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Here, an Au–Cu dual-electrode tip was designed to monitor the effect of Cu2+ on the membrane permeability of a single living cell in situ using scanning electrochemical microscopy. The probe approach curves (PACs) were obtained using potassium ferricyanide as a redox mediator. Meanwhile, according to the simulation, theoretical PACs could be acquired. Thus, the cell membrane permeability coefficient (Pm) values were obtained by overlapping the experimental PACs with the theoretical values. Cu2+ was directly generated by electrolyzing the Cu electrode of the dual-electrode tip to investigate its effect on the cell membrane permeability in situ. This work has potential value to improve the understanding of the mechanism of acute heavy metal damage on the cell membrane and will also help clarify the role of heavy metal ions in physiological or pathological processes.

Breast cancer is a heterogeneous disease, and it lacks special tumor markers. Exosomes, new noninvasive biomarkers, with the proteins on the exosome surface show potential for the diagnosis and prognosis of a tumor. However, assessing the variations of exosomal proteins still faces significant challenges. Herein, a magneto-mediated electrochemical sensor based on host-guest recognition has been developed for simultaneous analysis of breast cancer exosomal proteins. Magnetic beads (MB) modified with CD63 aptamer was first employed to capture exosomes. Silica nanoparticles (SiO2 NPs) was modified with MUC1, HER2, EpCAM, and CEA aptamers for specific exosomal proteins identification, respectively, and functionalized with N-(2-((2-aminoethyl)disulfanyl)ethyl) ferrocene carboxamide (FcNHSSNH2) as the signal molecule. The sandwich structure (MB-exosomes-SiO2 NPs probe) was separated by a magnet, and N-(2-mercaptoethyl) ferrocene carboxamide (FcNHSH) was released to the supernatant by the addition of reductants (dithiothreitol, DTT) that break the disulfide bond of FcNHSSNH2. FcNHSH and the graphene oxide-cucurbit [7](GO-CB[7]) modified screen-printed carbon electrode (SPCE) was employed to monitor the oxidation current signals. In this way, four tumor markers on different breast cancer cells (MCF-7, SK-BR-3, MDA-MB-231, and BT474) derived exosomes were sensitively detected. Furthermore, the present assay enabled accurate analysis of exosomes from breast cancer patients, suggesting the potential of exosome analysis in clinic diagnosis.

In this study, a novel all-solid-state ion selective electrode (ASS-ISE) was developed for the detection of Cu2+ by open circuit potential (OCP) based on phytic acid (PA)@Ag, which forms a thin anion layer interacting with positive charged Cu2+. Due to the excellent conductivity and complexing ability with multi-chelating features, PA can selectively recognize metal ions and execute ion-electron conduction. Moreover, with the assistance of Ag nanoparticles, multiple PA molecules were connected, forming a network structure with the improved conductivity and stability. After optimizing the thickness of PA@Ag film, this ASS-ISE gave a near Nernstian behavior (31.3 ± 1.8 mV/dec) towards the variation of Cu2+ concentration in the range of 1.0 × 10−5 to 1.0 × 10−3 mol/L with a detection limit of 2.7 × 10−6 mol/L, and presented excellent selectivity and stability. Furthermore, it was applied in the analysis of Cu2+ in environmental waters with the satisfactory recoveries, and the results were consistent well with those obtained by inductively coupled plasma atomic emission spectrometer, providing a promising alternative for the on-site detection of Cu2+ with simplicity and reliability.

Cigarette smoke contains abundant toxicants, and profiling its cytotoxicity represents a critical topic. In this study, cell-substrate impedance sensing (ECIS) was used to measure the cytotoxicity of 4‑(methylnitrosoamino)‑1‑(3‑pyridinyl)‑1‑butanone (NNK), nicotine, and the total particle material (TPM) of high-tar and low-tar cigarettes on CHO-K1 cells. Normalized impedance values at 3174 Hz were collected with microscopic imaging as an assistant, showing the dynamics of cell damage and the ability for cell self-recovery. The NI determination of the four toxicants indicated that, as the concentration of toxicants increased, cigarette smoke produced more intense toxic effects on the cells, and the ability of cell self-recovery worsened until there was permanent damage to the cells and the cells eventually died. Furthermore, the survival rate of the cells was obtained during treatment. NRU assays as a comparison were developed for evaluating the cytotoxicity by calculating the IC50. Both methods showed that the cytotoxicity decreased in the following order: the TPM of high-tar cigarettes, TPM of low-tar cigarettes, nicotine and NNK. The sensitivity of the ECIS method was higher. Our work provides a useful and convenient approach for determining the cytotoxicity of cigarettes in a real-time, label-free manner, contributing to the development of low-toxicity cigarettes.

Reactive oxygen species (ROS) have gained much attention as they are related to cellular homeostasis and functions. New insights into physiology of cancer cells are anticipated from the analysis of their ROS evolution at single cancer cells level. Herein, a novel scanning electrochemical microscopy (SECM) detection strategy with self-designed multi-potential step waveform (MPSW) has been developed for in situ monitoring ROS with no interference of dissolved oxygen (DO) at the single cells level. Using MPSW, the signals of three different electrochemical molecules – FcMeOH, DO and ROS could be obtained simultaneously in a single imaging process. Images of FcMeOH were used to determine cell morphology. ROS released by normal and cancer cells without the interference of DO were imaged and quantitatively determined using MPSW. ROS released by cancer cells under different pHs was further investigated, suggesting that an acid environment could stimulate the release of ROS. Thus, this work provides a new way for sensing ROS released by cells without interference, and can be applied in the cancer research based on ROS releasing.

In this study, distinguishing skin cancer cells (A431) and normal cells (HaCaT) was achieved using electrical impedance spectroscopy (EIS) with a novel developed device. The proliferation behaviors of the two cell types during a culture period of 5 days were characterized by the normalized impedance measured at 1465 Hz with simultaneous microscopic imaging for assistance. By fitting to the established equivalent circuits, A431 cells generated smaller resistance (Rc) values with smaller increasing variation, and comparable capacitance (Cc) values with similar decreasing variation compared with HaCaT cells. Moreover, Cc values were linearly correlated to the cell number. The results indicate that these two cell types can be distinguished with EIS based on the differences in the values and variation trends of Rc and Cc during the proliferation process in a real-time and label-free manner. Our work supplies a useful analytical approach for skin cancer cell research and may facilitate the early diagnosis of skin cancer.

