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Nonenzymatic Sensors

Moreover, Au NPs not only provide a large surface area, high loading efficiency, and fast electron transfer but also stabilize the enzyme through electrostatic interactions. The MPH biosensor shows a rapid response and high selectivity for detection of methyl parathion, with a linear range from 0. The obtained nanohybrid of Au—PPy—rGO not only increased the surface area of the modified electrode but also showed excellent conductivity. AChE molecules were protected by a biocompatible 3D porous silica matrix to prevent them from leaking out and to retain their bioactivity.

Furthermore, the fabricated AChE biosensor displayed high stability, excellent activity, and fast response to OPs. This assembly protocol is expected to be used for the immobilization of various enzymes and proteins, leading to robust biosensors. Zhang and co-workers reported a facile electrochemical biosensing interface for sensitive glucose determination based on a Pt BSA nanocomposite along with the covalent adsorption of GOx. Upon the successive injection of glucose, the GOx-based biosensor catalyzed the oxidation of glucose to gluconolactone in the presence of oxygen, with the reduction peak current gradually decreased, making it suitable for glucose determination.

In addition to the simple modification of enzymes on the surface of electrode materials, embedding them within a different matrix is widely reported. Kale and co-workers succeeded in immobilizing GOD in a mixture containing silica sol—gel and poly vinyl alcohol composite film. LBL assembly has been selected as a reliable method to immobilize enzymes with preserved activity due to its simplicity and versatility.

In addition, CNTs and Au NPs could enhance the electrochemical signal by catalyzing the response of H 2 O 2 and effectively facilitate the electron transfer due to the good conductivity, further improving the detection sensitivity and stability. Compared to the immobilization of enzymes onto a substrate surface, incorporation of enzymes into a 3D matrix has the potential to increase the enzyme loading as well as to protect the enzyme from the surrounding environment.

To increase the loading of GOx and simplify glucose biosensor fabrication, Yang and co-workers prepared a hydrogel from Fc modified amino acid phenylalanine, which was utilized for the incorporation of GOx. Due to the improved enzyme loading and efficient electron transfer, the as-prepared glucose biosensor exhibited good performance for the electrochemical detection of glucose, such as high sensitivity, wide linear range, short response time, and good stability. Leopold and co-workers reported the xerogel biosensors containing composite films of 3-mercaptopropyl trimethoxysilane xerogel embedded with GOx, doped with Au NPs, monolayer protected clusters MPCs , and coated with an outer polyurethane layer.

It is found that the MPC-doped sol—gel glucose biosensors of this study are equal to or exceed comparable glucose biosensors reported previously. On the other hand, the 3D porous structure of the PAni hydrogel favored the high density immobilization of the enzyme and the mass transfer of the glucose. The glucose enzyme sensor based on this heterostructure exhibited unprecedented sensitivity and low detection limit. Reprinted from ref Copyright American Chemical Society. Enzyme-based electrochemical biosensors, especially the third-generation amperometric glucose biosensors, are fascinating because they function as the ideal biosensing model in the absence of mediators.

The direct electrical communication of GOx can also contribute to the detection of glucose at low potentials which are slightly positive from the redox potential of GOx. Considerable attempts to overcome the long electron-tunneling distance were made to realize the direct electrochemistry of enzymes.

Ye and co-workers reported a DET glucose biosensor based on GOx self-assembled on electrochemically reduced carboxyl graphene. Carboxylic acid groups remained and can effectively link with GOx. Well-defined and quasi-reversible redox peaks could be obtained. However, it is worth noting that the determination of glucose based on electroreduction of enzyme-consuming O 2 at low potentials close to the redox potential of GOx should conceptually belong to the first-generation amperometric glucose biosensors, rather than the third-generation ones.

To address this issue, Gorski and co-workers systematically investigated the signal transduction and enzyme activity in biosensors based on the GOx and CNTs embedded in a bioadhesive film of chitosan CHIT. Two main issues including the role of reactions relevant to the electrochemical glucose sensing the effect of CNT on the retention and enzymatic activity of GOx in CHIT films and in aqueous suspensions were studied.

