electrospun microfibers as a template. The diameter range of these microtubes was from 0.63 to 2.93 μm. This accuracy was achieved by tuning the template microfiber's diameter. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to study the effect of these microtubes' diameters on their electroactivity. When the diameter is 1.43 μm, the PANI microtube exhibited the maximum electroactivity and a distinguished electrocatalytic oxidation of ascorbic acid (AA). Herein, the PANI microtube was fabricated into an amperometric AA sensor. At the working potential of 0.4 V (vs. SCE), the oxidation current was significantly enhanced by the addition of a low concentration of AA but no such current was observed with other selected small molecules. The AA detection limit was 0.28 μM at the signal-to-noise ratio of 3. A linear relationship was demonstrated between the oxidation current enhancement and the AA concentration ranging from 5.0 μM to 4.7 mM. The detection limit was better than a reported PANI nanotube (1.0 μM), nanofiber (1.7 μM) and nanoparticle (8.3 μM). The high sensitivity could be attributed to the PANI microtube's high conductivity, high electroactivity for the AA electrocatalytic oxidation, as well as its high surface-to-volume ratio resulting in an efficient interaction between the microtube and the AA molecule. A promising application of the PANI microtube-based sensor was offered.

A novel target-triggered DNA autonomous assembly pathway for the formation of one-dimensional DNA polymer chains was proposed based on the catalyzed hairpin assembly and sticky end self-assembly, which led to an enzyme-free and signal amplified gold nanoparticle (AuNP) based colorimetric nucleic acid assay with picomolar sensitivity and specificity.

An enzyme-free and label-free visual sensing strategy was developed for sensitively detecting thrombin using a plasmonic nanoplatform. Both the thrombin-triggered catalytic hairpin assembly (CHA) amplification reaction and G-quadruplex/hemin DNAzyme-controlled plasmonic signal readout were engineered on an electrospun nanofibrous membrane. Owing to its large specific surface area and porous structure, the nanofibrous membrane enhanced the loading capacity of B-H2 and the interface interaction efficiency. This plasmonic nanoplatform was used to perform the sensitive and naked-eye detection of thrombin as low as 1.0 pM in human serum samples. This visual strategy can discriminate thrombin from other co-existing proteins very well. Moreover, the visual sensing platform exhibited excellent reusability and long-term stability. The proposed enzyme-free and label-free plasmonic nanoplatform is low-cost, easy to operate and highly sensitive, and has potential applications in the point-of-care detection of protein biomarkers.

Identification of single-base mismatches has found wide applications in disease diagnosis, pharmacogenetics, and population genetics. However, there is still a great challenge in the simultaneous discrimination of single-base mismatch and full match. Combined with a nanopore electrochemical sensor, a shared-stem structure of molecular beacon was designed that did not need the labeling, but achieved an enhanced signal-to-background ratio of ∼104, high thermodynamic stability to bind with target sequences, and a fast hybridization rate. Fully matched and single-base mismatched sequences were simultaneously discriminated at the single-molecule level, which is expected to have potential applications ranging from the quick detection, early clinical diagnostics to point-of-care research.

A plasmonic nanoplatform to perform an enzyme-free, naked-eye, and trace discrimination of single-base mutation from fully matched sequence is reported. The nanoplatform showed great potential to enhance catalytic hairpin assembly (CHA) amplification efficiency and biocatalytic activity of hemin/G-quadruplex (DNAzyme). When human immunodeficiency virus (HIV) DNA biomarker was used as the model analyst, a naked-eye detection with high selectivity and high sensitivity down to 10–17 M in whole serum was achieved by observing red-to-blue color change. Single-base mismatch and two-base mismatch were detected at the low concentrations of 10–11 and 10–8 M, respectively. The naked-eye detection based on the enzyme-free plasmonic nanoplatform is expected to have potential applications ranging from quick detection and early diagnostics to point-of-care research.

