Herein, we describe a novel homogenous electrochemical aptasensor for the ultrasensitive detection of ractopamine (RAC) based on the signal amplification of a hairpin DNA cascade amplifier (HDCA) and electrocatalysis of dsDNA-templated copper nanoparticles. The present electrochemical aptasensor employs a label-free “turn-on” strategy with enzyme-free amplification. Briefly, the target RAC triggers the HDCA autocatalytic process of two ingeniously designed complementary hairpin DNA, which results in the formation of nucleic-acid-stabilized copper nanoparticles (dsDNA/CuNPs). Thus, with the mimic oxidase catalytic character of the dsDNA/CuNPs and enzyme-free hairpin DNA cascade amplifier reactions, a remarkable electrochemical response can be achieved, which is dependent on the concentration of the target RAC. Taking advantage of the highly amplified efficiency of trigger recycling and the excellent electrochemical catalytic response of dsDNA/CuNPs, the proposed strategy is capable of detecting RAC ultrasensitively. Under optimal conditions, the electrochemical signal increases with an increase in the target RAC concentration in the wide dynamic range from 1 × 10−12 M to 3 × 10−7 M with a detection limit (LOD) of 3 × 10−13 M. The proposed system does not need any sophisticated operation procedures such as electrode immobilization and expensive labeling. Therefore, the devised label-free and enzyme-free amplification homogeneous electrochemical aptasensor strategy may become an alternative method for the simple, sensitive, and selective detection of biological small molecules, proteins, nucleic acids and nuclease activity.

•A portable personal glucose meter was used to detect PDGF-BB sensitively.•Catalytic and molecular beacon system was used for amplify signal.•Cation exchange reaction (CX) is key for ultrasensitive PDGF-BB assay.•By CX, greater aptamer payload and thousands of released Zn2+ was achieved.•The released Zn2+ realized the cycle by the catalyzing cleavage of 8–17 DNAzyme.Sensitive and rapid detection of platelet-derived growth factor BB (PDGF-BB), a cancer-related protein, could help early diagnosis, treatment, and prognosis of cancers. Although some methods have been developed to detect PDGF-BB, few can provide quantitative results using an affordable and portable device that is suitable for home use or field application. In this work, we report the first use of a portable kind of personal glucose meter (PGM) combining a catalytic and molecular beacon (CAMB) system with a cation exchange reaction (CX reaction) for ultrasensitive PDGF-BB assay. It realized the amplification of the detection in three ways, including greater aptamer payload on nanoparticles, CX reaction releasing thousands of Zn2+ and the cycle by the catalyzing cleavage of 8–17 DNAzyme. In the process, with the addition of PDGF-BB into the aptasensor, the specific recognition between aptamer and protein was initiated resulting in the combination of ZnS NNC for further CX reaction to release thousands of Zn2+, which could cleave the substrate DNA in the CAMB system realizing multiple cycle. The cleaved DNA fragment was designed with invertase-labeled could convert sucrose into glucose which could be detected and quantified by PGM accompanying with the change of color of the control window from yellow to green. The enhanced signal of the PGM has a relationship with the concentration of PDGF-BB in the range of 3.16×10–16 M to  3.16×10–12 M, and the detection limit is 0.11 fM. Moreover, the catalytic and cleavage activities of 8–17 DNAzyme can be achieved in solution; thus, no enzyme immobilization is needed for detection. The triply amplified strategy showed high selectivity, stability, and applicability for detecting the desired protein.

A crowded TTE DNA structure for the preparation of Cu NPs with enhanced fluorescence was prepared and applied for the ultrasensitive detection of target DNA.

