In vitro drug screening models derived from different PC12 cell lines for exploring Parkinson's disease based on electrochemical signals of catecholamine neurotransmitters.

Gold nanostructures and a Nafion modified screen-printed carbon electrode (Nafion/AuNS/SPCE) were developed to assess the cell viability of Parkinson's disease (PD) cell models. The electrochemical measurement of cell viability was reflected by catecholamine neurotransmitter (represented by dopamine) secretion capacity, followed by a traditional tetrazolium-based colorimetric assay for confirmation. Due to the  capacity to synthesize, store, and release catecholamines as well as their unlimited homogeneous proliferation, and ease of manipulation, pheochromocytoma (PC12) cells were used for PD cell modeling. Commercial low-differentiated and highly-differentiated PC12 cells, and home-made nerve growth factor (NGF) induced low-differentiated PC12 cells (NGF-differentiated PC12 cells) were included in the modeling. This approach achieved sensitive and rapid determination of cellular modeling and intervention states. Notably, among the three cell lines, NGF-differentiated PC12 cells displayed the enhanced neurotransmitter secretion level accompanied with attenuated growth rate, incremental dendrites in number and length that were highly resemble with neurons. Therefore, it was selected as the PD-tailorable modeling cell line. In short, the electrochemical sensor can be used to sensitively determine the biological function of neuron-like PC12 cells with negligible destruction and to explore the protective and regenerative impact of various substances on nerve cell model.

Fenton reaction-mediated fluorescence quenching of N-acetyl-L-cysteine-protected gold nanoclusters: analytical applications of hydrogen peroxide, glucose, and catalase detection.

Given the importance of hydrogen peroxide (H2O2) in many biological processes and its wide application in various industries, the demand for sensitive, accurate, and economical H2O2 sensors is high. In this study, we used Fenton reaction-stimulated fluorescence quenching of N-acetyl-L-cysteine-protected gold nanoclusters (NAC-AuNCs) as a reporter system for the determination of H2O2. After the experimental conditions were optimized, the sensing platform enabled the analysis of H2O2 with a limit of detection (LOD) as low as 0.027 µM. As the glucose oxidase cascade leads to the generation of H2O2 and catalase catalyzes the decomposition of H2O2, these two biocatalytic procedures can be probed by the Fenton reaction-mediated quenching of NAC-AuNCs. The LOD for glucose was found to be 0.18 µM, and the linear range was 0.39-27.22 µM. The LOD for catalase was 0.002 U mL(-1), and the linear range was 0.01-0.3 U mL(-1). Moreover, the proposed sensing methods were successfully applied for human serum glucose detection and the non-invasive determination of catalase activity in human saliva, demonstrating their great potential for practical applications.

Ultrasensitive amperometric determination of hand, foot and mouth disease based on gold nanoflower modified microelectrode.

Given the widespread use of point-of-care testing for diagnosis of disease, micro-scale electrochemical deoxyribonucleic acid (DNA) biosensors have become a promising area of research owing to its fast mass transfer, high current density and rapid response. In this study, a gold nanoparticles modified gold microelectrode (AuNPs/Au-Me) was constructed to determine the hand, foot and mouth disease (HFMD)-related gene. The noble metal nanoparticles modification yielded ca. 7.4-fold increase in electroactive surface area of microelectrode, and the signal for HFMD-related gene was largely magnified. Under optimal conditions, the biosensor exhibited salient selectivity and sensitivity with a low detection limit of 0.3 fM (S/N = 3), which is sufficient for clinical diagnosis of HFMD. Additionally, the developed AuNPs/Au-Me was successfully applied to determining the polymerase chain reaction (PCR) amplified products of target gene. Thus, the electrochemical DNA biosensor possesses great potential in early-stage diagnosis and long-term monitoring of various disease.

A Y-shape-structured electrochemiluminescence biosensor based on carbon quantum dots and locked nucleic acid probe for microRNA determination with single-base resolution.

Since microRNAs (miRNAs) are predictors of tumorigenesis, accurate identification and quantification of miRNAs with highly similar sequences are expected to reflect tumor diagnosis and treatment. In this study, a highly selective and sensitive electrochemiluminescence (ECL) biosensor was constructed for miRNAs determination based on Y-shaped junction structure equipped with locked nucleic acids (LNA), graphene oxide-based nanocomposite to enrich luminophores, and conductive matrix. Specifically, two LNA-modified probes were designed for specific miRNA recognition, that is, a dual-amine functionalized hairpin capture probe and a signal probe. A Y-shaped DNA junction structure was generated on the electrode surface upon miRNA hybridizing across the two branches, so as to enhance the selectivity. Carbon quantum dots-polyethylene imine-graphene oxide (CQDs-PEI-GO) nanocomposites were developed to enrich luminophores CQDs, and thus enhancing the ECL intensity. For indirect signal amplification, an electrochemically activated poly(2-aminoterephthalic acid) (ATA) film decorated with gold nanoparticles was prepared on electrode as an effective matrix to accelerate the electron transfer. The fabricated ECL biosensor achieved sensitive determination of miRNA-222 with a limit-of-detection (LOD) as low as 1.95 fM (S/N = 3). Notably, Y-shaped junction structures equipped with LNA probes endowed ECL biosensor with salient single-base discrimination ability and anti-interference capacity. Overall, the proposed Y-shaped ECL biosensor has considerable promise for clinical biomarker determination.