A novel electrochemical DNA (E-DNA) biosensing strategy was designed and used for the detection of breast cancer susceptibility gene (BRCA-1). The biosensor was based on gold nanoparticles-reduced graphene oxide (AuNPs-GO) modified glass carbon electrode (GCE) covered with the layer of molecularly imprinted polymers (MIPs) synthesized with rhodamine B (RhB) as template, methacrylic acid (MAA) as the monomer, and Nafion as additive. The signal amplification tracing tag SiO2@Ag NPs were prepared by covering AgNPs on the surface of SiO2 nanoparticles in situ, and then DNA probes were modified on AgNPs by Ag-S bond, forming the composites SiO2@Ag/DNA. In presence of target DNA (T-DNA), homogeneous hybridization was performed with SiO2@Ag/DNA and RhB labeled DNA, and the resulting SiO2@Ag/dsDNA/RhB was specifically recognized by MIPs via the interaction between imprinting cavities and RhB. Under optimal conditions, the proposed biosensor exhibited wide linear range from 10 fM to 100 nM, low detection limit of 2.53 fM (S/N = 3), excellent selectivity, reproducibility, stability, and feasibility in serum analysis. Overall, these findings suggest the promising prospects of the proposed biosensing strategy in clinical diagnostics.

Herein we propose a multiple signal amplification strategy designed for ultrasensitive electrochemical detection of glycoproteins. This approach introduces a new type of boronate-affinity sandwich assay (BASA), which was fabricated by using gold nanoparticles combined with reduced graphene oxide (AuNPs-GO) to modify sensing surface for accelerating electron transfer, the composite of molecularly imprinted polymer (MIP) including 4-vinylphenylboronic acid (VPBA) for specific capturing glycoproteins, and SiO2 nanoparticles carried gold nanoparticles (SiO2@Au) labeled with 6-ferrocenylhexanethiol (FcHT) and 4-mercaptophenylboronic acid (MPBA) (SiO2@Au/FcHT/MPBA) as tracing tag for binding glycoprotein and generating electrochemical signal. As a sandwich-type sensing, the SiO2@Au/FcHT/MPBA was captured by glycoprotein on the surface of imprinting film for further electrochemical detection in 0.1 M PBS (pH 7.4). Using horseradish peroxidase (HRP) as a model glycoprotein, the proposed approach exhibited a wide ...

Different cells respond differently to environmental stimulation, resulting in different extracellular pH values. Therefore, the pH of the extracellular microenvironment (pHe) is an important manifestation of cell response to the environment. In this study, a pH-selective dual-microelectrode tip was used as the probe tip of a scanning electrochemical microscopy (SECM) instrument to in situ monitor the pHe changes of different cells after being electrically stimulated by the SECM potential method. pHe is the pH at a certain distance from the cells, and the distance is determined by substituting the current measured by the approach curve into a theoretical calculation. A gold microelectrode was used to determine the distance between the dual-microelectrode tip and the cell, and a pH-selective microelectrode was used to measure the pHe. It was found that the pHe value was related to the distance between the pH-measuring electrode and the cell, and different types of cells had different pHe changes after electrical stimulation. Through fluorescent staining with calcein-AM and propidium iodide, it was verified that this phenomenon was caused by changes in the permeability of the cell membrane due to electrical stimulation. In situ monitoring the pHe changes of different cells will be helpful for exploring the response of cells to environmental stimulation and the mechanism of these different responses.

As a kind of malignant tumor, breast cancer needs more simple and sensitive diagnosis methods. Exosomes, a novel cancer marker, have the potential to be used in cancer biopsy alternative surveillance. Thus, the determination of the correlation between cancer cells and their exosome composition is of significance. Herein, an electrochemiluminescence (ECL) aptasensor has been designed to detect the MUC1 protein expression on breast cancer cells and their derived exosomes. CD63 aptamer was immobilized on the electrode surface to capture the cells and exosomes, and Ru(bpy)32+@SiO2 nanoparticles (Ru@SiO2 NPs) modified with MUC1 aptamer could recognize specifically MUC1 protein on cells and exosomes. The amplified ECL signal from Ru@SiO2 NPs exhibited linear responses in the range of 7.53 - 753 μg/mL with a limit of detection (LOD) of 0.83 μg/mL for MCF-7 cells, as well as linear range of 3.22 × 10−4 - 156 μg/mL with a LOD of 2.73 × 10−4 μg/mL for their exosomes. The expression of MUC1 protein on the surface of MDA-MB-231 and SK-BR-3 cells and their exosomes, were further analyzed, indicating the consistency of MUC1 expression of breast cancer cells and their exosomes. Furthermore, this aptasensor was applied to the determination of MUC1 protein on exosomes in serum samples of patient and normal control, demonstrating its potential for clinical cancer surveillance.

Recently, many strategies have been developed to trace the breast cancer susceptibility gene (BRCA), which is closely related to the occurrence of breast cancer. In this work, a novel recyclable electrochemical sensing platform was constructed to detect BRCA DNA based on the homogeneous hybridization of this target sequence with ferrocene-labeled DNA/Au nanospheres (FcNS) and horseradish peroxidase-labeled DNA/Au nanospheres (HRPNS) concatamers, and the host-guest interaction between cucurbit [7]uril (CB [7]) adsorbed on the electrode and Fc on the hybridization complex. With the optimization of experimental conditions, the fabricated sensing platform showed improved sensitivity with a low detection limit of 25 pM (S/N = 3), contributed by the dual amplification of signal using FcNS and HRPNS concatamers, and high specificity for BRCA DNA. The captured complex was dissociated from CB [7]-modified electrode with the rise of pH value to recycle the sensing platform. Furthermore, this detection strategy displayed the reliability and applicability in the analysis of human serum samples, indicating great potential for applications in early diagnosis of breast cancer.