Because of the intrinsic importance of transcription factors (TFs) as targets in clinical diagnosis and drug development, the simple and sensitive detection of TFs is very essential for biological studies and medical diagnostics. However, most of the reported methods involve complicated operations or labelling processes and in addition, an electrochemiluminescence (ECL) method with various distinct advantages hasn't been developed for TF detection until now. In this work, we describe a novel simple and efficient strategy for label-free ECL detection of transcription factors with hybridization chain reaction (HCR) signal amplification. Based on the Ag+-stabilized self-assembly triplex DNA, in the presence of TFs, TFs specifically bond to the duplex DNA (dsDNA) recognition probes, resulting in the separation of target DNA from the triplex structure. With the SH-capture probe DNA assembled gold electrode, the presence of the target DNA and helper DNA-1 and helper DNA-2 leads to the formation of long chain dsDNA polymers on the gold electrode surface through hybridization chain reaction, which allows the intercalation of numerous ECL indicators Ru(phen)32+ (phen = 1,10-phenanthroline) into the dsDNA grooves, resulting in significantly amplified ECL signal output. The proposed strategy combines the amplification power of the HCR and the inherent high sensitivity of the ECL technique, resulting in the sensitive detection of transcription factors with a detection limit of as low as 0.017 nM and a broad dynamic range from 0.05 to 2 nM. The distinct advantage of the method is that it is label-free, has high sensitivity and requires no separation of the signal generation strand, which boosts the potential application for real sample detection.

Determination of nucleotide kinase activity is valuable due to its importance in regulating nucleic acid metabolism. In this work, a novel and simple colorimetric method for discriminative detection of T4 polynucleotide kinase (PNK) through a MnO2 nanosheet-3,3′,5,5′-tetramethylbenzidine (TMB) colorimetric system is developed. In aqueous solution at pH 7.4, the MnO2 nanosheet, a mimic enzyme, rapidly converts colourless TMB to a blue oxidized form. In the presence of T4PNK, a hairpin DNA could be phosphorylated and further cleaved by λ exonuclease (λ exo) and the ssDNA fragments would adsorb on the surface of the MnO2 nanosheet, resulting in the inhibition of the catalytic effect of the MnO2 nanosheet to the oxidation of TMB and weakening of the blue color of oxidized TMB with the absorption peak centred at 652 nm. The presence of T4PNK can in this way easily be detected with the naked eye, based on the obvious colour change. The experimental results revealed that the advanced strategy was sensitive for detecting T4PNK in the concentration range from 0.005 to 10 U mL−1 with a detection limit of 0.005 U mL−1. In addition, the relative standard deviation was 3.92% in 3 repetitive assays of 10 U mL−1 T4PNK, indicating that the reproducibility of this strategy was acceptable. By comparison with other relevant fluorogenic assays, the sensitivity of the developed chromogenic assays was satisfactory. The present approach opens the possibility for the design of simple and sensitive colorimetric assays for polynucleotide kinase using inexpensive and available TMB as a universal chromogenic compound. Therefore, besides desirable sensitivity and selectivity, due to the colorimetric system, the developed method for T4PNK detection also showed cost-effective, reliable and simplified operations and strategy, which has considerable potential for biological process research.

•A label-free, enzyme-free, ultrasensitive TFs biosensor was constructed.•AT-rich dsDNA/Cu NPs was used as the signal in the assay.•Hairpin DNA cascade reaction amplification (HDCA) was employed in the biosensor.•It is a separation-free procedure conducted in a homogeneous solution.Detection and quantification of specific protein with ultralow concentration play a crucial role in biotechnological applications and biomedical diagnostics. In this paper, a label-free and enzyme-free amplified fluorescent biosensor has been developed for transcription factors detection based on AT-rich double-stranded DNA-templated copper nanoparticles (ds DNA/Cu NPs) and hairpin DNA cascade reaction. This strategy was demonstrated by using nuclear factor-kappa B p50 (NF-κB p50) and specific recognition sequences as a model case. In this assay, a triplex consists of double-stranded DNA containing NF-κB p50 specifically binding sequences and a special design single-stranded DNA (Trigger) which is able to activate the hairpin DNA cascade amplifier (HDCA). In the presence of NF-κB p50, the triplex became unstable since the target bound to the recognition sequences with strong affinity. The selective binding event confirmed that the Trigger was successfully released from the triplex and initiated HDCA to yield the product which could effectively template the formation of fluorescent Cu NPs. The experimental results revealed that the advanced strategy was ultra-sensitive for detecting NF-κB p50 in the concentration range from 0.1 to 1000 pM with a detection limit of 0.096 pM. In addition, the relative standard deviation was 4.08% in 3 repetitive assays of 500 pM NF-κB p50, which indicated that the reproducibility of this strategy was acceptable. Besides desirable sensitivity, the developed biosensor also showed high selectivity, cost-effective, and simplified operations. In addition, the proposed biosensing platform is versatile. By conjugating with various specific recognition units, it could hold considerable potential to sensitive and selective detect various DNA-binding proteins and might find wide applications in biomedical fields.