The biosensor was sensitive to glucose in air-equilibrated solutions based on the O 2 -mediated enzymatic oxidation of glucose. The signal transduction relied on the net drop in a biosensor current that was caused by a decrease in a 4e — O 2 reduction current and an increase in a 2e — H 2 O 2 reduction current. The third-generation biosensors based on enzyme DET were also reported in these two years. In a typical example, Cui and co-workers utilized functionalized planar boron-doped diamond BDD electrode as a biosensing platform for biomolecule immobilization with GOx as a test model.

Amperometric responses of the GOx electrode to glucose were illustrated in both aerated and deaerated buffer solution to address whether the signal response to glucose can be attributed to DET. Different from the other reports, the result confirmed the bioelectrocatalytic activity of the electrical contacted GOx. Because of its larger surface area and negatively charged surface, the polycrystalline ITO film was a suitable electrode for the adsorption of CYP proteins while maintaining efficient DET and enzymatic activity.

On this basis, the simple ITO interface was applied to drug metabolism and inhibitor evaluation. Wang and co-workers for the first time employed small molecular hydrogel as a surrounding matrix to stabilize Cytochrome c Cyt c , further facilitating electron transfer between redox enzyme and electrode. In addition, Zhao et al. This two year period witnessed some novel electrochemical detection strategies and methods in developing new enzyme-based biosensors.

Vagin and co-workers reported a single-enzyme and membrane-free self-powered biosensor, in which both cathodic and anodic bioelectrocatalytic reactions are powered by cholesterol. Among them, ChOx was immobilized in a sol—gel matrix on both electrodes. Compared to either of the two individual electrodes, the self-powered sensor formed on the high surface-area carbon cloth electrodes, resulting in enhanced sensitivity.

On this basis, simultaneous determination of glucose and ATP concentrations by two independent bioselective elements holds great promise in novel sensing devices. Reed and co-workers demonstrated electronic field-effect transistors FETs as sensitive devices. An Al 2 O 3 -passivated Si nanowire used to mimic transistor operation was created for measuring enzyme—substrate interactions via the monitoring of pH change.

The enzyme kinetics can also be analyzed to accurately determine the kinetic constant. This direct, rapid, and label-free detection method can be readily generalized to many unrelated classes of substrates and enzymes. It was determined that the presence of the additional CNT layer was needed to achieve a urea-based EIS sensor with enhanced properties. Development of a novel electrochemical interface along with functional materials and enzyme immobilization plays a critical role for the rational design and construction of bioelectronic devices.

The change of resistance to charge transfer or amperometric current in the presence of GOx or urease resulted from the change of glucose or urea concentration, thus realizing simultaneous detection of glucose and urea based on in situ pH-switchable enzyme-catalyzed reactions. Karra and Gorski studied the nafion-induced current amplification in dehydrogenase-based biosensors. The increase in the biosensor current was attributed to the pH-driven increase in the enzyme activity inside the two-film interface.

The combination of the two-film interface with enzyme engineering to modify enzyme activity—pH profiles can lead to the enzyme-based biosensor devices with highly amplified current output. In addition, Liu and co-workers adopted an in-site immobilizing method to embed GOx in copolymer involving N , N -diethylacrylamide and methyl acrylic acid.

This multiresponsive electrochemical behavior of the system could be further employed to maximize the electrochemical oxidation of glucose catalyzed by GOx entrapped in the films with Fc COOH 2 as the mediator in solution. Future efforts were aimed at further miniaturization and integration of the electronic interface, further facilitating the development of advanced electroanalytical devices. For example, Wang and co-workers describe the first example of real-time noninvasive lactate sensing in human perspiration during exercise events using a flexible printed temporary-transfer tattoo electrochemical biosensor.

The lactate biosensor was used for the electrochemical detection of sweat lactate, thereby substantiating its utility for the noninvasive assessment of lactate levels and degree of physical exertion. Mao and co-workers demonstrated a microfluidic chip-based online electrochemical system for in vivo continuous and simultaneous monitoring of glucose, lactate, and ascorbate in rat brain. In electrochemical genosensors, single stranded DNA ssDNA fragments are immobilized onto the electrode surface as recognition probes for capturing the target DNA through hybridization.

In the presence of hybrids, signals are generated via various mechanisms and then detected electrochemically. According to electrochemical detection principles, several important factors should be considered for the achievement of good sensitivity and selectivity in the biodetection. The past two years has witnessed substantial advances toward the development of a high performance electrochemical sensing platform for DNA detection.