Due to their unique physical and chemical properties, 2D materials have attracted intense interest for applications in filtration, sensing, nanoelectronics, and biomedical devices. Peptoids are a class of biomimetic sequence-defined polymers for which certain amphiphillic sequences self-assemble into 2D crystalline materials with properties that mimic those of cell membranes. In this study the ability of these membrane-like materials to self-repair following damage on a range of substrates is explored. In situ atomic force microscopy (AFM) is used to both create damage and image the subsequent repair process. Damage is induced by using the AFM to scribe peptoid-free patterns within a preassembled membrane. The results show here that, upon introduction of a peptoid-containing solution, for a suitable range of pH conditions, the damage is eliminated through assembly of the peptoids at the newly created edges, regardless of whether the substrates are negatively or positively charged and even in the absence of an underlying surface. The rate of the advancing edge depends on the edge orientation, the pH, and the composition of the substrate. Moreover, if the solution contains a second peptoid having an identical sequence in the hydrophobic block, repair of the defects results in nanoscale patterns of the new peptoid, even if the hydrophilic regions are distinct. Consequently, this ability to self-repair can be exploited to create nm-sized patterns of distinct functional groups within a single coherent membrane.

In this work, a recyclable electrochemical sensing platform to detect breast cancer susceptibility gene (BRCA) was constructed by pillararenes based host–guest recognition and homogeneous DNA hybridization. BRCA target DNA (T-DNA) formed sandwich-type DNA via homogeneous hybridization with methylene blue labeled signal DNA and alkylamino modified capture DNA, which could form complexes with pillararenes. Such sandwich-type DNA was captured by a novel trithiocarbonate modified pillar[5]arene (P5A-CTA) which was immobilized on the Au electrode. With the help of enzyme amplification, the electrochemical detection signal of target DNA in sensing platform was apparently amplified. This sensing platform exhibited wide linear range and excellent specificity, and was particularly recyclable after simple washing with hot acetonitrile due to the reversible host–guest complexation between P5A-CTA and alkylamino modified DNA.

A novel label-free electrochemical strategy was designed for multi-sites recognizing the guanine-rich DNA (G-rich DNA) with surface molecular imprinting polymer composite, in which the multi-walled carbon nanotubes (MWNCTs) were regarded as supporting material and the molecularly imprinted polymer (MIP) with guanine sites of DNA was applied as recognition element. Firstly, the vinyl groups were grafted onto the surface of carboxylic acid-functionalized MWNCTs (MWCNTs-COOH) through chemical modification. Then, the composite of MWCNTs-MIP was fabricated by the selective copolymerization of methacrylic acid, ethylene glycol dimethacrylate and guanine on the vinyl group-functionalized MWNTs surface. MWCNTs-MIP was characterized by different techniques, including Fourier transform infrared (FTIR) spectroscopy, scanning electronic microscopy (SEM), cyclic voltammogram (CV) and electrochemical impedance spectroscopy (EIS), showing that the MWCNTs-MIP composite was successfully prepared. Under the optimized conditions, the imprinting factor was nearly 5.68 according to the comparative slopes obtained on MWCNT-MIP and MWCNTs-non-imprinted polymer (MWCNTs-NIP), indicating that MWCNTs-MIP exhibited high selectivity for G-rich DNA. Moreover, the MWCNTs-MIP composite had a relatively wide linear range over G-rich DNA concentration of 0.05–1 μM and 5–30 μM with a detection limit of 7.52 nM (S/N = 3). Furthermore, the novel electrochemical strategy based on imprinted composite has very excellent performance in real samples of human serum and human urine.

The long DNA concatemers have been well developed to fabricate various biosensing platforms for the signal amplification. Herein, this signal amplification strategy was firstly used for an ultrasensitive scanning electrochemical microscopy (SECM)-based DNA biosensing platform. This platform was constructed through the hybridization of target DNA (TD) with thiol-tethered DNA capture probes (CP), immobilized on the gold substrate surface, and biotinylated DNA signal probes (SP), which formed then the long DNA concatemers through the continuous self-assembly with alternating DNA auxiliary probes (AP). The streptavidin-horseradish peroxidase (HRP) was linked to the long DNA concatemers through biotin-streptavidin interaction. In the HRP-catalyzed reaction, hydroquinone (H 2 Q) was oxidized to benzoquinone (BQ) with H 2 O 2 at the modified substrate surface where sequence-specific hybridization had occurred, and the BQ generated could be monitored by a SECM tip. This platform exhibited a low detection limit of 0.18 aM estimated by the 3σ rule. Combined with DNA microarrays, four kinds of TDs (100 fM) as the models were detected simultaneously by using this proposed strategy, which also demonstrated sufficient selectivity to distinguish specific DNA sequences and good reproducibility. This method opens a promising direction to improve the SECM sensitivity for high-throughput DNA detection.

A novel Congo red (CR) derivatized silica stationary phase was prepared and packed into a fused silica capillary tube for nano-flow HPLC. A variety of analytes including poly-aromatic hydrocarbons, parabens, acids, sulfonamides, bases, and nucleosides were successfully separated using the CR. In comparison with commercial ODS columns, this new stationary phase has a different separation mechanism (hydrophobically-assisted ion-exchange), which was evident in the separation of benzoic acid derivatives and sulfonamides. The successful application of CR-bonded silica stationary phase in the HILIC and PALC modes demonstrates the effectiveness of this potential chromatographic material in nano flow HPLC.

Amyloid-β oligomers (AβOs) are highly toxic species involved in Alzheimer's disease (AD). Hence, a reliable detection method for AβOs, which are promising potential therapeutic targets and biomarkers for AD, is of great significance for improving the diagnosis of AD. Herein, a novel sandwich assay electrochemical biosensor was developed for highly sensitive and selective detection of AβO, using molecularly imprinted polymers (MIPs) and aptamer as the recognition element. Instead of using an antibody to recognize the AβO target molecules, the AβO in the samples were captured by the film of MIPs and the AβO-specific aptamer, forming a MIPs/target/aptamer sandwich assay for the highly selective detection of AβO. The AβO-specific aptamer was immobilized on the surface of core-shell nanoparticles that combined silver nanoparticles with silica nanoparticles (SiO2@Ag NPs). The highly sensitive electrochemical signal from the sandwich assay was generated by using a small amount of AβO to trigger a large number of electrochemically active Ag NPs. Under the optimized conditions, the developed biosensor showed good linearity in the concentration range of 5 pg mL−1 to 10 ng mL−1 with a limit of detection of 1.22 pg mL−1. The biosensor also showed excellent specificity, reproducibility and stability. In addition, the feasibility of detecting AβO in human serum was successfully verified, demonstrating the promising potential of this approach for clinical research and the early diagnosis of AD.