•A simple label-free biosensor was developed for NAD+ detection with DNA/AgNCs probe.•The specific NAD+-ligation triggered nanocluster beacon from split to intact.•Logic gate operation was investigated based on the NAD+ sensor.•The construction of ATP aptasensor showed the platform universal.•The platform is highly sensitive and selective.A novel fluorescent label-free “turn-on” NAD+ and adenosine triphosphate (ATP) biosensing strategy is proposed by fully exploiting ligation triggered Nanocluster Beacon (NCB). In the presence of the target, the split NCB was brought to intact, which brought the C-rich sequence and enhancer sequence in close proximity resulting in the lightening of dark DNA/AgNCs (“On” mode). Further application was presented for logic gate operation and aptasensor construction. The feasibility was investigated by Ultraviolet–visible spectroscopy (UV–vis), Fluorescence, lifetime and High Resolution Transmission Electron Microscopy (HRTEM) etc. The strategy displayed good performance in the detection of NAD+ and ATP, with the detection limit of 0.002 nM and 0.001 mM, the linear range of 10–1000 nM and 0.003–0.01 mM, respectively. Due to the DNA/AgNCs as fluorescence reporter, the completely label-free fluorescent strategy boasts the features of simplicity and low cost, and showing little reliance on the sensing environment. Meanwhile, the regulation by overhang G-rich sequence not relying on Förster energy transfer quenching manifests the high signal-to-background ratios (S/B ratios). This method not only provided a simple, economical and reliable fluorescent NAD+ assay but also explored a flexible G-rich sequence regulated NCB probe for the fluorescent biosensors. Furthermore, this sensing mode was expanded to the application of a logic gate design, which exhibited a high performance for not only versatile biosensors construction but also for molecular computing application.

•DNA–gold nanoparticles network was as signal amplification unit.•An easy assembly method by the linkage of benzenedithiol bridge.•It avoided the direct incubation of the solid electrode with the DNA.•A highly sensitive DNA methyltransferase activity assay was fabricated.In this work, a highly sensitive electrochemical DNA methyltransferase (MTase) activity assay was fabricated with DNA–gold nanoparticles (Au NPs) network as signal amplification unit and an easy assembly method by the linkage of benzenedithiol bridge. By two complementary AuNPs modified single-stranded DNA, DNA–gold nanoparticles network was self-assembled. With the linkage of benzenedithiol bridge, the DNA network structure was immobilized on the surface of gold electrode through the covalent Au–S bond. In the presence of Dam MTase, the special sites of DNA–AuNPs network were methylated and could not be digested by restriction endonuclease Mbo I. Thus the loaded electrochemical indicator Methylene blue (MB) was MB molecules still remained on the DNA–Au NPs network. The electrochemical response depended on the methylated degree, which could be used to detect MTase activity. By the differential pulse voltammetry (DPV), it was demonstrated that a linear relationship between the DPV response and logarithm of Dam concentration ranged from 0.075 to 30 U/mL, achieving a low detection limit of 0.02 U/mL. The use of benzenedithiol avoided the direct incubation of the solid electrode with the capture DNA probe under complex and harsh conditions. Therefore the immobilization of DNA–AuNPs network was easy to be carried out, which is favorable for the specially high stability and reproducibility of the electrochemical biosensor.Benzenedithiol linked DNA–gold nanoparticles network based electrochemical biosensors for DNA MTase activity.