Recent review articles have focused on new methods and new signal amplification based on functional nanomaterials and enzymes in the DNA and RNA assays. Xu and co-workers recently evaluated the methods related to photoelectrochemical DNA biosensors. In the another review article, the sandwich assay based on the biotechnologies and nanotechnologies for nucleic acids was also introduced. Meanwhile, some other new methods and related progress were also investigated in the development of electrochemical DNA sensors. The electrochemical DNA sensing platform consists of capture probes immobilized on the sensing surface for capturing targets and signal probes with electrochemical tags for signal generation.

In the nucleic acid assays, good detection sensitivity can be achieved by optimizing hybridization conditions and improving hybridization efficiency. High detection specificity relies on the design of specific probes and the elimination of nonspecific binding on the sensing surface.


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Due to the proper distance between the nucleobases, the rigid amido bonds, the high flexibility of the aminoethyl linkers, and intramolecular hydrogen bonding, the peptide nucleic acid PNA probe has great sequence specific affinity and stability and has received great interest in DNA sensors. The regulation of the thermodynamic stability of the stem-loop structure decreases the background signal and increases the specificity as well.

The attached enzymes bring the electrocatalytic signal to amplify the detection. The combination of these effects improves the sensitivity of the sensor and can be applied to other miRNA detection methods. At the same time, DNA tetrahedral nanostructures containing a partially self-complementary region with a stem-loop hairpin structure were also innovatively designed.

As mentioned above, the immobilization of the probe DNA on the surface of the electrode dictates the performance of the resulting sensor. However, it is very difficult to precisely control the DNA spatial orientation and position on solid surfaces via formation of the self-assembled monolayer using thiolated ssDNA molecules.

A bovine serum albumin-monolayer-based probe carrier platform has been reported to improve the performance in comparison to a conventional thiolated ssDNA probe self-assembled monolayer-based electrochemical DNA hybridization biosensor. As is well understood, the application of redox labels is the simplest way to produce an electrochemical signal. However, a redox label can only transfer one or a few electrons to or from the electrode surface. The limitation of the number of electrons transferred directly affects the sensitivity of the DNA sandwich assay.

High sensitivity is highly desired since the DNA levels are low in some real problems, such as in pathogen DNA detection and cancer or infectious disease DNA detection. Enzymes have been successfully used for detection of analytes providing both recognition and amplification of the binding event with a detectable readout.

In this enzyme-based amplification system, DNA sensors based on enzymatic catalytic reactions, such as HRP 94 , 95 and alkaline phosphatase ALP , 96 , 97 were used as a substitute for the redox label, thus providing high, steady, and reproducible signal amplification. Besides, loading HRP onto various nanomaterials has become a promising way to further amplify the detection signal and achieve a lower detection limit for the analyte.

The activity of captured HRP was monitored by square-wave voltammetry measuring the electroactive enzymatic product in the presence of 2-aminophenol and hydrogen peroxide. The use of such labels greatly amplifies hybridization signals and enables the detection of full-length p BCR-ABL transcripts at subfemtomole levels, which corresponds to picograms of the target gene.

The resulting assay allowed robust discrimination between the perfect match and a three-base mismatch sequence. Ju et al. On the other hand, several novel electrochemical label-free methods using an enzyme-amplification strategy have been reported. The deposition of the insulating polymer film, poly 3,3-dimethoxybenzidine , was then carried out by the HRP-catalyzed polymerization of 3,3-dimethoxybenzidine in the presence of H 2 O 2.

Such a tool may open a new paradigm in routine miRNA analysis with a detection limit of 2. Besides this postamplification strategy toward signal production by a hybridization event, there are target recycling and other strategies via nuclease. They have drawn more and more concerns owing to its striking improvement for the detection sensitivity toward target analytes.

Ju and co-workers combined circular strand-displacement polymerization with silver enhancement to achieve a dual signal amplification. The released target found another molecular beacon to trigger the polymerization cycle, resulting in the multiplication of the reporter Au NPs on the sensor surface. Sequentially, the Au NPs-promoted silver deposition afforded a signal trace for electrochemical stripping analysis of target DNA.