It is known that DNA molecules can be used to build a various of complicated geometrical DNA nanostructures with programmable sequence design, and these DNA nanomaterials show a promising application in biotechnology and biomedicine. However, the construction of large-sized three dimensional DNA-based nanomaterials still remains a challenge. In this work, we propose a new strategy that only employs one target DNA to trigger multiple catalyzed hairpin assembly (CHA) reactions and sticky ends self-assembly to prepare hundreds of nanometer-sized DNA nanoparticles. Moreover, the obtained DNA nanoparticles can be served as efficient biosensors for sensitive colorimetric nucleic acids detection with a detection limit of 7.7 pM

G-quadruplex ligands have been accepted as potential therapeutic agents for anticancer treatment. Thioflavin T (ThT), a highly selective G-quadruplex ligand, can bind G-quadruplex with a fluorescent light-up signal change and high specificity against DNA duplex. However, there are still different opinions that ThT induces/stabilizes G-quadruplex foldings/topologies in human telomere sequence. Here, a sensitive single-molecule nanopore technology was utilized to analyze the interactions between human telomeric DNA (Tel DNA) and ThT. Both translocation time and current blockade were measured to investigate the translocation behaviors. Furthermore, the effects of metal ion (K+ and Na+) and pH on the translocation behaviors were studied. Based on the single-molecule level analysis, there are some conclusions: (1) In the electrolyte solution containing 50 mmol/L KCl and 450 mmol/L NaCl, ThT can bind strongly with Tel DNA but nearly does not change the G-quadruplex form; (2) In the presence of high concentration K+, the ThT binding induces the structural change of hybrid G-quadruplex into antiparallel topology with an enhanced structural stability; (3) In either alkaline or acidic buffer, G-quadruplex form will change in certain degree. All above conclusions are helpful to deeply understand the interaction behaviors between Tel DNA and ThT. This nanopore platform, investigating G-quadruplex ligands at the single-molecule level, has great potential for the design of new drugs and sensors.

Rapid and low-cost diagnosis of multiple infectious diseases is of great significance especially in densely populated or resource-constrained settings. Herein, we developed a one-step fast and label-free imaging array for multiplexed detection of trace avian influenza virus (AIV) DNA biomarkers. By designing a series of specific and efficient catalytic hairpin assembly (CHA) amplification reactions and utilizing thioflavin T, a specific G-quadruplex fluorescence probe, three subtypes of AIV DNA biomarkers (H1N1, H7N9 and H5N1) were simultaneously and quickly detected within only 20 min, which just needed a small reagent volume of 50 μL and a smartphone instead of a spectrometer. With the combination of fluorescence imaging output and grey-level analysis, the array sensor can be on-site with the limit of detection of 136 pM, 141 pM and 129 pM for H1N1, H7N9 and H5N1, respectively. The imaging array also displayed good mismatch discrimination, excellent anti-interference, and real sample application. In view of its advantages of fast detection, low cost and multiplexed analysis, the imaging array is expected to have potential applications for early infectious disease diagnosis in resource-constrained settings.

Self-assembly of DNA nanostructures is of great importance in nanomedicine, nanotechnology and biosensing. Herein, a novel target-catalyzed autonomous assembly pathway for the formation of dendrimer-like DNA nanostructures that only employing target DNA and three hairpin DNA probes was proposed. We use the sticky-ended Y shape DNA (Y-DNA) as the assembly monomer and it was synthesized by the catalyzed hairpin assembly (CHA) instead of the DNA strand annealing method. The formed Y-DNA was equipped with three ssDNA sticky ends and two of them were predesigned to be complementary to the third one, then the dendrimer-like DNA nanostructures can be obtained via an autonomous assembly among these sticky-ended Y-DNAs. The resulting nanostructure has been successfully applied to develop an enzyme-free and signal amplified gold nanoparticle (AuNP)-based colorimetric nucleic acids assay.