In the present study, based on the mimic oxidase catalytic character of nucleic-acid-stabilized silver nanoclusters (DNA/AgNCs) and hybridization chain reactions for signal amplification, the fabrication of a label-free sensitive “turn-on” electrochemical aptasensor for the amplified determination of lysozyme was demonstrated. First, the designed DNA duplex was modified on the electrode. With the specific binding of the target, lysozyme and its aptamer, the lysozyme-binding DNA sequence was liberated, exposing the induced DNA sequence, which in turn triggered the formation of the supersandwich DNA structure. Because the cytosine-rich sequence was designed ingeniously on the DNA sequence, DNA/AgNCs were formed on the supersandwich DNA structure. The peroxidase-like character of DNA/AgNCs produced detectable electrochemical signals for the lysozyme aptasensor, which showed a satisfying sensitive detection of lysozyme with a low detection limit of 42 pM and a wide linear range of 10−10 M to 10−5 M.

Thioflavin T (ThT), as one of the most exciting fluorogenic molecules, boasts the “molecular-rotor” ability to induce DNA sequences containing guanine repeats to fold into G-quadruplex structures. It has been demonstrated to sense this change by its remarkable fluorescence enhancement. In this work, taking T4 polynucleotide kinase (PNK) as a model, the ThT/G-quadruplex based platform and λexonuclease (λexo) cleavage reaction were combined to design a label-free “turn-on” strategy for fast, simple and accurate detection of T4 PNK activity and its inhibition. In the presence of T4 PNK, the designed thioflavin T based molecular beacon (TMB) DNA probe could be phosphorylated and then digested by the cleavage of λexo, releasing the G-quartets. These then bound to ThT to form ThT/G-quadruplexes with an obvious fluorescence generation, for the “turn-on” detection of T4 PNK. In comparison to traditional methods, the proposed TMB probe is convenient, requiring no sophisticated labeling and separation processes and displaying high analytical performance. It exhibits a satisfying detection result for the activity of T4 PNK with a low detection limit of 0.001 U mL−1. This is not only meaningful for further research on disease-related biochemical processes, but also valuable for molecular-target therapies.

Development of strategies for the sensitive and selective detection of the folate receptor (FR) that are simple and low cost is of great importance for assessing cancer therapeutics due to its crucial role in physiological, pharmacological and pathological processes. In this paper, gold nanoparticle (AuNP)-based novel ratiometric colorimetry for the detection of the folate receptor (FR) is proposed based on terminal protection of small-molecule-linked DNA. The single-stranded DNA (ssDNA) terminally tethered to folic acid (FA) is protected from degradation by exonuclease I (Exo I) when the FA moiety is bound to FR. The hybridization between FR-protected DNA and DNA-functionalized Au NPs generated a red-to-purple colour change, allowing the visual detection of FR. The detection limit of FR can be as low as 0.33 ng mL−1 with the naked eye. It provides a promising strategy for visual detection of the binding event of FA to its protein receptor-FR with advantages such as simplicity, high selectivity, and a wide linear range.

Phosphorylation of DNA by polynucleotide kinase (PNK) plays an important role in a variety of cellular processes, such as nucleic acid metabolism and DNA damage repair. Strategies for a simple and effective PNK assay are in urgent need. Here we report the development of a simple label-free electrochemical method for a PNK assay based on the mimic peroxidase character of a prepared TiO2 nanotube array (NTA) and its specific attachment to phosphorylated DNA. The concentration of T4 PNK was successfully detected by the TiO2 NTA electrode with a detection limit of 0.15 U mL−1. In the process, only a one-step incubation process was used and no labeling processes were involved. In addition, the treated TiO2 foil was anodized into the nanotube array and it was directly used as the electrode without any other complex modification processes which avoided the leakage of an electrode modifier, and so the TiO2 nanotube array with excellent stability and flexibility is worthy of use in real time monitoring.

