Investigation of Active and Inactivated Yeast Cells by Scanning Electrochemical Impedance Microscopy

Scanning electrochemical microscopy (SECM), electrochemical impedance spectroscopy (EIS) and scanning electrochemical impedance microscopy (SEIM) were used to investigate electrochemical activity of active and inactivated yeast Saccharomyces cerevisiae cells. SEIM experiment was performed using a unique electrochemical impedance spectrometer with a fast Fourier transform (FFT‐EIS) function, which enabled simultaneously perturb/evaluate electrochemical system at 50 frequencies. This allowed very quick observing the differences between impedance spectra, which were taken every few seconds. Therefore, we were able to apply SEIM for relatively fast determination of electrochemical impedance dependence on the distance between ultramicroelectrode (UME) and surface modified by immobilized yeast cells. It was determined that electrochemical activity and ‘breathing’ (a consumption of dissolved oxygen) of yeast can be electrochemically observed when the distance between UME and surface of yeast cells is in the range from 0 μm to 25 μm. Therefore, 25 μm is the maximum distance suitable for efficient investigation of yeast cell activity when experiments are performed in FFT‐SEIM mode. Charge transfer resistance of active and inactivated yeast cells was determined using EIS. It was calculated that charge transfer resistance of active yeast cells is 1.5 times lower than that of inactivated yeast cells. Lipophilic vitamin K₃ (Vit‐K₃) and hydrophilic vitamin K₁ (Vit‐K₁) were mixtured and used as redox mediators for charge transfer from yeast cells.     

Microelectrode Combinations of Gold and Polypyrrole Enable Highly Stable Two-electrode Electrochemical Impedance Spectroscopy Measurements under Turbulent Flow Conditions

An electrochemical impedance spectroscopy (EIS) sensor design is proposed based on a standard interdigitated electrode layout in which the smaller working electrode consists of gold (Au) whereas the larger combined counter and reference electrode is coated with a porous layer of polypyrrole (PPy) doped with polystyrene sulfonate (PSS) (PPy : PSS). Each electrode material was first characterized by EIS in a standard 3-electrode setup with subsequent spectra fitting by a modified Randles equivalent circuit. The differences in the spectra obtained by the PPy : PSS coated electrodes can be explained by an increased electroactive surface area due to the porous polymer film. The changes in morphology of the film are discussed with respect to the evolution of the elements of the electric equivalent circuit. When applying the Au/PPy : PSS electrode combination to a standard 2-electrode arrangement, the enlarged highly electroactive surface area of the PPy : PSS coating lowers the interfacial impedance in a way that mainly the gold working electrode contributes to the overall system impedance. Therefore, obtaining reproducible EIS signals depends only on the electrode's open-circuit potential (OCP) and on additional adsorption events at the gold electrode/electrolyte interface. We present a protocol for microelectrode coating with PPy : PSS, which enables highly stable 2-electrode EIS experiments without the need of a reference electrode. This combination is believed to be very useful if an integration of sensing electrodes inside Micro Total Analysis Systems is aspired.     

Electrochemical impedance spectroscopic measurements of FCCP-induced change in membrane permeability of MDCK cells

This study demonstrates a new electrochemical impedance spectroscopic (EIS) method for measurements of the changes in membrane permeability during the process of cell anoxia. Madin-Darby canine kidney (MDCK) cells were employed as the model cells and were cultured onto gelatin-modified glassy carbon (GC) electrodes. EIS measurements were conducted at the MDCK/gelatin-modified GC electrodes with Fe(CN)(6)(3-/4-) as the redox probe. The anoxia of the cells grown onto electrode surface was induced by the addition of carbonycyanide p-(trifluoromethoxy) phenylhydrazone (FCCP) into the cell culture, in which the MDCK/gelatin-modified GC electrodes were immersed for different times. The EIS results show that the presence of FCCP in the cell culture clearly decreases the charge-transfer resistance of the Fe(CN)(6)(3-/4-) redox probe at the MDCK/gelatin-modified GC electrodes, and the charge-transfer resistance decreases with increasing time employed for immersing the MDCK/gelatin-modified GC electrodes into the cell culture containing FCCP. These results demonstrate that the EIS method could be used to monitor the changes in the cell membrane permeability during the FCCP-induced cell anoxia. To simulate the EIS system, a rational equivalent circuit was proposed and the values of ohmic resistance of the electrolyte, charge-transfer resistance and constant phase elements for both the gelatin and the cell layers are given with the fitting error in an acceptable value. This study actually offers a new and simple approach to measuring the dynamic process of cell death induced by anoxia through monitoring the changes in the cell membrane permeability.     

