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Current Medicinal Chemistry, 2008, 15, ????-???? 1

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Trends in Cell-Based Electrochemical Biosensors

Lin Ding 1, Dan Du 1, Xueji Zhang* ,2 and Huangxian Ju* ,1 1Key Laboratory of Analytical Chemistry for Life Sciences (Ministry of Education of China), Department of Chemistry,

Nanjing University, Nanjing 210093, P. R. China2 Department of Chemistry, Universi ty of South Florida, 4202 East Fowler Avenue, CHE 305, Tampa, FL 33620-5250,

USAAbstract: Cell-based electrochemical biosensors have contributed tremendously to the fields of biology, medicine, chem-istry, pharmacology, and environmental science. With electrochemical transducers and whole cells as the recognition ele-ments, these biosensors provide new horizons for biosensing and life science research. This review focuses on the re-search accomplishments on this topic over the last three years, and is divided into three sections according to the types ofcellular responses. Our aim is to highlight how simple and sensitive electrochemical methods can be coupled with cells byvirtue of the integration of interface control, nanotechnology and genetic engineering to generate new enabling technolo-gies. Some specific examples to demonstrate how these sensors are useful in medicinal chemistry and drug design havealso been discussed. It is hoped that this review can provide inspiration for the development of fast, selective, sensitive,and convenient detection and diagnosis platforms.

Keywords: Electrochemical, cells, biosensors, impedance, microelectrode array, review.

1. INTRODUCTION

Electrochemical biosensing technology, with intimatelycoupled biological recognition elements and electrochemicaltransduction units, has been extensively used to obtain quan-titative or semi-quantitative information of plenty of analytes[1,2]. Advantages include high sensitivity, selectivity, rapidanalysis and the ability to operate in turbid solutions [3].Furthermore, the electrochemical equipments are more ame-nable to miniaturization, and the continuous response of anelectrode system allows for on-line control. According to thetype of recognition elements, electrochemical biosensors can

be classified into immunological , enzymatic, non-enzymaticreceptor, DNA and whole-cell electrochemical biosensors.

Whole cells, as more complex biological recognition ele-

ments, hold the promise of presenting the intact sub-cellularmachinery for the identification of physiologically relevantevents, and providing functional and analytical information[4-7]. The internal amplification cascades of cells can beused to increase the sensitivity of the devices. In addition,compared with enzyme, the self-sustaining whole cells areless expensive and more stable during the production proc-ess. Over the past decades, different types of cells have beenincorporated into electrochemical biosensor, including mi-croorganisms [7,8], mammalian cells [4-6] and recombinantcells (see Glossary) [9] in the immobilization format of sin-gle cell, cell layer and cell network. These cell-basedelectrochemical biosensors have a wide range of applicationsin pharmacology, medicine, cell biology, toxicology,neuroscience, and environmental monitoring [5,6,10].Furthermore, they are of paramount importance for medici-nal chemistry, because cellular electrochemical responses tovarious medicinal compounds provide “true” pharmacologyof the medicinal components. They a lso give comprehensive

*Address correspondence to these authors at the Key Laboratory of Analyti-cal Chemistry for Life Sciences (Ministry of Education of China), Depart-ment of Chemistry, Nanjing University, Nanjing 210093, P. R. ChinaDepartment of Chemistry, University of South Florida, 4202 East FowlerAvenue, CHE 305, Tampa, FL 33620-5250, USA; Tel: +86 25 8359 3593;Fax: +86 25 8359 3593; E-mail: [email protected]; [email protected]

information of the mechanistic aspects of drug activity, such

as stereochemistry, diffusion, solubility, metabolism, membrane permeability, etc. This has tremendous implications fo rthe drug discovery process and can expedite the research inthis field. Moreover, as drug costs are directly tied to candi-date failures, it is beneficial to predict drug toxicity andassess drug effects in situ , in real-time and on a large popula-tion of cells but with the resolution of a single cell at veryearly stages of development [11]. This can be readilyachieved by cell-based electrochemical biosensors. Current

problems facing the applications of the devices are analyticalstrategies, batch-to-batch reproducibility, and lack of selec-tive placement of biological cells on the sensing sites [5].Advancement in technology integration, nanomaterials utili-zation, interface control, and genetic engineering has pro-

vided a promising opportunity for meeting these challenges.Although cell-based biosensing technology is still at its