•Photoinduced electron transferred between Ag nanoclusters and HRP-DNAzyme.•A label-free fluorescent aptasensor was fabricated for PDGF-BB.•A logic gate of hemin and PDGF-BB also shows satisfying result.•The process only involved one-step incubation.Platelet-derived growth factor-BB (PDGF-BB) is often overexpressed in human malignant tumors as an indicator for tumor angiogenesis. Here by the photoinduced electron transfer (PET) between DNA-Ag fluorescent nanoclusters (NCs) and G-quadruplex/hemin complexes, we present a sensitive label-free fluorescent sensor for PDGF-BB. In the presence of PDGF-BB, the specific conjugation with its aptamer induced the conformational change of the duplex-like DNA sequence, releasing the G-quadruplex sequence part. Then in the presence of hemin and K+, the horseradish peroxidase mimicking DNAzyme (HRP-DNAzyme) was formed. With the electron transfer between the DNA-Ag NCs to the hemin Fe (III) center of HRP-DNAzyme, the PET occurred with a decrease in the fluorescence intensity of the DNA-Ag NCs. The detection performance such as selectivity, linear dynamic range, sensitivity, and the quenching capability of HRP-DNAzyme were estimated. The detection range for PDGF-BB is from 5×10−13 to 1×10−8 M and the detection limit is 1×10−13 M. The experimental results confirmed that the novel fluorescent aptasensor possessed a good sensitivity and high selectivity for PDGF-BB. In addition, the developed probe is nontoxic, label-free only involving one-step hybridization without sophisticated fabrication process. Furthermore, based on this quenching mode occurred by PDGF-BB and hemin, using PDGF-BB and hemin as inputs and the fluorescence signal as an output, a logic gate has been fabricated.

In this paper, horseradish peroxidase-mimicking DNAzyme (HRP-DNAzyme) and Prussian blue (PB)–gold (Au) nanocomposites were designed as versatile electrochemical sensing platforms for the amplified detection of DNA, Hg2+ and adenosine triphosphate (ATP). By the conjugation of the target probe with the capture probe, a conformational change resulted in the formation of HRP-DNAzyme on the PB–Au modified electrode. The redox of HRP-DNAzyme (red) was efficiently carried out in the presence of H2O2, in which PB acted as a mediator stimulating the biocatalytic functions of HRP-DNAzyme and actuated a catalytic cycle bringing an amplified signal. Specific recognition of the target DNA, Hg2+ and ATP allowed selective amperometric detection of the target molecule. The detection limits of DNA, Hg2+ and ATP were 50 nM, 30 pM and 3 nM, respectively. The highlight of this work is that the catalytic cycle between PB–Au nanocomposites and HRP-DNAzyme was adequately utilized in the amplification platform for versatile sensing. The novel electrocatalytic biosensor involving only one-step incubation exhibited a wide linear range, low detection limit, and satisfactory selectivity and operational stability. The proposed approach provided an ease-of-use and universal reporting system with a simple design and easy operations.