Electrochemical monitoring the effect of drug intervention on PC12 cell damage model cultured on paper-PLA 3D printed device.

Three-dimensional (3D) cell culture system, as an alternative approach for traditional cell culture, attracts great attention because of physiological relevance and great microenvironment similarity to human conditions. Herein, a facile paper-polylactic (PLA) platform that was fabricated by wax printing and 3D printing, coupled with electrochemical sensor, was designed for the construction and intervention of 3D cell damage model. Pheochromocytoma cells (PC12) and bone marrow mesenchymal stem cells (BMSCs) were seeded on the paper-PLA 3D platforms and displayed the features of uniform distribution, good adhesion and perfect proliferation, as well as decreased circularity when compared to those grown on the two-dimensional (2D) interfaces. The electrochemical sensors revealed cell viability by monitoring dopamine released by cell models, ascertaining the applicability of the paper-PLA platform to a long-term 3D cell culture and drug assessment. Additionally, the results revealed that donepezil and BMSCs-secreted active molecules exhibited stronger cytoprotective effect against amyloid-beta oligomers-induced cell damage on the paper-PLA 3D printed platforms, indicating the cell damage model and the cell intervention model were achieved successfully in the simulated in vivo physiological microenvironment. Thus, the proposed paper-PLA platform may serve as a promising candidate for efficient drug screening and toxicity evaluation due to its simple structure, low cost, and convenient integration of 3D cell culture and activity evaluation.

Simultaneous voltammetric determination of norepinephrine, ascorbic acid and uric acid on polycalconcarboxylic acid modified glassy carbon electrode.

A novel polycalconcarboxylic acid (CCA) modified glassy carbon electrode (GCE) was fabricated by electropolymerization and then successfully used to simultaneously determine ascorbic acid (AA), norepinephrine (NE) and uric acid (UA). The characterization of electrochemically synthesized Poly-CCA film was investigated by atomic force microscopy (AFM), electrochemical impedance spectroscopy (EIS) and voltammetric methods. It was found that the electrochemical behavior of the polymer-modified electrode depended on film thickness, i.e., the electropylmyerization time. Based on the electrochemical data, the charge transfer coefficient (alpha) and the surface coverage (Gamma) were calculated. This poly-CCA modified GCE could reduce the overpotential of ascorbic acid (AA), norepinephrine (NE) and uric acid (UA) oxidation in phosphate buffer solution (pH 6.0), while it increases the peak current significantly. The current peak separations of AA/NE, NE/UA and AA/UA on this modified electrode are 91mV, 256mV and 390mV in CV at 100mVs(-1), respectively. Therefore, the voltammetric responses of these three compounds can be well resolved on the polymer-modified electrode, and simultaneously determination of these three compounds can be achieved. In addition, this modified electrode can be successfully applied to determine AA and NE in injection and UA in urine samples without interferences.

Off-line form of the Michaelis-Menten equation for studying the reaction kinetics in a polymer microchip integrated with enzyme microreactor.

We firstly transformed the traditional Michaelis-Menten equation into an off-line form which can be used for evaluating the Michaelis-Menten constant after the enzymatic reaction. For experimental estimation of the kinetics of enzymatic reactions, we have developed a facile and effective method by integrating an enzyme microreactor into direct-printing polymer microchips. Strong nonspecific adsorption of proteins was utilized to effectively immobilize enzymes onto the microchannel wall, forming the integrated on-column enzyme microreactor in a microchip. The properties of the integrated enzyme microreactor were evaluated by using the enzymatic reaction of glucose oxidase (GOx) with its substrate glucose as a model system. The reaction product, hydrogen peroxide, was electrochemically (EC) analyzed using a Pt microelectrode. The data for enzyme kinetics using our off-line form of the Michaelis-Menten equation was obtained (K(m) = 2.64 mM), which is much smaller than that reported in solution (K(m) = 6.0 mM). Due to the hydrophobic property and the native mesoscopic structure of the poly(ethylene terephthalate) film, the immobilized enzyme in the microreactor shows good stability and bioactivity under the flowing conditions.

Novel coupling mechanism-based imaging approach to scanning electrochemical microscopy for probing the electric field distribution at the microchannel end.

A novel coupling mechanism-based imaging approach to scanning electrochemical microscopy (SECM) was used to image the distribution of electric field at the end channel of a poly(dimethylsiloxane) (PDMS) capillary electrophoresis (CE) microchip in the absence of redox species. The coupling imaging mechanism was systematically investigated and qualitatively illustrated. It was proved that the distribution of solution potentials within the scanning plane caused a different reduction rate of water at the tip electrode, which led to the variation in tip current. Within the scanning plane, the solution potentials measured in the central area of the microchannel were usually higher than those measured outside. The SECM images showed a strong dependence on tip potential, tip-to-channel distance, and separation potential. According to the Tafel equation, SECM images were converted to parameters that directly showed the distribution of solution potential. Change in the solution potential along the central axial line of the microchannel was also continuously sensed by allowing the tip to approach the microchannel in the presence of high voltage. Using dopamine as a model compound, the effect of solution potential on electrochemical detection was estimated by detecting separation parameters.