Nanoscale transport through nanopores and live-cell membranes plays a vital role in both key biological processes as well as biosensing and DNA sequencing. Active translocation of DNA through these nanopores usually needs enzyme assistance. Here we present a nanopore derived from truncated helicase E1 of bovine papillomavirus (BPV) with a lumen diameter of c.a. 1.3 nm. Cryogenic electron microscopy (cryo-EM) imaging and single channel recording confirm its insertion into planar lipid bilayer (BLM). The helicase nanopore in BLM allows the passive single-stranded DNA (ssDNA) transport and retains the helicase activity in vitro. Furthermore, we incorporate this helicase nanopore into the live cell membrane of HEK293T cells, and monitor the ssDNA delivery into the cell real-time at single molecule level. This type of nanopore is expected to provide an interesting tool to study the biophysics of biomotors in vitro, with potential applications in biosensing, drug delivery and real-time single cell analysis., Active translocation of DNA through nanopores usually needs enzyme assistance. Here authors present a nanopore derived from helicase E1 of bovine papillomavirus (BPV) which acts as a conductive pore embedded in lipid membrane to allow the translocation of ssDNA and unwinding of dsDNA.

Early diagnosis of hepatitis C virus (HCV) infection is still urgently desired as there is a global healthy burden and no vaccine available. In this work, a plasmonic nanoplatform was engineered with catalytic hairpin assembly (CHA) amplification reaction specifically of HCV core protein (HCVcp), G-quadruplex/hemin DNAzyme and nanofibrous membrane together. HCVcp was detected in whole serum at the ultralow concentration of 1.0 × 10−4 pg/mL with naked eye. By testing serum samples from 30 donors with different viral loads, detection sensitivity of the plasmonic nanoplatform turned out to be much better than that of the commercial ELISA kit. In addition, the plasmonic nanoplatform exhibited high specificity, excellent reusability and long-term stability. Naked-eye detection based on the plasmonic nanoplatform is expected to have potential applications in point-of-care testing (POCT) and early diagnosis of hepatitis C and other infectious diseases.

The toxic oxidative damage of G-quadruplexes (G4), linked to neurodegenerative diseases, may arise from their ability to bind and oxidatively activate cellular hemin. However, there have been no precise studies on how telomeric G4 enhances the low intrinsic peroxidase activity of hemin. Herein, a label-free and nanopore-based strategy was developed to explore the enhancement mechanism of peroxidase activity of hemin induced by telomeric G4 (d(TTAGGG)n ). The nanopore-based strategy demonstrated that there were simultaneously two different binding modes of telomere G4 to hemin. At the single-molecule level, it was found that the hybrid structural telomeric G4 directly binds to hemin (the affinity constant (Ka )≈106 m-1 ) to form a tight complex, and some of them underwent a topological change to a parallel structure with an enhancement of Ka to approximately 107 m-1 . Through detailed analysis of the topology and peroxidase activity and molecular modeling investigations, the parallel telomere G4/hemin DNAzyme structure was proven to be preferable for high peroxidase activity. Upon strong π-π stacking, the parallel structural telomere G4 supplied a key axial ligand to the hemin iron, which accelerated the intermediate compound formation with H2 O2 in the catalytic cycle. Our studies developed a label-free and single-molecule strategy to fundamentally understand the catalytic activity and mechanism of telomeric DNAzyme, which provides some support for utilizing the toxic, oxidative-damage property in cellular oxidative disease and anticancer therapeutics.

Electrospun Ru(bpy)32+/carbon nanotube (CNT)/room temperature ionic liquid (IL: 1-butyl-3-methylimidazolium hexafluorophosphate (BMIMPF6)) composite polyacrylonitrile (PAN) nanofibers (CNT/IL/PAN) were successfully fabricated by a one-step electrospinning technique. Utilizing high binding capacity of both carboxylic acid functionalized multiwalled CNTs and large structure anion PF6− (derived from BMIMPF6) to Ru(bpy)32+, the composite nanofibers can serve as an excellent Ru(bpy)32+ immobilization nanomatrix for electrochemiluminescence (ECL) sensing. Using electrochemistry and fluorescence techniques, the composite nanofibers demonstrated superior electron transfer capability and ECL significant amplification. These results might be attributed to a synergistic effect between CNTs and ILs which both have unique electronic and catalytic properties. The ECL intensity from the composite nanofibers was 23.4-fold larger than that of PAN nanofibers, 1.7-fold greater than CNT/PAN nanofibers, and 2.4-fold greater than IL/PAN nanofibers. Furthermore, the composite nanofibers exhibited the largest ECL quenching efficiency, 8.2 times more than that of the PAN nanofibers in the presence of the same phenol concentration (20 μM). Moreover, a nanofiber-based ECL sensor has great selectivity, stability and reusability. In a word, an ECL sensor based on the CNT/IL/PAN nanofibers was developed to sensitively detect phenolic compounds with a low detection limit of 1.0 nM for phenols. A novel ECL amplification strategy based on the synergistic effect between doped CNTs and doped ILs within nanofibers was demonstrated and developed to a promising ECL sensing nanoplatform for ultrasensitive and fast detection.