nascent development stage, several reviews have been pub-lished [1,5,6,8,10,12-17] and mostly focused on narrowclasses of cells or specific applications. This paper reviewsthe trends of cell-based electrochemical biosensors with awide range of whole cells in the last three years, includingsensors not only in the narrowly defined sense, in which thecells act as recognition elements [16], but also for measuringcell functions. According to the cellular responses, this re-view discusses electrochemical biosensors based on (1) cel-lular activity and function, (2) cellular barrier behavior and(3) recording/stimulation of electric potential of electrogeniccells. In contrast to the first two types, where non-neuralcells are usually used, neural cells, which have unique ad-vantage of high specificity through receptor-interaction [10],are incorporated for the third type of biosensors. In view ofthe fact that cell immobilization constitutes an important firststep toward the fabrication of sensors, a brief discussion inthis regard is also presented.

2. CELL IMMOBILIZATION

As biocompatible matrices for preparation of biosensors[18], a successful matrix for cell immobilization can stably



2 Current Medicinal Chemistry, 2008 Vol. 15, No. 14 Ding et al.

integrate cells and efficiently maintain their functionality.Identification of artificial surfaces is critical for preparationof cell biosensors. Whole cell immobilization on artificialsurfaces often mimics what occurs naturally. For cells grow-ing on a surface, the surface properties, such as roughness,hydrophobicity, topography, positive/negative charges, sur-face chemistry and specific protein or cell-surface interac-tions, can affect the activity and adhesion of cells to theartificial surfaces [19,20]. Recently, biocompatible nanopar-ticles have been extensively used to construct non-toxic

biomimetic interface for immobiliza tion of living cells[21,22], which allows adsorption of cells by the electrostaticforce or weak interaction between the nanoparticles and cellmembrane. Sol-gel matrices and natural polymers can entrapcells on the electrode surface. The major problem of cellentrapment is the instability of cell immobilization duringcontinuous use and the additional diffusion barrier resultingfrom the entrapped materials, which can be minimized byincreasing the porosity of the matrix. Although covalent

binding usually leads to the decrease of cell viability when

the cells are exposed to reactive groups and harsh reactionconditions, the surface modification of supports with func-tional groups such as amino groups, hydroxyl group andcarboxylic side chain has been developed for proper anchor-age of viable cells to improve cell attachment [23,24]. Thegrafted monolayer film with excellent hydrophilicity and

biocompatibility can provide an appropriate biomimeticinterface for cell adhesion, proliferation and electrochemicalstudy [23].

3. ELECTROCHEMICAL BIOSENSORS BASED ONCELLULAR ACTIVITY AND FUNCTION

3.1. Voltammetric Behaviours of Immobilized Cells A living cell can be properly described as an electro-

chemical dynamic system [25]. Electron generation andcharge transfer exist in all living cells due to the redox reac-tions and the changes of ionic composition and concentrationin life processes [26], which have been used to characterizethe viability of cells in homogeneous solution.

The tumor cells attached to a modified electrode can ex-hibit an irreversible voltammetric response around +0.8 V inthe first scan (solid curve in Fig. ( 1)) [27], which is related tothe oxidation of guanine [27-30]. However, at a bare carbon

paste or glassy carbon electrode, the tumor cells do not showany detectable electrochemical signal, which demonstratethat the nanomaterials such as gold nanoparticles, carbonnanotubes, or carbon nanconofiber can significantly rein-force the response [27,28,30]. No corresponding reductionwave was observed in the reverse scan (from positive poten-tial to negative potential). This oxidation peak has been em-

ployed for developing a simple and rapid method to e lectro-chemically investigate the exogenous effect, and a promisingapproach for an electrochemical anti-tumor drug sensitivitytest [27,29]. The optimal exogenous factors that affect cellviability, such as solution pH, ionic strength, temperatureand components, are just consistent with cell growth condi-tions in culture. Comparing with conventional methods suchas the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium

bromide (MTT) assay, the developed drug sensitivity testexhibits good performance characteristics, such as high sen-sitivity, desirable accuracy, low cost and simplified proce-dures.

Fig. (1). Cyclic voltammograms of AsPC-1 cells immobilized ongold nanoparticles modified carbon paste electrode in 10 mM pH7.4 PBS at scan rate of 80 mV/s using a conventional three-electrode system with gold as working, platinum wire as auxiliary,and saturated calomel electrode as reference electrodes. The solidand dotted lines represent first and second scan, respectively, andthe direction of initial scan is positive.