This signal showed high selectivity and can be performed from 10 —16 to 10 —12 mol L —1 with a detection limit down to the subfemtomolar level. Given the limitation of the enzyme i. Because of their biological compatibility, high surface area, chemical stability, nontoxicity, excellent catalytic activity, and conductivity, the introduction of nanomaterials has greatly improved the analytical performance, amplified the detection signal, and stabilized the recognition probes or biosensing interface. It is well-known that the electrode materials as the key component are most widely used in electroanalytical investigations and play an important role in constructing high performance electrochemical sensing platforms to detect target molecules through different analytical principles.

Various nanomaterials, especially Au NPs and carbon nanomaterials, have been used as excellent carriers for loading numerous signal elements such as enzymes, oligonucleotides, and redox labels. Yu and co-workers adopted a hairpin sequence as the capture probe with a restriction site introduced into its stem segment. In contrast, the capture probe is opened by the target hybridization, deforming the restriction site and forcing the biotin tag away from the electrode. Au NPs modified with a large number of Fc-signaling probes are captured on the basis of the biotin—streptavidin complexation.

Furthermore, Fc tags can be dragged in close proximity to the electrode surface via hybridization between the signaling probes and the capture probe residues after Eco RI treatment, facilitating interfacial electron transfer and further enhancing the signal. This sensor achieves an ultralow detection limit to the zeptomole region and a wide dynamic range over 7 orders of magnitude. The presence of nonmatched probe reduces the cross-reaction between target DNA and matched probe on the Au NPs, resulting in increased sensitivity of the sandwich-type DNA biosensor.

Design of the cooperative amplification-based electrochemical sensor for the zeptomole detection of DNA. The newly developed nanomaterials can act as electroactive tracers for signal amplification by numerous signal species directly from themselves. Combined with effective methods for determination of nanotracers, ultrasensitive electrochemical DNA-based assays have been easily developed. Liu and co-workers presented a novel strategy for simultaneous detection of multiple DNA targets based on the use of different encoding metal ions as tags.

Metal ions bound to metallothionein molecules can be released after hybridization with DNA targets and then detected by stripping voltammetry. A novel dendritic QD nanocluster was constructed and used as versatile electrochemiluminescence ECL and electrochemical probes for the detection of DNA and cancer cells. Nanomaterials with enzyme-like characteristics were also used as a new method for signal amplification in genosensors.

Huang and co-workers fabricated a sensitive gap-electrical biosensor based on self-catalytic growth of unmodified Au NPs as conductive bridges for amplifying DNA hybridization events. Such catalytic activity can enlarge the diameters of Au NPs in the glucose and HAuCl 4 solution and result in a connection between most of the Au NPs and a conductive gold film formation with a dramatically increased conductance. It is of great significance to explore the interaction between functional materials with electrochemical probes in a genosensor system, which provides wide opportunities to develop a novel sensing platform.

Zuo and co-workers demonstrated an ultrasensitive detection platform for miRNA by combining hybridization chain reaction HCR amplification and the tetrahedral DNA nanostructure probes. In fact, most of the reports on genosensors involve multiple magnification approaches mentioned above. Tang and co-workers combined HCR amplification with silver nanotags to electrochemically monitor nucleic acids with high sensitivity.

Under optimal conditions, the target-triggered long-range DNA nanostructures presented good electrochemical behavior for the detection of human immunodeficiency virus DNA at a concentration as low as 0. Nanopore analysis has emerged as the simplest single-molecule technique. Various ssDNA with similar lengths can slide through a-hemolysin a-HL -based protein nanopore at a bias voltage and lead to an indistinguishable signal for DNA detection.

Kang et al. Furthermore, the fabricated sensor is specific and able to distinguish two base mismatches in the target sequence as well as capable of capturing and recording the target sequence from a heterogeneous mixture. DNA damage occurs frequently in organisms. Some endogenous and exogenous chemicals have been found to induce structural damages to nuclear DNA by base oxidation or modification.

If unrepaired, these damaged DNA may lead to gene mutation and even tumor generation. Electrochemical genosensors are well qualified for the rapid screening of industrial and environmental chemicals for their potential geno-toxicity. DNA aberrant methylation represses gene transcription, deregulates gene expression, and causes various human diseases.