Some Properties of Sodium Dodecyl Sulfate Functionalized Multiwalled Carbon Nanotubes Electrode and Its Application on Detection of Dopamine in the Presence of Ascorbic Acid

A sodium dodecyl sulfate (SDS) functionalized multiwalled carbon nanotubes (MWNTs) electrode (SDS/MWNTs) was successfully constructed in this study. The electrochemical property of the SDS/MWNTs electrode has been characterized by electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Nyquist plots suggest that the immersion time of SDS affects the resistances of the MWNTs electrodes. The thickness of adsorbed SDS on MWNTs surface is estimated to be 1.23 nm, which is close to the value of SDS monolayer. CV results demonstrate a 5‐fold enhanced response for dopamine (DA) at the SDS/MWNTs electrode compared to the bare MWNTs one. DPV results illustrate that DA can be selectively determined in the presence of high concentration ascorbic acid (AA) with a linear range from 20 μM to 0.20 mM and a sensitivity of 0.024 μA μM^?1 at the SDS/MWNTs electrode.     

Single-wall carbon nanotube-based proton exchange membrane assembly for hydrogen fuel cells

A membrane electrode assembly (MEA) for hydrogen fuel cells has been fabricated using single-walled carbon nanotubes (SWCNTs) support and platinum catalyst. Films of SWCNTs and commercial platinum (Pt) black were sequentially cast on a carbon fiber electrode (CFE) using a simple electrophoretic deposition procedure. Scanning electron microscopy and Raman spectroscopy showed that the nanotubes and the platinum retained their nanostructure morphology on the carbon fiber surface. Electrochemical impedance spectroscopy (EIS) revealed that the carbon nanotube-based electrodes exhibited an order of magnitude lower charge-transfer reaction resistance (R(ct)) for the hydrogen evolution reaction (HER) than did the commercial carbon black (CB)-based electrodes. The proton exchange membrane (PEM) assembly fabricated using the CFE/SWCNT/Pt electrodes was evaluated using a fuel cell testing unit operating with H(2) and O(2) as input fuels at 25 and 60 degrees C. The maximum power density obtained using CFE/SWCNT/Pt electrodes as both the anode and the cathode was approximately 20% better than that using the CFE/CB/Pt electrodes.     

钡添加剂对二次碱性锌电极性能的影响

采用直接化学合成法制备以钡作添加剂的二次碱性锌电极活性物质,样品的表面形貌及其晶态结构由SEM和XRD表征,并以循环伏安、充放电循环及电化学阻抗谱测定样品的电化学性能.实验表明,锌电极因Ba的添加而生成一种化学式为BaZn(OH)4·xH2O的锌酸钡,改善了电极的电化学性能.此外发现,Zn电极的电化学阻抗谱含有一个对应于电极表面放电产物覆盖率的时间常数.     

Development of impedimetric and optical calcium biosensor by using modified gold electrode with porcine S100A12 protein

We describe the development of a label free method to analyze the interactions between Ca(2+) and the porcine S100A12 protein immobilized on polyvinyl butyral (PVB). The modified gold electrodes were characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), scanning electron microscopy (SEM) and surface plasmon resonance (SPR) techniques. SEM analyses of PVB and PVB-S100A12 showed a heterogeneous distribution of PVB spherules on gold surface. EIS and CV measurements have shown that redox probe reactions on the modified gold electrodes were partially blocked due the adsorption of PVB-S100A12, and confirm the existence of a positive response of the immobilized S100A12 to the presence of calcium ions. The biosensor exhibited a wide linear response to Ca(2+) concentrations ranging from 12.5 to 200mM. The PVB-S100A12 seems to be bound to the gold electrode surface by physical adsorption; we observed an increase of 1184.32m° in the SPR angle after the adsorption of the protein on the PVB surface (in an indication that 9.84ng of S100A12 are adsorbed per mm(2) of the Au-PVB electrode), followed by a further increase of 581.66m° after attachment of the Ca(2+) ions. In addition, no SPR response is obtained for non-specific ions. These studies might be useful as a platform for the design of new reusable and sensitive biosensing devices that could find use in the clinical applications.     