Exosomes (30–150 nm) secreted by cells play an important role in intercellular communication. Herein, a novel double imprinting-based electrochemical method has been developed for analyzing the particle size distribution (PSD) of mimetic exosomes – a mixture of SiO2@HRP with the diameters in 50 nm, 100 nm and 150 nm at a ratio similar to the exosomes. Double imprinting polymers (DIP) were synthesized with SiO2@HRP at different sizes as template, providing the both recognition of size and morphology. With the capture of the target and the binding of the signal amplification tag – SiO2@Ag/MPBA on the DIP film, a sandwich structure was constructed on Au NPs-GO modified glassy carbon electrode. Under the optimal conditions, the DIP film exhibits the linear correlations between the current intensity and the logarithmic concentration of SiO2@HRP as ΔI50 = −1.52 + 0.50 × lgc50 (2.89 × 104–2.89 × 109 particles/mL), ΔI100 = −1.61 + 0.48 × lgc100 (2.89 × 104–2.89 × 109 particles/mL) and ΔI150 = −1.54 + 0.49 × lgc150 (5.75 × 104–5.75 × 109 particles/mL) with the limit of detection of 1.44 × 103 particles/mL, 5.68 × 102 particles/mL and 7.70 × 102 particles/mL, respectively. Furthermore, the DIP film was applied to the PSD analysis of mimetic exosomes – SiO2@HRP at mixed sizes with a ratio of 50 nm:100 nm:150 nm = 5.0%:42.5%:52.5% detected by nanoparticle tracking analysis (NTA), obtaining a similar result of 50 nm:100 nm:150 nm = 3.6%:43.0%:53.4% with small relative concentration errors. It also exhibited excellent reproducibility and stability. This double imprinting-based method shows high potential in the separation and detection of complex biosamples.

Herein, a novel and convenient electrochemically active-inactive switching molecular beacon based on hemin (Hs-MB) has been designed for easy discrimination of single nucleotide polymorphisms (SNPs) and sensitive detection of insulin. The electrochemically active changing capability of Hs-MB is based on two identical hemin groups labeled at both ends of MB sequence in dimer or monomer forms depending on the conformation of MB which is in stem-loop structure or line shaped structure. The Hs-MB assay permits discrimination of SNPs and the highly sensitive and specific detection of insulin with detection limit successively as low as 0.5 pmol/L. Even at a very low target concentration, the Hs-MB assay also shows a good specificity in the presence of other potentially interfering components. The experimental results also show that Hs-MB can also be used for the accurate and rapid monitoring of insulin secretion by glucose-stimulated from MIN6 cells at different time periods, demonstrating that Hs-MB has potential in monitoring of biomarker variation in vivo.

A simple and sensitive method for glucose determination was described based on the host-guest inclusive complex of cucurbit[7]uril and ferrocenemonocarboxylic acid. In this work, the mixture of cucurbit[7]uril and graphene oxide, ferrocenemonocarboxylic acid as electron transfer mediator and glucose oxidase as catalytic material were modified successively on the electrode. Under the optimized conditions, a linear response between current intensity and glucose concentrations over a range of 0.1–10 mM was obtained with a relatively low detection limit (27 µM, S/N=3). Meanwhile, this method exhibited high selectivity in the interference investigation, and the reliability and applicability in human serum samples analysis.

Here, polydopamine derivative monolayer was generated by electro-polymerization of thiol dopamine derivatives modified on gold electrode, and the adhesion of polydopamine made it being a platform for secondary reaction. The influence of pH on formation of polydopamine derivative was investigated. Horseradish peroxidase (HRP) was adhered on the polydopamine derivative monolayer electrode catalyzing the oxidation of H2O2. Cyclic voltammetry (CV) was used to detect the polydopamine monolayer as well as electrochemical catalysis of H2O2. Besides, electrochemical impedance spectroscopy (EIS) and atomic force microscope (AFM) were also employed to characterize the polymeric monolayer.

Exosomes, lipid bilayer membrane vesicles, can guide various pathological and physiological processes. However, reliable, convenient and sensitive methods for exosome determination for early cancer diagnosis are still technically challenging. Herein, an electrochemical aptasensor based on click chemistry and the DNA hybridization chain reaction (HCR) for signal amplification has been developed for the ultrasensitive detection of tumor exosomes. CD63 aptamer was first immobilized on a glassy carbon electrode for capturing exosomes, and 4-oxo-2-nonenal alkyne (alkynyl-4-ONE) molecules, functionalized lipid electrophiles, were conjugated to the exosomes via the reaction of amino and aldehyde groups. Azide-labeled DNA probe as an anchor was then connected to the exosomes by copper (I)-catalyzed click chemistry. Signal amplification was achieved by HCR, and the numerous linked horseradish peroxidase (HRP) molecules could catalyze the reaction of o-phenylenediamine (OPD) and H2O2. The concentration of exosomes could be quantified by monitoring the electrochemical reduction current of 2,3-diaminophenazine (DAP). Under the optimal conditions, this method allowed the sensitive detection of exosomes in the range of 1.12 × 102 to 1.12 × 108 particles/μL with a limit of detection (LOD) of 96 particles/μL. Furthermore, the present assay enabled sensitive and accurate quantification of exosomes in human serum, and it has high potential for exosome analysis in clinical samples.

In this study, a novel and ultrasensitive biosensor strategy based on gold nanoparticles and aptamer segments was designed to detect two biomolecule targets: DNA and thrombin. Two DNA probes were designed that included aptamer segments and nucleic acids. The aptamer segments assembled into a G-quadruplex structure in the presence of thrombin, and the nucleic acids formed a double helix structure with the target DNA. The two gold nanoparticle-labeled DNA probes were effectively aggregated by specific recognition of the target molecules. Methylene blue, a redox-active indicator of single-stranded DNA, was incorporated in the unrecognized segment of the probe on the gold nanoparticle aggregation. Furthermore, the gold nanoparticle aggregation not only provided an excellent conductive matrix for methylene blue, but also abundantly enriched the methylene blue molecule. Using this method, DNA and thrombin were detected in low concentration ranges with both probes. These gold nanoparticle aggregation-based elect...