In this paper, we have designed a signal amplified method for the electrochemical determination of polynucleotide kinase activity. It is based on (a) the peroxidase-like activity of magnetite microspheres (MNPs), (b) the specific recognition capabilities of titanium dioxide (TiO2) with the phosphate groups of the capture probe and (c) the DNA dendrimer structure for signal amplification. MNPs coated with TiO2 (TMNPs) were prepared and characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. TMNP–DNA dendrimers were formed by the hybridization of captured nucleic acids with a link probe. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were carried out to study the electrocatalytic process. The formation of the TMNP–DNA dendrimer structures was related to the phosphorylated capture probe and further to the activity of polynucleotide kinase, which was the base of the polynucleotide kinase detection. The TMNP–DNA dendrimer based biosensor showed sensitive detection of polynucleotide kinase with a satisfying result; a low detection of 0.003 U mL−1 and wide linear range of 0.01 to 30 U mL−1 were achieved. Additionally, the present TMNP–DNA dendrimer based biosensor also demonstrated excellent selectivity, stability and reproducibility.

We report on the detection of trace quantities of melamine (MA) by a colorimetric method that exploits the conformational change of hemin G-quadruplex-DNAzyme. The addition of MA to hemin G-quadruplex-DNAzyme structure containing thymine bases causes the thymine in the DNAzyme to interact with MA via a stable triple H-bond and leads to a conformational change. This, in turn, affects the peroxidase-like activity of hemin which is determined colorimetrically at 450 nm by adding 3,3’,5,5’-tetramethylbenzidine and hydrogen peroxide. The method was applied to the colorimetric determination of MA over a wide range of concentrations (0.2 to 24 μM) with a detection limit of 80 nM. The effect also can be detected with bare eyes. The method was successfully applied to the determination of MA in spiked milk powder.

•Potassium ions sensor was fabricated by the conformational change of DNA.•Signal amplification was achieved by the biocatalytic and mediated co-effect.•Different metal types (iron, nickel and copper) of different sizes were tested and compared first time.•Fc unit stimulated the biocatalysis of HRP-DNAzyme.•The system realized the electrical contact of HRP-DNAzyme by Fc as relay.Sensitive and selective sensors need to be explored to detect the physiological potassium level due to its important role in the living organisms. In the present system, a novel electron transfer mediator actuated electrocatalytical biosensor was demonstrated to assay K+ based on the conformational change of DNA. With the hybridization between the complementary bases and the self-folding of guanine-rich nucleic acid sequence, the horseradish peroxidase-mimicking enzyme (HRP-DNAzyme) was formed and brought to approach the ferrocene (Fc) unit on Au nanoparticles (AuNPs). Thus, in the system, Fc unit acted as the relay, stimulating the electrical contact of HRP-DNAzyme with the electrode to obtain the bioelectrocatalyze reduction signal. Under the Fc actuated catalysis of HRP-DNAzyme and amplification of Au nanoparticles, the obtained biosensor exhibited a sensitive detection for K+. A satisfying result of a wide linear range and low detection limit were obtained with the novel electrocatalytical biosensor which was then applied in real samples.

•A dual hairpin-like MB was built for the selective and specific detection of MA.•The sensor was more stable and lower toxicity than typical MB structure.•The system has high quenching efficiency based on contact quenching mechanism.•The detection limit of MA is about 4000 times lower than the MA safety limit.This study presents a novel dual hairpin-like molecular beacon (MB) for the selective and sensitive detection of melamine (MA) based on the conjugation of MA and thymine. In this protocol, the coordination between coralyne and adenosine (A) leaded a dual hairpin-like MB and the fluorophore–quencher pair is close proximity resulting in the fluorescence quenching. With the addition of MA, it conjugated with thymine in the loop part of dual hairpin-like MB by triple H-bonds, triggering the dissociation of the dual hairpin-like MB. The resulting spatial separation of the fluorophore from quencher induced the enhancement in fluorescence emission. Under the optimized conditions, the sensor exhibited a wide linear range of 8×10−9–1.6×10−5 M (R2=0.9969) towards MA, with a low detection limit of 5 nM, approximately 4000 times lower than the Drug Administration and the US Food estimated MA safety limit. The real milk samples were also investigated with a satisfying result.