Cell detection and counting through cell lysate impedance spectroscopy in microfluidic devices

Cell-based microfluidic devices have attracted interest for a wide range of applications. While optical cell counting and flow cytometry-type devices have been reported extensively, sensitive and efficient non-optical methods to detect and quantify cells attached over large surface areas within microdevices are generally lacking. We describe an electrical method for counting cells based on the measurement of changes in conductivity of the surrounding medium due to ions released from surface-immobilized cells within a microfluidic channel. Immobilized cells are lysed using a low conductivity, hypotonic media and the resulting change in impedance is measured using surface patterned electrodes to detect and quantify the number of cells. We found that the bulk solution conductance increases linearly with the number of isolated cells contributing to solution ion concentration. The method of cell lysate impedance spectroscopy is sensitive enough to detect 20 cells microL(-1), and offers a simple and efficient method for detecting and enumerating cells within microfluidic devices for many applications including measurement of CD4 cell counts in HIV patients in resource-limited settings. To our knowledge, this is the most sensitive approach using non-optical setups to enumerate immobilized cells. The microfluidic device, capable of isolating specific cell types from a complex bio-fluidic and quantifying cell number, can serve as a single use cartridge for a hand-held instrument to provide simple, fast and affordable cell counting in point-of-care settings.     

Micromachined impedance spectroscopy flow cytometer for cell analysis and particle sizing

A new cytological tool, based on the microCoulter particle counter (microCPC) principle, aimed at diagnostic applications for cell counting and separation in haematology, oncology or toxicology is described. The device measures the spectral impedance of individual cells or particles and allows screening rates over 100 samples s(-1) on a single-cell basis. This analyzer is intended to drive a sorting actuator producing a subsequent cell separation. Size reduction and integration of functions are essential in achieving precise measurements and high throughput. 3D finite element simulations are presented to compare various electrode geometries and their influence on cell parameters estimation. The device is based on a glass-polyimide microfluidic chip with integrated channels and electrodes microfabricated at the length scale of the particles to be investigated (1-20 microm). A laminar liquid flow carries the suspended particles through the measurement area. Each particle's impedance signal is recorded by a differential pair of microelectrodes using the cell surrounding media as a reference. The micromachined chip and processing electronic circuit allow simultaneous impedance measurements at multiple frequencies, ranging from 100 kHz to 15 MHz. In this paper, we describe the microfabrication and characterisation of an on-chip flow-cytometer as the first building block of a complete cell-sorting device. We then discuss the signal conditioning technique and finally impedance measurements of cells and particles of different sizes and types to demonstrate the differentiation of subpopulations in a mixed sample.     

Impedimetric biosensor for cancer cell detection

Electrochemical impedance spectroscopy was evaluated for the label free detection of MCF-7 cancer cell in which c-erbB-2 receptor is overexpressed on the cell surfaces. Anti-c-erbB-2, used as a specific antibody, was immobilized on electrogenerated polypyrrole-NHS on electrodes via covalent linking. The polymer formation, the grafting of the antibody, and the recognition event with the cancer cells using MCF-7 as a model cell line, were characterized by using cyclic voltammetry and fluorescence microscopy. The impedimetric sensor showed high sensitivity from 100 to 10 000 cell/mL without needing any labeling step and represents an efficient transduction method for cell selective detection.     

钛/活细胞界面原位EIS研究

设计了原位电解池用于生物材料/活细胞界面电化学交流阻抗谱(EIS)的测量, 原位观测了钛/MG63细胞界面EIS行为及其随时间的变化. 原位电解池中引入Ag/AgCl作为参比电极, 解决了EIS测量过程中高频部分信号失真的问题. 钛/MG63细胞界面EIS研究初步结果表明: 活细胞在钛电极表面附着, 可改变界面双电层结构和电极的表面状态, 进而影响界面的电化学性质. 实验发现, 由于钛电极表面氧化层的阻抗信号处于中低频段, 而细胞膜层自身的电化学阻抗响应位于高频范围, 由此, 可分别分析不同的电极过程, 解析其相互关系, 研究活细胞在生物材料表面的作用机理.     