Qualitative and quantitative detection of DNA was achieved by a "sandwich" DNA sensor through SG/TC (substrate generation and tip collection) mode of scanning electrochemical microscopy (SECM). The "sandwich" DNA structure was formed by the hybridization of thiol-tethered oligodeoxynucleotide probes (capture probe), assembled on the gold substrate surface, with target DNA and biotinylated indicator probe. HRP (horseradish peroxidase)-wrapped SiO2 nanoparticles were linked to the sandwich structure through biotin-streptavidin interaction. Hydroquinone (H2Q) was oxidized to benzoquinone (BQ) at the modified substrate surface where sequence-specific hybridization had occurred through the HRP-catalyzed reaction in the presence of H2O2. The detection was based on the reduction of BQ generated on the modified substrate by SECM tip. For SECM imaging experiment, we structured the microsensor platform through localized desorption of 1-dodecanethiol monolayer. Approach curves were employed for quantitative detection of DNA concentration. The detection limit of complementary DNA was as low as 0.8pM. This technique is promising for the application on electrochemical DNA chip.

A novel magnetic ordered mesoporous carbon composite was prepared. Electrochemical measurements showed that the ordered mesoporous carbon composite provided an excellent matrix for the co-adsorption of horseradish peroxidase (HRP). HRP could be separated and collected by the application of a magnetic field and its direct electron-transfer could be achieved in the solution, not on the electrode thereby preventing the degradation of the enzyme. The cyclic voltammetric experimental results of HRP indicated that HRP displayed a pair of stable peaks with a formal potential of −0.306 V in PBS. The resulting biosensor exhibited fast amperometric response to hydrogen peroxide.

A palladium nanoparticle/graphene/chitosan/glassy carbon electrode (PdNPs/GR/CS GCE) was prepared and characterized in this paper. This chemical modified electrode displayed excellent eletrochemical catalytic activities towards ascorbic acid (AA), dopamine (DA) and uric acid (UA) compared with bare GCE, PdNPs/GCE and GR/CS GCE by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). All oxidation potentials of AA, DA and UA at the PdNPs/GR/CS GCE shifted negatively and the peak currents were much larger than other electrodes. DPV was used for simultaneous determination of AA, DA and UA in their mixture, and the peak potential separations between AA and DA, DA and UA, AA and UA were 252 mV, 144 mV and 396 mV, respectively. The limits of detection ( S / N = 3) for AA, DA and UA were 20 μM, 0.1 μM and 0.17 μM, respectively. This PdNPs/GR/CS GCE showed many merits in stability, sensitivity, facility and economy.

In this work, a simple and sensitive electrochemical method was developed to determine Sudan I by cyclic voltammetry and differential pulse voltammetry using a glassy carbon electrode modified with a chitosan/carbon nanotube composite. In cyclic voltammetry, Sudan I exhibited a well-defined oxidation peak located at 0.72 V at the multi-walled carbon nanotube (MWCNT)/chitosan-modified GCE. The determination conditions, including pH, scan rate, and chitosan: MWCNT mass ratio at the modified electrode, were optimized. Under the optimum experimental conditions, Sudan I could be linearly detected by differential pulse voltammetry with a detection limit of 3.0 × 10-8 mol?L-1.

A sensitive electrochemical aptasensor for thrombin detection is presented based on the host-guest recognition technique. In this sensing protocol, a 15 based thrombin aptamer (ab. TBA) was dually labeled with a thiol at its 3′ end and a 4-((4-(dimethylamino)phenyl)azo) benzoic acid (dabcyl) at its 5′ end, respectively, which was previously immobilized on one Au electrode surface by Au S bond and used as the thrombin probe during the protein sensing procedure. One special electrochemical marker was prepared by modifying CdS nanoparticle with β-cyclodextrins (ab. CdS-CDs), which employed as electrochemical signal provider and would conjunct with the thrombin probe modified electrode through the host–guest recognition of CDs to dabcyl. In the absence of thrombin, the probe adopted linear structure to conjunct with CdS-CDs. In present of thrombin, the TBA bond with thrombin and transformed into its special G-quarter structure, which forced CdS-CDs into the solution. Therefore, the target-TBA binding event can be sensitively transduced via detecting the electrochemical oxidation current signal of Cd of CdS nanoparticles in the solution. Using this method, as low as 4.6 pM thrombin had been detected.

The electrochemical monitoring of glucose consumption is relevant for cell biology studies because of its wide detection range, high sensitivity and easy implementation. Whereas the glucose consumption and cell growth rate can be tightly correlated, they should also be cell population density dependent. In this work, we fabricated high sensitive enzyme electrodes for accurate monitoring of glucose consumption of cells in different growth stages. The performance of the fabricated device was firstly evaluated by cyclic voltammetry (CV) with p-benzoquinone (PBQ) as redox mediator, showing a linear response over a wide detection range (0.3–60 mM), a high sensitivity (1.61 ± 0.10 μA mM−1 mm−2 (n = 5)) and a low detection limit (80 μM). Then, daily glucose consumptions of NIH 3T3 cells in 24-well plates were determined for a period of 7 days. The results could be compared to the cell population growth curve, showing a close correlation but different behavior. We found that the increase of the glucose consumption took place prior the cell number increase but the glucose consumption per cell decreases linearly in the exponential growth stage of cells.

A label-free ECL aptamer biosensor based on CdS QDs/ACNTs electrode for the sensitive detection of thrombin was fabricated. CdS QDs and Chitosan (CTS) complex films were coated and filled the tubes of ACNTs by carrying out electrodeposition reaction of CTS-CdS QDs on ACNTs electrode, the resulting electrode showed high ECL intensity and good biocompatibility. After aptamer was bound to the film via glutaric dialdehyde, the modified electrode could be used as an ECL aptamer sensor for the thrombin detection. The specific reaction between thrombin and aptamer resulted in the decrease in ECL intensity. The change of the ECL intensity was found to be linear with the logarithm of thrombin concentration in the range from 1.0×10−13 M to 1.0×10−9 M. The ECL aptamer sensor had the advantages of speed, high sensitivity, specificity and stability, which could be a promising technique for protein detection.