The term hydrogel describes a type of soft and wet material formed by crosslinked hydrophilic polymers. The distinct feature of hydrogels is their ability to absorb a large amount of water and swell. The properties of a hydrogel are usually determined by the type of polymer and crosslinker, the degree of crosslinking, and the water content. However, a group of hydrogels, called “smart hydrogels”, changes properties in response to environmental changes or external stimuli. Recently, DNA or DNA-inspired responsive hydrogels have attracted considerable attention in construction of smart hydrogels because of the intrinsic advantages of DNA. As a biological polymer, DNA is hydrophilic, biocompatible, and highly programmable by Watson-Crick base pairing. DNA can form a hydrogel by itself under certain conditions, and it can also be incorporated into synthetic polymers to form DNA-hybrid hydrogels. Functional DNAs, such as aptamers and DNAzymes, provide additional molecular recognition capabilities and versatility. In this review, we discuss DNA-based hydrogels in terms of their stimulus response, as well as their applications.

The inducible coactivator PGC-1αplays master regulator in mitochondrial biogenesis and thermogenesis in brown adipose tissues (BATs). BAT is a natural antiobesity organ which dissipates chemical energy in the form of heat through specialized mitochondrial protein UCP-1. Eletroacupuncture (EA) has been widely used as an alternative treatment for obesity and its related disorders such as type 2 diabetes. The molecular mechanism of electroacupuncture on treatment of obesity is still unclear. We hypothesized that electroacupuncture induced PGC-1αexpression to increase the energy expenditure in BAT. Rats were randomly divided into control group and electroacupuncture treatment group. We investigated the effects of electroacupuncture at Zusanli (ST36) acupoint on the expressions of PGC-1αand its associated genes in the BAT of rats using real-time PCR and western blotting. We found that electroacupuncture effectively induces the expression of PGC-1αand UCP-1 by 4-fold and 5-fold in the BAT of rats, respectively. Our results indicated that the molecular mechanism of electroacupuncture for the treatment of obesity may be, or at least partially, through induction of both PGC-1αand UCP-1 expressions to increase energy expenditure in BAT.

Methodologies to detect disease biomarkers at ultralow concentrations can potentially improve the standard of living. A facile and label-free multi-amplification strategy is proposed for the ultrasensitive visual detection of HIV DNA biomarkers in real physiological media. This multi-amplification strategy not only exhibits a signficantly low detection limit down to 4.8 pM but also provides a label-free, cost-effective and facile technique for visualizing a few molecules of nucleic acid analyte with the naked eye. Importantly, the biosensor is capable of discriminating single-based mismatch lower than 5.0 nM in human serum samples. Moreover, the visual sensing platform exhibits excellent specificity, acceptable reusability and a long-term stability. All these advantages could be attributed to the nanofibrous sensing platform that 1) has a high surface-area-to-volume provided by electrospun nanofibrous membrane and 2) combines glucose oxidase (GOx) biocatalysis, DNAzyme-catalyzed colorimetric reaction and catalytic hairpin assembly (CHA) recycling amplification together. This multi-amplification nanoplatform promises label-free and visual single-based mismatch DNA monitoring with high sensitivity and specificity, suggesting wide applications that range from virus detection to genetic disease diagnosis.