Multiplexing microelectrodes for dielectrophoretic manipulation and electrical impedance measurement of single particles and cells in a microfluidic device

This work presents a microfluidic device, which was patterned with (i) microstructures for hydrodynamic capture of single particles and cells, and (ii) multiplexing microelectrodes for selective release via negative dielectrophoretic (nDEP) forces and electrical impedance measurements of immobilized samples. Computational fluid dynamics (CFD) simulations were performed to investigate the fluidic profiles within the microchannels during the hydrodynamic capture of particles and evaluate the performance of single‐cell immobilization. Results showed uniform distributions of velocities and pressure differences across all eight trapping sites. The hydrodynamic net force and the nDEP force acting on a 6 μm sphere were calculated in a 3D model. Polystyrene beads with difference diameters (6, 8, and 10 μm) and budding yeast cells were employed to verify multiple functions of the microfluidic device, including reliable capture and selective nDEP‐release of particles or cells and sensitive electrical impedance measurements of immobilized samples. The size of immobilized beads and the number of captured yeast cells can be discriminated by analyzing impedance signals at 1 MHz. Results also demonstrated that yeast cells can be immobilized at single‐cell resolution by combining the hydrodynamic capture with impedance measurements and nDEP‐release of unwanted samples. Therefore, the microfluidic device integrated with multiplexing microelectrodes potentially offers a versatile, reliable, and precise platform for single‐cell analysis.     

Novel photoelectrochemical immunosensor for MCF-7 cell detection based on n-p organic semiconductor heterojunction

In this work, we developed a novel photoelectrochemical（PEC） sensor based on n-p organic semiconductor heterojunction for sensitive detecting MCF-7 cancer cells. BTA-C4 Ph and PM6 were designed as photoactive materials to form n-p heterojunction, which greatly enhanced the photoelectric conversion efficiency. Antibody-modified magnetic nanoparticles were utilized to capture and separate MCF-7 cells from samples. Detection of MCF-7 is ascribed to the loading of MCF-7 onto BTA-C4 Ph-PM6 modified e...     

An electrochemical impedance immunoanalytical method for detecting immunological interaction of human mammary tumor associated glycoprotein and its monoclonal antibody

An electrochemical impedance immunosensor has been developed for the specific detection of immunological interaction between human mammary tumor associated glycoprotein and its monoclonal antibody (GP1D8). Antibody proteins were immobilized by spontaneous adsorption of antibody on gold. Consequently, electrochemical impedance spectroscopy (EIS) measurements of a gold electrode coated with the antibody showed changes in a.c. current response after the addition of the specific antigen. The successful immunological reaction between the immobilized antibody–antigen at the electrode surface could be monitored.     

Construction of an electrochemical sensor based on the electrodeposition of Au-Pt nanoparticles mixtures on multi-walled carbon nanotubes film for voltammetric determination of cefotaxime

Mixtures of gold-platinum nanoparticles (Au-PtNPs) are fabricated consecutively on a multi-walled carbon nanotubes (MWNT) coated glassy carbon electrode (GCE) by the electrodeposition method. The surface morphology and nature of the hybrid film (Au-PtNPs/MWCNT) deposited on glassy carbon electrodes is characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS), X-ray diffraction (XRD), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) techniques. The modified electrode is used as a new and sensitive electrochemical sensor for the voltammetric determination of cefotaxime (CFX). The electrochemical behavior of CFX is investigated on the surface of the modified electrode using linear sweep voltammetry (LSV). The results of voltammetric studies exhibited a considerable improvement in the oxidation peak current of CFX compared to glassy carbon electrodes individually coated with MWCNT or Au-PtNPs. Under the optimized conditions, the modified electrode showed a wide linear dynamic range of 0.004-10.0 μM with a detection limit of 1.0 nM for the voltammetric determination of CFX. The modified electrode was successfully applied for the accurate determination of trace amounts of CFX in pharmaceutical and clinical preparations.     

交流阻抗法研究四羧基酞菁锌掺杂的二氧化钛半导体电极

用电沉积和丝网印刷法制备了纳米二氧化钛膜电极及四羧基酞菁锌(ZnPcTc)掺杂的多孔纳米二氧化钛半导体电极. 采用交流阻抗法(EIS)对二氧化钛膜的电子传输性能以及界面性质进行了表征, 确定了各阻抗弧对应的电极过程. 采用合理的模型计算了电极的电子传输动力学参数. 结果表明, 掺杂ZnPcTc后, 膜电阻明显降低, 且电极-电解液界面电容有所增大, 有利于TiO2电极在染料敏化太阳能电池器件中的应用.     