A novel G-quadruplex-based DNAzymes aptasensor for the amplified electrochemical detection of thrombin has been described. The aptasensor utilized a combination of hemin and guanine-rich thrombin-binding aptamer (TBA) to form horseradish peroxidase (HRP)-mimicking DNAzymes with peroxidase catalytic activity. In the presence of thrombin, the enzyme activity could be extensively promoted, thereby providing the amplified electrochemical readout signals for detecting thrombin. This aptasensor exhibited high sensitivity and selectivity for thrombin determination, which enabled the analysis of thrombin with a detection limit of 6×10–11 M. On the basis of results, this method could have broad applications in the detection of proteins and other biomolecules.

A new electrochemical aptamer molecular beacon (MB) was designed by the carminic acid (CA) covalently linking at the each end of a special single-stranded stem-loop shaped oligonucleotide and named as CAs-MB. CA is an electrochemically active molecule and two CA molecules at the ends of molecular beacon stem were closed enough to associate each other to be as CA dimer. The dimer was electrochemically inactive. It separated into two CA monomers and produced the electrochemical signal while CAs-MB combined with target. In this protocol, the detection strategy of CAs-MB for thrombin is based on electrochemical active–inactive switching between monomer and dimer forms of CA. In order to enhance the electrochemical signal, magnetic nanobeads (MNB) was applied by connecting CAs-MB with MNB through a duplex of DNA. With the magnetic enrichment, the detection limit for thrombin reached to 42.4 pM. The experiment results showed that this type of electrochemical active–inactive switching aptamer molecular beacon allowed the direct detection of target proteins in the solution with no requirement of removing uncombined CAs-MB. Besides, CAs-MB/MNB can be easily regenerated by using 2 M NaCl solution to cleave the thrombin from the aptasensor.

We herein constructed a sensor that converts target DNA hybridization-induced conformational transformation of the probe DNA to electrochemical response based on host-guest recognition and nanoparticle label. In the sensor, the hairpin DNA terminal-labeled with 4-((4-(dimethylamino)phenyl)azo)benzoic acid (dabcyl) and thiol group was immobilized on Au electrode surface as the probe DNA by Au-S bond, and the CdS nanoparticles surface-modified with β-cyclodextrins (CdS-CDs) were employed as electrochemical signal provider and host-guest recognition element. Initially, the probe DNA immobilized on electrode kept the stem-loop configuration, which shielded dabcyl from docking with the CdS-CDs in solution due to the steric effect. After target hybridization, the probe DNA underwent a significant conformational change, which forced dabcyl away from the electrode. As a result, formerly-shielded dabcyl became accessible to host-guest recognition between β-cyclodextrin (β-CD) and dabcyl, thus the target hybridization event could be sensitively transduced to electrochemical signal provided by CdS-CDs. This host-guest recognition-based electrochemical sensor has been able to detect as low as picomolar DNA target with excellent differentiation ability for even single mismatch.

This communication reports on a new electrochemical method to detect the hybridization specificity by using host–guest recognition technique. A hairpin DNA with dabcyl-labeled at its 3′ and NH2 group at 5′ terminal was combined with CdS nanoparticle to construct a double-labeled probe (DLP), which could selectively hybridize with its target DNA in homogeneous solution. A β-CD modified Poly(N-acetylaniline) glassy carbon electrode was used for capturing the dabcyl label in DLP. When without binding with target DNA, the DLP kept its stem-loop structure which shielded the dabcyl molecule due to the loop of the hairpin DNA and CdS nanoparticle blocking dabcyl enter into the cavity of these β-CD molecules on the electrode. However, in present of complementary sequence, the target-binding DLP was incorporated into double stranded DNA, causing the DLP’s loop-stem structure opened and then the dabcyl was easily captured by the β-CD modified electrode. During electrochemical measurement, the signal from the dissolved Cd2+ was used for target DNA quantitative analysis. Keywords: DNA, Electrochemical, Homogeneous, Cyclodextrins, Double-labeled probe

With a view of simultaneous determination of physio-active ingredients in oolong tea infusion: sugars, amino acids, epigallocatechin gallate and ascorbic acid, a novel CZE with amperometric detection method was studied. Operated in a wall-jet configuration, 100 mmol/L NaOH was used in detecting cell to lead the electrocatalysis oxidation behaviors of the analytes on a 300 mum diameter copper-disc electrode (working electrode), while in separating capillary, a mild alkaline running buffer consisting in a mixture of 30 mmol/L borate and 40 mmol/L phosphates charged and carried analytes to detecting end. The methodology research was performed for system stability and suitability. Under the optimal CE conditions, analytes could be separated within moderate time period. Good linearity between peak area and concentration existed over three orders of magnitude; lower RSD and LOD were achieved. The oolong tea infusion was assayed and result was satisfactory.

Polythymine oligonucleotide (PTO)-modified gold electrode (PTO/Au) was developed for selective and sensitive Hg2+ detection in aqueous solutions. This modified electrode was prepared by self-assembly of thiolated polythymine oligonucleotide (5′-SH-T15-3′) on the gold electrode via AuS bonds, and then the surface was passivated with 1-mercaptohexanol solution. The proposed electrode utilizes the specific binding interactions between Hg2+ and thymine to selectively capture Hg2+, thereby reducing the interference from coexistent ions. After exchanging the medium, electrochemical reduction at −0.2 V for 60 s, voltammetric determination was performed by differential pulse voltammetry using 10 mM HEPES; pH 7.2, 1 M NaClO4 as supporting electrolyte. This electrode showed increasing voltammetric response in the range of 0.21 nM Hg2+, with a relative standard deviation of 5.32% and a practical detection limit of 60 pM. Compared with the conventional stripping approach, the modified electrode exhibits good sensitivity and selectivity, and is expected to be a new type of green electrode.