A chemiluminescence (CL) amplification platform based on HCC/Lucigenin&GOx (HLG) film was developed. Hollow structural calcium carbonate (HCC) particles were used as alternative materials for carrying both enzyme and CL reagent. The model enzyme (GOx), immobilized in confined space of HCC particles, exhibited an improved biocatalysis. The Michaelis constant (Km) and the enzymatic rate constant (kcat) were determined to be 0.209 μM and 2.21 s−1, respectively, which are much better than those of either free GOx in aqueous solution or the GOx immobilized on common nanomaterials. Based on the HLG platform, CL signal was effectively amplified and visualized after adding trace glucose, which could be attributed to the HCC particles’ high biocompatibility, large specific surface area, attractive interfacial properties and efficient interaction with analyses. The visual CL bioplatform showed an excellent performance with high selectivity, wide linear range and low detection limit for sensing trace glucose. Because it eliminates the need of complicated assembly procedure and enables visualization by the naked eye, the sensitive and selective CL bioplatform would provide wide potential applications in disease diagnosis and food safety.

Enzyme-free signal amplification has enabled sensitive in vitro detection of biomolecules such as proteins and nucleic acids. However, monitoring targets of interest in live cells via enzyme-free amplification is still challenging, especially for analytes with low concentrations. To the best of our knowledge, this paper reports the first attempt to perform mRNA imaging inside live cells, using a nonenzymatic hairpin DNA cascade reaction for high signal gain, termed a hairpin DNA cascade amplifier (HDCA). In conventional nucleic acid probes, such as linear hybridization probes, mRNA target signaling occurs in an equivalent reaction ratio (1:1), whereas, in HDCA, one mRNA target is able to yield multiple signal outputs (1:m), thus achieving the goal of signal amplification for low-expression mRNA targets. Moreover, the recycled mRNA target in the HDCA serves as a catalyst for the assembly of multiple DNA duplexes, generating the fluorescent signal of reduced MnSOD mRNA expression, thus indicating amplified intracellular imaging. This programmable cascade reaction presents a simple and modular amplification mechanism for intracellular biomarkers of interest, providing a significant boost to the search for clues leading to the accurate identification and effective treatment of cancers.

The application of cancer theranostics depends on the development of multifunctional nanostructured platforms for accurate cell targeting and controlled drug release, imaging, and therapy. Herein, a comprehensive, easily fabricated anticancer theranostic platform with a high drug-loading capacity, termed an aptamer-functionalized calcium carbonate (CaCO3 ) nanostructure (apt-CCN), is reported. Flow cytometry and confocal fluorescence microscopy studies demonstrate that apt-CCNs can specifically bind to target cancer cells, but not to control cells, and that they possess highly efficient internalization to target cancer cells. This smart nanostructure selectively reaches the lysosomes through receptor-mediated endocytosis and is responsive to the relatively low lysosome pH (4.5-5.5), which facilitates the release of doxorubicin. The apt-CCN platform offers targeted and efficient drug transport, as well as target-specific delivery of imaging agents for cancer diagnosis and therapy.

Review: the most recent and important developments in nucleic acid-based logical devices and their evolution from in vitro, through cellular, even towards in vivo biological applications; 44 refs.

Researchers increasingly visualize a significant role for artificial biochemical logical systems in biological engineering, much like digital logic circuits in electrical engineering. Those logical systems could be utilized as a type of servomechanism to control nanodevices in vitro, monitor chemical reactions in situ, or regulate gene expression in vivo. Nucleic acids (NA), as carriers of genetic information with well-regulated and predictable structures, are promising materials for the design and engineering of biochemical circuits. A number of logical devices based on nucleic acids (NA) have been designed to handle various processes for technological or biotechnological purposes. This article focuses on the most recent and important developments in NA-based logical devices and their evolution from in vitro, through cellular, even towards in vivo biological applications.