Hence, detecting the DNA aberrant methylation level benefits the early diagnosis of some tumors and the epigenetic therapy for DNA methylation-related diseases. Then, an antihis tag antibody was captured to further inhibit the photocurrent and increase the detection sensitivity through the immunoreactions. This Bi 2 S 3 -based photoelectrochemical sensor possessed excellent photoelectron property and presented high detection specificity, even distinguishing single-base mismatched sequences.

Xie and co-workers reported the highly sensitive detection of DNA methylation, methyltransferase activity, and inhibitor screening based on DNA-Au NPs signal amplification. On the basis of this principle, DNA methylation could be determined on the basis of the voltammetric signal change of MB. Immunoassays are the detection platforms based on specific antigen—antibody recognition. They are well established standard biodetection methods used in clinical laboratories for disease diagnosis, in the food industry for food safety testing, and in monitoring environmental contamination.

Two recent review articles summarize the recent trends of electrochemical immunosensors toward POC diagnostics. Voltammetry and amperometry, such as linear sweep, differential pulse, square-wave, and stripping, are the most widely used electrochemical methods for immunoassay. Most of the strategies discussed below employ the sandwich immunoassay approach, in which the target antigen Ag is captured by its specific antibody Ab 1 and detected by labeled secondary antibody Ab 2.

In a sandwich-type immunoassay format, subsequent anodic-stripping voltammetric detection of cadmium released under acidic conditions from the coupled QDs was conducted at an in situ prepared mercury film electrode. The new immunoassay is promising for enzyme-free and cost-effective analysis of low-abundance biomarkers. Singh et al.

Electroanalytical Sensor Technology

Benefiting from this, the detection limit for carbonhydrate antigen CA was slightly higher than 0. Ren et al. More importantly, the in situ recycling of the proximate complex could be achieved with nicking endonuclease Nt. The proposed assay showed a wide detection range from 0. Parshetti et al. That enhanced signal amplification. Eletxigerra et al. The sensor was constructed by using magnetic microbeads and disposable screen-printed carbon electrodes, with the addition of hydroquinone as the electron transfer mediator and H 2 O 2 as the enzyme substrate.

After a thorough optimization of the assay, extremely low limits of detection were achieved: 2. Bhimji et al. Excellent performance relative to a commercial gold standard test was obtained, which was based on the surface functionalization of SU-8 and oxidation current of p -aminophenol. Because of the heterogeneous nature of the assay, there is no interference by electroactive substances or electrode fouling.

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The electrochemical immunoassay has also been used in other fields besides cancer biomarkers. The key architectural improvements were made, including: i avoiding use of secondary antibodies and ii utilizing yeast-scFv cell membrane fragments. Tlili et al. One involves screening and viability assays, employing bacteriophage as the recognition element in label-free electrochemical impedance spectroscopy.

The other approach is a confirmation by loop-mediated isothermal amplification to amplify specifically the E. In another study, biotinylated full antibody-based immunosensors have been optimized to enable the specific detection of pathogenic bacteria S. Electrodeposited polytyramine was used as a base layer for the conjugation of biotinyl antibodies via a biotin-Neutr Avidin bridge. Simultaneously, AChE enzyme activity of postexposure is also determined. In another work, we presented the first report on the development of the Fe 3 O 4 at TiO 2 magnetic NPs-based disposable electrochemical immunosensor with quantum dot-linked antibodies for sensitive and selective detection of the OP-butyrylcholinesterase adduct in human plasma.

Fe 3 O 4 at TiO 2 magnetic NPs not only selectively capture phosphorylated adduct by metal chelation but also directly separate it from biological matrices by simply exerting an external magnetic field. C Parallel measurement of AChE activity in a postexposure sample with and without reactivation.

Online Electrochemical Detectors: Fundamental Aspects And Analytical Applications 1984

The second strategy involved the use of mono- and dithiolated self-assembled monolayers carrying hydrazide functional groups. The immunosensors based on either a direct assay using electrochemical impedance spectroscopy or a sandwich-assay using differential pulse voltammetry for MS 2 phage detection were investigated. Their results showed that both immobilization protocols efficiently controlled the orientation of antibodies and the permeability of the electroactive species in solution, resulting in strategies that can be easily tailored to prepare highly sensitive electrochemical immunosensors.