In the paper, a new kind of vitamin B 12 (acquo-cobalamine) chemically modified electrode was fabricated and applied in capillary zone electrophoresis coupled with amperometric detection (CZE-AD) for simultaneous determination of six antioxidants in fruits and vegetables. The catalytic electrochemical properties of the chemically modified electrode could obviously enhance oxidation peak heights responses by about five times to glutathione, ascorbic acid, vanillic acid, chlorogenic acid, salicylic acid, and caffeic acid compared with common carbon disk electrode. Furthermore, the effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CZE-AD were investigated. Under the optimum conditions, the six analytes could be completely separated and detected in a borate–phosphate buffer (pH 8.4) within 15 min. Their linear ranges were from 2.5 × 10 −7 to 1.0 × 10 −4 mol L −1 and the detection limits were as low as 10 −8 mol L −1 magnitude ( S / N = 3). The proposed method has been successfully employed to monitor the six analytes in practical samples with recoveries in the range 96.0–106.0% and RSDs less than 5.0%. Above results demonstrate that capillary zone electrophoresis coupled with electrochemical detection using vitamin B 12 modified electrode as detector is of convenient preparation, high sensitivity, good repeatability, and could be used in the rapid determination of practical samples.

In general capillary zone electrophoresis (CZE) separation models, o-, m-, and p-phenylenediamine isomers can be separated in a weak acidic running buffer for their pK(a) values being 4.52, 5.64, 6.04, respectively, while o-, m-, and p-dihydroxybenzene isomers can be separated in a weak basic buffer for their pK(a) values being 9.40, 9.40 and 10.04, respectively. So, it is hard to find a suitable running buffer at a fixed pH in normal CZE for simultaneous separation of these two groups of positional isomers. In this paper, a novel method based on alternately running two different pH buffers in CZE coupled with amperometric detection (CZE-AD) was designed to simultaneously determine these two groups of positional isomers. It is found that when two different pH running buffers were employed alternately under appropriate order and time, the six analytes could be separated perfectly in less than 20 min and the detection limits were as low as 10(-7) mol/L. Furthermore, the effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CZE-AD were investigated. Experimental results demonstrated that the introduced method was practical to simultaneously determine two groups of positional isomers with different pK(a) and had some advantages of high sensitivity, good repeatability and small sample requirement.

A simple and efficient electrochemical method is utilized to functionalize aligned carbon nanotubes (ACNTs) by the electrochemical reduction of 4-carboxyphenyl diazonium salt. Thus hemoglobin (Hb) molecules were covalently immobilized on the diazonium-ACNTs surface via carbodiimide chemistry. Direct electrochemistry and bioelectrocatalytic activity of the immobilized Hb were then investigated by cyclic voltammetry (CV) and amperometry techniques. It is showed that the Hb film on the diazonium-ACNTs electrode had well-defined redox peaks with a formal potential (E°) at −312 mV (vs. Ag/AgCl), and the Hb-ACNTs electrode displayed good electrocatalytic activity to H2O2 reduction. Owing to the high Hb covering on the ACNTs surface (Γ*=2.7×10−9 mol cm−2), the catalytic current were significantly improved when compared to the current measured at an Hb-tangled carbon nanotubes electrode. The Hb-ACNTs electrode exhibited high sensitivity, long-term stability and wide concentration range from 40 μM to 3 mM for the amperometric detection of H2O2. The heterogeneous reaction rate constant (ks) was 0.95±0.05 s−1 and the apparent Michaelis–Menten constant (K was 0.15 mM.

A solid-state electrochemiluminescence (ECL) biosensing switch system based on special ferrocene-labeled molecular beacon aptamer (Fc-MBA) has been developed successfully for thrombin detections. Such special switch system includes two main parts, an ECL substrate and an ECL intensity switch. The ECL substrate was made by modifying the complex of Au nanoparticle and Ruthenium (II) tris-(bipyridine) (Ru(bpy)(3)(2+)-AuNPs) onto Au electrode. A molecular beacon aptamer labeled by ferrocene acted as the ECL intensity switch. The loop bases of the ECL intensity switch are designed with special anti-thrombin aptamer sequence which could be combined with its target protein via the reaction between aptamer and thrombin. During the reactions, the molecular beacon aptamer opened its stem-loop, and the labeled Fc was consequently kept away from the ECL substrate. Such structural change resulted in an obvious ECL intensity increment due to the decreased quenching effect of Fc to the ECL substrate. The analytic results are sensitive and specific.

The combinatorial chemical synthesis offers in vitro aptamer generation with limitless aptamer selection for various analytes. Additionally, aptamer has quite many outstanding properties as the biomolecular recognition elements, thus competing with traditional affinity ligands (enzyme, antibodies, lectins etc.) in biological and medicine employments. This article reviews the advancements in electrochemical aptasensors for biomolecule monitoring, emphasizing the efficient detection strategies of sandwich structure formation using two recognition elements to capture and label target respectively, EIS measuring the electronic transfer resistance change, the aptamer conformation change triggering the electrochemical signal on/off and target molecules displacing electroactive markers.

A versatile method for preparing nanoporous polypyrrole (PPy) film using electrogenerated nanobubbles as templates on highly oriented pyrolytic graphite (HOPG) is presented using in situ electrochemical atomic force microscopy (EC-AFM). Keywords: Polypyrrole, Nanobubbles, Highly oriented pyrolytic graphite, Electrochemical atomic force microscopy

Tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy) 3 2+ ) has been successfully immobilized onto electrode through the electrodeposition of Ru(bpy) 3 2+ /AuNPs/chitosan composite film. In the experiments, chitosan solution was first mixed with Au nanoparticles (AuNPs) and Ru(bpy) 3 2+ . Then, during chronopotentiometry experiments in this mixed solution, a porous 3D network structured film containing Ru(bpy) 3 2+ , AuNPs and chitosan has been electrodeposited onto cathode due to the deposition of chitosan when pH value is over its p K a (6.3). The applied current density is crucial to the film thickness and the amount of the entrapped Ru(bpy) 3 2+ . Additionally, these doping Ru(bpy) 3 2+ in the composite film maintained their intrinsic electrochemical and electrochemiluminescence activities. Consequently, this Ru(bpy) 3 2+ /AuNPs/chitosan modified electrode has been used in ECL to detect tripropylamine, and the detection limit was 5 × 10 −10 M.