Ruthenium nanoparticle tris(2,2'-bipyridyl)ruthenium(II) bis(hexafluorophosphate) (Ru(bpy)3(PF6)2, RuNP) was self-assembled on polystyrene (PS) electrospun microfibers. The formation of RuNP is attributed to the sulfonated PS (SPS) microfibers' high adsorptive capability of 94% for Ru(bpy)3(2+), as well as the strong interaction between the Ru(bpy)3(2+) and ionic liquid (1-butyl-3-methylimidazolium hexafluorophosphate, BMIMPF6). The RuNP/SPS microfibers exhibited an enhanced electrochemiluminescence (ECL) emission, 2.3 times higher than that from Ru(bpy)3(2+)/SPS microfibers and 6.6 times higher than that from Ru(bpy)3(2+)/SPS continuous thin films. It is worthy of note that, as a result of the hydrophobic nature of the RuNP, the transfer of water-insoluble α-naphthol is accelerated, and thus the α-naphthol ECL quenching efficiency is enhanced. An ECL sensor based on the RuNP/SPS microfibers was fabricated and used to detect low concentrations of α-naphthol. The detection limit was of 1.0 nM (S/N3), and the linear response ranged from 0 to 18 μM. This sensor has been successfully applied to measure the α-naphthol content in pesticide carbaryl samples. Our work provides a very simple and cost-effective method to fabricate RuNP on polymer microfibers with great potential in the field of chemo/biosensors.

Polystyrene electrospun optical fibrous membrane (EOF) was fabricated using a one-step electrospinning technique, functionalized with glucose oxidases (GOD/EOF), and used as a quick and highly sensitive optical biosensor. Because of the doped iridium complex, the fibrous membrane emitted yellow luminescence (562 nm) when excited at 405 nm. Its luminescence was significantly enhanced with the presence of extremely low concentration glucose. The detection limit was of 1.0 × 10(-10) M (S/N = 3), superior to that of reported glucose biosensor with 1.2 × 10(-10) M. A linear range between the relative intensity increase and the logarithm of glucose concentration was exhibited from 3.0 × 10(-10) M to 1.3 × 10(-4) M, which was much wider than reported results. Notably, the response time was less than 1 s. These high sensitivity and fast response were attributed to the high surface-area-to-volume of the porous fibrous membrane, the efficient GOD biocatalyst reaction on the fibers surface, as well as the fast electron or energy transfer between dissolved oxygen and the optical fibrous membrane.

A water-soluble covalently linked C60-porphyrin compound–C60Por is reported to show enhanced DNA-cleaving activity. The highly enhanced DNA-cleaving activity is expected to be ascribed to the high water-solubility and affinity of C60Por to DNA, as well as to the accessibility to the charge-separated states C60˙−–Por˙+ formed under radiation.

A water-soluble covalently linked C(60)-porphyrin compound-C(60)Por is reported to show enhanced DNA-cleaving activity. The highly enhanced DNA-cleaving activity is expected to be ascribed to the high water-solubility and affinity of C(60)Por to DNA, as well as to the accessibility to the charge-separated states C(60)˙(-)-Por˙(+) formed under radiation.

Electrospun Ru(bpy)(3)(2+)-doped Nafion (ERDN) nanofibers were successfully fabricated by a one-step electrospinning technique. The diameters of the nanofibers range from 120 nm to 220 nm. The ERDN nanofibers, evenly distributed on the substrate with their random orientations, form a porous 3-D structure nanofibrous membrane. Compared to continuous thin films, the obtained nanofibrous membrane maintains a higher surface-area-to-volume ratio, a larger amount of immobilized-Ru(bpy)(3)(2+), and a better stability of the Ru(bpy)(3)(2+) immobilization. As a result, the nanofibers' ECL signal is enhanced approximately 15-fold and stable (does not change during continuous CV scans for 50 cycles). As a sensing platform, the nanofibers can sensitively detect low concentration phenolic compounds by monitoring the phenol-dependent ECL intensity change. The detection limit for phenol is 1.0 nM based on a signal/noise ratio3, which is comparable or better than that in reported phenol assays. In addition, the nanofibers exhibit excellent ECL behaviors on Au, Pt, GC and ITO electrodes. A great potential for ERDN nanofibers-based ECL sensors is offered.