The excellent features of the biosensors make the proposed method a valuable tool for gluten detection in foods. The combination of nanomaterials and immunosensors shows great potential for monitoring biomolecules and sensitive detection of target analytes. Nanomaterials can be used as carriers to load signal markers or directly as signal reporters for sensitively detecting biomarkers, and they can accelerate electron transfer when they are used as functional materials on electrode surfaces.

Graphene plays an important role in recent trends for immunosensors fabrication. Immobilization of the bioactive species is crucial for proper detection. Graphene offers an easy way to protect and stabilize these species. In addition to other nanomaterials, graphene-based immunosensors have exhibited good analytical characteristics and shown great promise for clinical applications. Recently, Wu et al.

Zhu et al. For recent examples, Lin et al. This reusable biosensor utilized a magnetic GO-modified gold electrode as the detection substrate. The fabrication method allows for reproducibility, ease of production and use, and storage stability enabling potential clinical use for rapid vascular endothelial growth factor detection. Lu et al.

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Moreover, in recent years, the study of the graphene-based derivative, such as heteroatom-doped graphene, GO, and graphene nanoribbon, has been popular and extensive, particularly with respect to electrochemical applications. The design of the immunosensor also involved a polydopamine functionalized GO hybrid conjugated to horseradish peroxidase-secondary antibodies by covalent bonds as a multilabeled and biocompatible probe to increase the electrochemical response.

The biomolecule—nanomaterials hybrid system has excellent prospects for interfacing biological recognition events with electronic signal transduction so as to design a new generation of bioelectronic devices with high sensitivity. Li et al. Lou et al. In the meantime, they also introduced an electrically heated carbon electrode in the detection procedure of the immunosensor and further improved the sensitivity.

After performing a sandwich immunoreaction, the quantitatively captured HRP-Au NP nanoprobes could catalyze oxidation of aniline to produce electroactive PAni on the immunosensor surface. The electrochemical measurement of PAni enabled a novel detection strategy for the HRP-based immunoassay. In the past two years, ECL has received much attention and become an important detection method. The resultant functionalized material has a high specific surface area, good electrocatalytic ability, and efficient photocatalytic activity and has been used to construct an ECL immunosensor for the detection of Streptococcus suis Serotype 2.

Electrochemical sensors and monitoring techniques | SpringerLink

A wide linear detection range of 0. Qi et al. The principle was based on the idea of encompassing heavy labels in larger carriers and on polyvalent binding motifs, employing the Ru1-encapsulated liposome peptides and the magnetic capture peptides. With the use of the catalyst with high loading of hemin as a signal tag of the secondary antibody, a novel ultrasensitive immunoassay for the carcinoembryonic antigen detection was demonstrated.

Photoelectrochemistry is a newly developed analytical method and now attracts substantial research scrutiny in various fields. It should be noted that the CdSe QDs acted as both photosensitizer and an alternative multivalent form for carbohydrate antigen with high binding affinity. This design would facilitate testing for disease-related sugar markers as well as evaluate the immunogenic properties of carbohydrate vaccine candidates. To achieve clinical or POC use, multiplexed electrochemical target detection has been investigated intensively in the last two years. Signal amplification was achieved through graphene modification of the immunodevice surface to accelerate the electron transfer and also the use of silica NPs as a tracing tag to label the signal antibodies.

Using the horseradish peroxidase-O-phenylenediamine-H 2 O 2 electrochemical detection system, the potential clinical applicability of this biosensor was demonstrated in the detection of four candidate cancer biomarkers in serum samples from cancer patients. The successful adaption of a bubble-based cartridge to the screen printed electrode system leads to automatic and rapid sample delivery at the electrode surface in one step with minimal user intervention. They have performed sensitive and selective detections of several biological targets, including tumor biomarkers and H1N1 split influenza vaccine.

Inorganic—organic poly 3,4-ethylenedioxythiophene -poly styrenesulfonate heterostructured light-emitting diodes LEDs based on ZnO nanorods were presented by Zhang et al. As a proof-of-concept, an advanced multiplexed photoelectrochemical immunosensor array was fabricated using the prepared LEDs as an excitation light source, and excellent performance for the detection of three different cancer biomarkers was achieved. Kong et al. The branched sensing electrode fabricated via photolithography on an ITO electrode consists of two separate circular areas 2 mm in diameter, named as W1 and W2 , which joined to a rectangular area for the electrical contact.