Silver oxide nanowire arrays (Ag2O NWAs) were first synthesized on a copper (Cu) rod by a simple and facile wet-chemistry approach without using any surfactants. The as-synthesized Ag2O NWA/Cu rod not only can be used as an integrated electrode (called a Ag2O NWA/CRIE) to detect hydrazine (HZ) but also can serve as the catalyst layer for a direct HZ fuel cell. The current density of HZ oxidation on Ag2O NWA (94.4 mA cm–2) is much bigger than that on a bare Cu rod (3.9 mA cm–2) at −0.6 V, and other Ag2O NWAs have the lowest onset potential (−0.85 V). This suggests that a Ag2O NWA integrated electrode has potential application in catalytic fields that contain the HZ fuel cell.Keywords: amperometric sensor; electrocatalysis; hydrazine; integrated electrode; silver oxide;

In the present work, sensitive and stable sandwiched electrochemical immunosensing strategies with graphene (Gr)/gold nanoparticles (GNP) composite as the immobilization platform and ferrocene (Fc) derivatives as labels were proposed. Under the optimal choice and the proposed immunosensing strategy, with 1,1′-ferrocenedicarboxylic acid label, the human IgG could be detected specifically with a low detection limit of 0.4 ng/mL and a wide linear dynamic range from 1 to 300 ng/mL. Meanwhile, the electrochemical behavior of three different ferrocene derivatives labels was studied.Highlights► A sensitive electrochemical immunosensor was fabricated with new platform and label. ► Gold nanoparticles/graphene platform was fabricated with an easy electrodeposition method. ► 1,1′-ferrocenedicarboxylic acid was first introduced as the label. ► Three different ferrocene derivatives labels were compared and discussed.

Determination of nucleotide kinase activity is valuable due to its importance in regulating nucleic acid metabolism. Herein, we describe a strategy for simply and accurately determining nucleotide kinase activity by TiO2 nanotubes mediated signal transition and Au nanoparticles amplification. In this method, DNA containing 5′-hydroxyl group is self-assembled onto a gold electrode and used as a substrate for T4 polynucleotide kinase (PNK). By the specific immobilization affinity of TiO2 nanotubes with the phosphorylated DNA, TiO2 nanotubes were linked with phosphorylated substrate DNA on the electrode. And then Au nanoparticles modified 5′-phosphate DNA was conjugated with the TiO2 nanotubes and hybridized with methylene blue labeled signal DNA. Because gold nanoparticles have high loading of signal indicator methylene blue, the electrochemical signal is generated and amplified. It presents an excellent performance with wide linear range and low detection limit. Additionally, inhibition effects of some salts have also been investigated. The developed method is a potentially useful tool in researching the interactions between proteins and nucleic acids and provides a diversified platform for a kinase activity assay.Highlights► TiO2 nanotubes for the immobilization of phosphorylated DNA were synthesized by a novel method. ► T4 polynucleotide kinase (PNK) activity was determined in the sandwich DNA structure. ► Au nanoparticles were used for signal amplification. ► Wide linear range and low detection limit were obtained in the present system.

In the present study, based on multifunctional Dual-Hairpin DNA structure, a simple, fast and high sensitive assay for the detection of DNA, thrombin and adenosine triphosphate (ATP) was demonstrated. DNA sequence labeled with methylene blue (MB), which was designed as single-stranded DNA (ssDNA) matching with target DNA, thrombin, or ATP aptamer, hybridized to the adjunct probe and formed the dual-hairpin structure on the electrode. With the hybridization of adjunct probe and the hairpin-like capture probe in the stem region, the dual-hairpin was formed with outer and inner hairpins. By the conjugation of the target probe with the adjunct probe in the outer hairpin, the adjunct probe divorced from the dual-hairpin structure. The adjunct probe with signal molecules MB, attaching near or divorcing far from the electrode, produced electrochemical signal change and efficient electron transfer due to the fact that it was in proximity to the electrode. However, upon hybridization with the perfect match target, the redox label with the target probe was forced away from the modified electrode, thus resulting in the change of the Dual-Hairpin DNA conformation, which enables impedance of the efficient electron transfer of MB and, consequently, a detectable change of the electrochemical response. In addition, another highlight of this biosensor is its regenerability and stability owing to the merits of structure. Also, based on this Dual-Hairpin platform, the detection limits of DNA, thrombin, and ATP were 50 nM, 3 pM, and 30 nM, respectively. Moreover, this pattern also demonstrated excellent regenerability, reproducibility, and stability. Additionally, given to its ease-of-use, simplicity in design, easy operations, as well as regenerability and stability, the proposed approach may be applied as an excellent design prompter in the preparation of other molecular sensors.