纳米碳管作为氧扩散电极催化层第二相碳载体的电极特性

A novel gas diffusion electrode using binary carbon supports (carbon nanotubes and active carbon) as the catalyst layer was prepared. The electrochemical properties for oxygen reduction reaction (ORR) in alkaline electrolyte were investigated by polarization curves and electrochemical impedance spectroscopy. The results show that the binary-support electrode exhibits higher electrocatalytic activity than the single-support electrode, and the best performance is obtained when the mass ratio of carbon nanotubes and activated carbon is 50 ∶50. The results from their electrode kinetic parameters indicate that the introduction of carbon nanotubes as a secondary support provides high accessible surface area, good electronic conductivity and fast ORR kinetics. The electrocatalytic activity of binary-support electrodes is obviously improved by the deposition of Pt nanoparticles on carbon nanotubes, even at very low Pt loading (45.7 μg/cm2). In addition, the EIS analysis results show that the process of ORR may be controlled by diffusion of oxygen in the thin film for binary-support electrodes with or without Pt catalyst.     

Development and Evaluation of a Nanoporous Iron (Hydr)oxide Electrode for Phosphate Sensing

Nanoporous iron (hydr)oxide electrodes are evaluated as phosphate sensors using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The intensity of the reduction peak current (I_cp) of the ferrihydrite working electrode is tied to phosphate concentration at low pH; however, a hematite electrode combined with the use of EIS provided reliable sensing data at multiple pH values. Nanoporous hematite working electrodes produced an impedance phase component (θ) that shifts with increasing phosphate, and, at chosen frequencies, θ values were fitted for the range 1 nM to 0.1 mM phosphate at pH 4 and pH 7 in 5 mM NaClO₄.     

Application of Impedimetric and Voltammetric Genosensor for Detection of a Biological Warfare: Anthrax

A strategy for the detection of anthrax, which is a potential biological weapon by using an electrochemical genosensing technology, is investigated. An alkanathiol‐linked or unlabeled capture probe related to B. anthracis is immobilized onto gold or graphite electrode surface. A 101‐mer anthrax target is used for hybridization. The extent of hybridization between probe and target sequences is determined by using differential pulse voltammetry (DPV) and electrochemical impedance spectrometry (EIS). EIS analysis are based on electron transfer resistance (R_ct) in the presence of [Fe(CN)₆]^3?/4? and DPV measurements are based on transduction of both guanine oxidation and Meldola's blue (MDB) reduction signal as hybridization indicator. The response of the probe‐modified electrodes which was interacted with a noncomplementary sequence was the same as the responses of probe‐modified surface and proved the specifity of the hybridization with the target. According to these results the developed genosensors based on EIS and DPV techniques can be employed for rapid and selective detection of B. anthracis.     

Dielectrophoretic cell trapping for improved surface plasmon resonance imaging sensing

The performance of conventional surface plasmon resonance (SPR) biosensors can be limited by the diffusion of the target analyte to the sensor surface. This work presents an SPR biosensor that incorporates an active mass‐transport mechanism based on dielectrophoresis and electroosmotic flow to enhance analyte transport to the sensor surface and reduce the time required for detection. Both these phenomena rely on the generation of AC electric fields that can be tailored by shaping the electrodes that also serve as the SPR sensing areas. Numerical simulations of electric field distribution and microparticle trajectories were performed to choose an optimal electrode design. The proposed design improves on previous work combining SPR with DEP by using face‐to‐face electrodes, rather than a planar interdigitated design. Two different top‐bottom electrode designs were experimentally tested to concentrate firstly latex beads and secondly biological cells onto the SPR sensing area. SPR measurements were then performed by varying the target concentrations. The electrohydrodynamic flow enabled efficient concentration of small objects (3 μm beads, yeasts) onto the SPR sensing area, which resulted in an order of magnitude increased SPR response. Negative dielectrophoresis was also used to concentrate HEK293 cells onto the metal electrodes surrounded by insulating areas, where the SPR response was improved by one order of magnitude.