By equipping each branched sensing pad with different redox active substrates and different antibodies, the sensor can simultaneously respond to multiple targets in the sample. Liu et al. Cd II and Pb II ions were released from the surface of the corresponding nanoclusters by treatment with acid and detected by square wave anodic stripping voltammetry. There has been substantial progress in the development of electrochemical immunosensors; however, a major challenge for electrochemical sensors is still simultaneous detection of multiple targets in complex biological samples with reliability, portability, low cost, rapid response, and excellent selectivity and sensitivity.

Hence, the integration of electrochemical immunoassays into a disposable format will have great potential in the applications of clinical diagnostics, particularly for POC. Considering the vital role of cells in life science and human health, cell-related bioassaying has become a hot research topic within the past decades. Developing highly sensitive cytosensors will have a great impact in health care. Notably, electrochemical cytosensing approaches play a more and more important role in the analysis and detection of target cells due to the inherent advantages such as miniaturization, easy operation, rapid response, satisfied sensitivity, high selectivity, affordability, and real-time and nondestructive analysis.

Different electrochemical methods have been used in the investigation, including common electrochemical methods, the ECL method, and the photoelectrochemical method in label-free or sandwich assays. For label-free electrochemical cytosensing, the key is to fabricate a biocompatible recognition interface onto the electrode, coupled with highly sensitive read-out electrical signal to report the cell-recognition events with the help of biofunctionalized nanomaterials.

These microspheres were employed to develop the sensing layer with the conjugation of monoclonal anti-CEA antibody anti-CEA. With its excellent conductivity, stability, and biocompatibility, the 3D architectural Au BSA microspheres were used to develop a label-free cytosensor. Highly specific detection of BXPC-3 cells with a broader detection range and a detection limit of 18 cells mL —1 has been achieved.

Small molecules are also used for specific recognition of cells. Electrochemical impedance spectroscopy EIS detection results have showed that the detection limit for HL cells was achieved at cells per mL. In addition, aptamers can be useful molecular probes for cell detection. Quantitative determination of human colon cancer DLD-1 cells were performed by an electrochemical aptasensor. Carbon nanospheres not only accelerated electron transfer but also supplied a highly stable matrix for the efficient immobilization of target MUC-1 aptamer, considerably amplifying the electrochemical signals and resulting in sensitive detection performance toward DLD-1 cells.

The developed electrochemical cytosensor showed great reliable performance with satisfied sensitivity and specificity in detecting single MEAR cancer cells in 10 9 blood cells from a WBC sample. Real-time intracellular sensing is of great significance for advancing fundamental biological and clinical science. Competition strategy is another choice adopted for developing electrochemical cytosensors.

It has been reported that ultrasensitive electrochemical detection of leukemia cells achieved a detection limit down to 10 cells. Thereafter, the residual Au NPs on the nanoprobes will perform Au NP-catalyzed silver deposition for amplified signal read-out. This allows real time monitoring intracellular events. Owing to the advantage of multiplexing ability, high through-output performance, and low reagent consumption, microfluidic devices were used to study the on-site real-time assay of the proliferation and apoptosis of HeLa cells.

By virtue of its controllability and low background, ECL detection was widely used in cytosensing. A label-free ECL cytosensor was developed for specific determination of early apoptosis. That resulted in a steric effect on the interaction between sensor and coreactants. More captured cells would lead to a lower ECL signal. A therapeutical effect could be evaluated by quantifying an early apoptotic HepG2 cell induced by resveratrol.

The papers in this volume deal with an extraordinarily wide range of topics but all have the common focus of electrochemical detection as a practical chromatographic tool. While it is certainly not essential to be familiar with the theoretical principles in order to utilize it successfully, the determined user of electrochemical detector will seek to have an understanding of the background. Some of the following pages will provide an excellent grounding as well as pointing the potential user in the direction of proven and possible applications in a variety of fields.

The meeting, of which this book is a record, was the fifth event in the biannual Anglo-Czech Symposia in Electrochemistry. The contributions of the visitors to the scientific content of the meeting were enthusiastically received and their participation in the informal and social activities can only have furthered the cause of cooperation and good will between our two countries.

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