A supersandwich DNA structure was fabricated and used for the amplified detection of Hg2+.

This communication reports on a novel amperometric hydrazine sensor of CuO nanoarray based on a Cu substrate. Copper oxide nanoarray was directly grown on Cu substrates using a one-step facile hydrothermal method and was characterized using scanning electron microscopy and X-ray powder diffraction. The electrochemical study has shown that the CuO nanoarray exhibits higher catalytic effect on the hydrazine than the normal CuO nanoparticles. This may be attributed to the special structure of the nanomaterials esp. the substrate of the electric Cu. And the amperometric response showed that the CuO nanoarray modified glassy carbon electrode has a low detection and a high sensitivity for hydrazine.

A crowded TTE DNA structure for the preparation of Cu NPs with enhanced fluorescence was prepared and applied for the ultrasensitive detection of target DNA.

Herein, we describe a novel homogenous electrochemical aptasensor for the ultrasensitive detection of ractopamine (RAC) based on the signal amplification of a hairpin DNA cascade amplifier (HDCA) and electrocatalysis of dsDNA-templated copper nanoparticles. The present electrochemical aptasensor employs a label-free “turn-on” strategy with enzyme-free amplification. Briefly, the target RAC triggers the HDCA autocatalytic process of two ingeniously designed complementary hairpin DNA, which results in the formation of nucleic-acid-stabilized copper nanoparticles (dsDNA/CuNPs). Thus, with the mimic oxidase catalytic character of the dsDNA/CuNPs and enzyme-free hairpin DNA cascade amplifier reactions, a remarkable electrochemical response can be achieved, which is dependent on the concentration of the target RAC. Taking advantage of the highly amplified efficiency of trigger recycling and the excellent electrochemical catalytic response of dsDNA/CuNPs, the proposed strategy is capable of detecting RAC ultrasensitively. Under optimal conditions, the electrochemical signal increases with an increase in the target RAC concentration in the wide dynamic range from 1 × 10−12 M to 3 × 10−7 M with a detection limit (LOD) of 3 × 10−13 M. The proposed system does not need any sophisticated operation procedures such as electrode immobilization and expensive labeling. Therefore, the devised label-free and enzyme-free amplification homogeneous electrochemical aptasensor strategy may become an alternative method for the simple, sensitive, and selective detection of biological small molecules, proteins, nucleic acids and nuclease activity.

Phosphorylation of DNA by polynucleotide kinase (PNK) plays an important role in a variety of cellular processes, such as nucleic acid metabolism and DNA damage repair. Strategies for a simple and effective PNK assay are in urgent need. Here we report the development of a simple label-free electrochemical method for a PNK assay based on the mimic peroxidase character of a prepared TiO2 nanotube array (NTA) and its specific attachment to phosphorylated DNA. The concentration of T4 PNK was successfully detected by the TiO2 NTA electrode with a detection limit of 0.15 U mL−1. In the process, only a one-step incubation process was used and no labeling processes were involved. In addition, the treated TiO2 foil was anodized into the nanotube array and it was directly used as the electrode without any other complex modification processes which avoided the leakage of an electrode modifier, and so the TiO2 nanotube array with excellent stability and flexibility is worthy of use in real time monitoring.