We report a new kind of electrochemical molecular beacon, termed “electrochemically active–inactive switching molecular beacon”, for direct detection of DNA in homogenous solution. The electrochemical molecular beacon consists of a stable stem-loop oligonucleotide carrying two carminic acid moieties (acting as electrochemical reporter) attached at its termini. In a close form, the electrochemical signal is quenched because two carminic acid moieties are close enough to associate into dimer. In the presence of the complementary DNA target, the electrochemical molecular beacon undergoes a conformational transformation from closed (hairpin) to open (linear) structure, which is associated with an increase in electrochemical signal. We found that the electrochemical molecular beacon is as effective as conventional molecular beacon in signaling the presence of complementary target and discriminating targets that differ by a single nucleotide. The proposed electrochemical molecular beacon has a great potential for investigating the interactions of DNA-protein and developing electrochemical real-time polymerase chain reaction. Keywords: DNA, Electrochemical molecular beacon, Molecular recognition, Nucleotides, Probe

Aligned carbon nanotubes (ACNTs) electrode has been developed for the direct protein electrochemistry and enzyme-biosensor study involving two types of nanoparticles. Pt nanoparticles (Ptnano) were electro-modified on the ACNTs’ each tube, greatly increasing the electrode surface area for locating protein and also its electronic transfer ability. Glucose oxidase (GOD) with chitosan (CS) and CdS nanoparticles electrochemically coated on each tube of ACNTs–Ptnano by the electrodeposition reaction of CS when pH value passing its pKa. The CdS nanoparticles between ACNTs electrode and GOD have stimulated the GOD’s direct electron transfer during its redox reaction of FAD/FADH2. The CS–GOD–CdS/ACNTs–Ptnano electrode also offer sensitive response to the substrate of glucose with detection limit of 46.8 μM (S/N = 3) and apparent Michaelis–Menten constant of 11.86 mM. Keywords: Aligned carbon nanotubes, Glucose oxidase, Chitosan, Direct electron transfer, Biosensor

A sensitive electrochemical aptasensor for detection of thrombin based on target protein-induced strand displacement is presented. For this proposed aptasensor, dsDNA which was prepared by the hybridization reaction of the immobilized probe ssDNA (IP) containing thiol group and thrombin aptamer base sequence was initially immobilized on the Au electrode by self-assembling via AuS bind, and a single DNA labeled with CdS nanoparticles (DP-CdS) was used as a detection probe. When the so prepared dsDNA modified Au electrode was immersed into a solution containing target protein and DP-CdS, the aptamer in the dsDNA preferred to form G-quarter structure with the present target protein resulting that the dsDNA sequence released one single strand and returned to IP strand which consequently hybridized with DP-CdS. After dissolving the captured CdS particles from the electrode, a mercury-film electrode was used for electrochemical detection of these Cd2+ ions which offered sensitive electrochemical signal transduction. The peak current of Cd2+ ions had a good linear relationship with the thrombin concentration in the range of 2.3×10−9–2.3×10−12 mol/L and the detection limit was 4.3×10−13 mol/L of thrombin. The detection was also specific for thrombin without being affected by the coexistence of other proteins, such as BSA and lysozyme.

A carbon paste electrode modified with multiwall carbon nanotubes and copper(I) oxide (MWCNT–Cu2O CPME) was fabricated, and the electrochemical behaviors of 19 kinds of natural amino acids at this modified electrode were studied. The experimental results showed that the various kinds of amino acids without any derivatization displayed obvious oxidation current responses at the modified electrode. It was also found that the current response values of amino acids were dependent mainly on pH values of buffer solutions. The phenomenon could be explained by the fact that the amino acids suffered complexation or electrocatalytic oxidation processes under different pH values. Six kinds of amino acids (arginine, tryptophan, histidine, threonine, serine, and tyrosine), which performed high-oxidation current responses in alkaline buffers, were selected to be detected simultaneously by capillary zone electrophoresis coupled with amperometric detection (CZE–AD). These amino acids could be perfectly separated within 20 min, and their detection limits were as low as 10−7 or 10−8 mol L−1 magnitude (signal/noise ratio = 3). The above results demonstrated that MWCNT–Cu2O CPME could be successfully employed as an electrochemical sensor for amino acids with some advantages of convenient preparation, high sensitivity, and good repeatability.

A biocompatible nanocomposite film was fabricated for hemoglobin (Hb) molecules immobilization. This film consists of multiwalled carbon nanotubes (MWNTs), 1-pyrenebutanoic acid, succinimidyl ester (PASE), hemoglobin (Hb) and Au nanoparticles (AuNPs). Herein, PASE molecules physically adsorbed onto MWNTs, and its groups then covalently bond with Hb. AuNPs were then linked with Hb/PASE/MWNTs via electrostatic adsorption force. UV-visible adsorption spectra, Fourier transform infrared spectra, scanning electron microscope and electrochemical impedance spectroscopy have characterized the film. Cyclic voltammetry (CV) scans showed that in the film Hb has well-defined redox reaction, with the formal potential (E°) at about −0.27 V (vs. Ag/AgCl). Herein, differential pulse voltammetry (DPV) was employed to electrochemically detect the Hb in the film with a detection limit of 9.3×10−8 M. The proposed method was also succeeded for Hb detection in clinical blood samples. Furthermore, the Hb in the film showed good electrocatalytic activities to the reduction of H2O2, TCA, NaNO2 and O2.

A novel Au nanoparticles (Au-NPs)-based protocol for DNA hybridization detection based on assembly of alternating DNA and poly(dimethyldiallylammonium chloride) (PDDA) multilayer films by layer-by-layer (LBL) electrostatic adsorption has been studied. Electrochemical impedance spectroscopy (EIS) and UV–vis absorbance measurements were used to study the film assembly. All the results indicate that the uniform multilayer can be obtained on the polypyrrole (PPy) coated electrode surface and the hybridization reaction can be amplified by the layer-by-layer progress. The hybridization was detected by the reductive signal of Au-NPs and nonspecific adsorption was greatly eliminated by an unrelated DNA sequence to the target DNA. Under optimum conditions, a significant sensitivity enhancement had been obtained, and the detection limit was down to 3.20 × 10 −14 M when 6 layers assembled. The DNA biosensor has good stability and reproducibility.