Glossary

Recombinant cells: the sensing elements prepared by fusing a reporter gene to the cells for eliciting a response to an analyte.

Potentiometry: involving potential determination between indicator and reference electrodes at zero current under equilibrium conditions for monitoringthe accumulation of charge due to selective binding at the indicator electrode surface.

Amperometry: measuring the current resulting from the electrochemical oxidation or reduction of an electroactive species at a constant applied potential.

Conductometriy: measuring the change in the conductance of the biological component arising between a pair of metal electrodes.

DOX electrochemical sensor: Comprised of 96 sets of three-electrode configuration commensurate with the dimensions of conventional 96-well plate inwhich the cells are cultured. During cell respiration, oxygen is consumed, and the measured current arising from the reduction of oxygen at negative poten-

tials reduces to a threshold value. The time to achieve this threshold current is proportional to the initial cell concentration.Impedance spectroscopy: a transient electrochemical method by applying a small amplitude of perturbing sinusoidal voltage to the electrochemical celland measuring the resulting response, including non-Faradaic and Faradaic impedance. Non-Faradaic impedance is performed in the absence of any redox

probe, while Faradaic measurement requires redox-active species .

Action potential: the sequential, electrochemical polarization and depolarisation that travels across the membrane of a nerve cell in response to stimulation.

Microfluidics: fluidics in structures on micron and smaller-length scales, resulting in low turbulence, with laminar flows.

Fi . ( 1) containsthe dotted and solid



Cell-Based Electrochemical Biosensors Current Medicinal Chemistry, 2008 Vol. 15, No. 14

3.2. Electrochemical Biosensors Based on Cellular Func-tion

Metabolism in cells, which depends on the intracellular physiological state and extracellular environment, leads tothe changes of pH, oxygen and ion concentrations. A varietyof cell-based electrochemical biosensors based on measure-ment of these changes have been fabricated [31-49]. Micro-organisms such as E coli , yeast, fungi and so on are the mostused recognition elements for this type of cell-based biosen-sor. However, recent research has focused on more complexsystems such as using animal cells or recombinant cells[7,32-34]. The detection of cellular metabolism can givedirect evidence of the activation of specific receptors, whichis valuable for drug screening and contains much more in-formation than just measurement of the recognizing/bindingevents [35,36]. Different electrochemical methods such as

potentiometry, amperometry or conductometry (see Glos-sary) have been used to perform the measurements (Fig. ( 2)).

A. Potentiometric Cell-Based Biosensors

Conventional potentiometric cell-based biosensors con-sist of an ion-selective electrode (ISE) or gas-sensing elec-trode (GSE) coated with a cell-immobilized layer [8]. Theimmobilized cells consume analyte to generate a potentialchange due to ion accumulation or depletion on electrodesurface [50]. This method requires a very stable referenceelectrode, which is a limitation of these transducers [8]. Awhole-cell potentiometric biosensor for screening of toxinshas been proposed by attaching endothelial cells to a K +-selective membrane, on which ion transport is almost com-

pletely inhibited by a confluent cell monolayer [32]. Whenthe biosensor is exposed to a specific class of compounds,the permeability of cell monolayer increases, and more K cation can penetrate through the attached monolayer, which

produces a potential response. Interestingly, this potenti-ometric biosensor can be easily adapted to small-scale appli-cations.

B. Amperometric Cell-Based Biosensors

Amperometric cell biosensors have been widely devel-oped for determination of oxygen, glucose, lactose, and soon (Table 1) [35-48]. Most of them use oxygen sensors todetect cell respiration [37-40,42-44,46], which offer theadvantage of continuous monitoring. Besides oxygen, anelectroactive product or intermediate produced by specificenzymatic reaction of cells can also be detected. Amongthese analytes, nitric oxide (NO), adenosine triphosphate(ATP, 1, Scheme 1), glutamate, histamine ( 2, Scheme 1) andcatecholamine family (dopamine ( 3, Scheme 1), norepineph-rine ( 4, Scheme 1), and epinephrine ( 5, Scheme 1)) are par-ticularly important, because they act as signals in many bio-logical systems. They can also probably be employed to

determine the effects of clinical drugs and measure extracel-lular stress factors in various types of cells. For example, anendothelial cellular biosensing system has been constructed

by immobilizing human umbilica l vein endothelial cells(HUVEC) on an electrode to detect nitric oxide, an indicatorof blood vessel relaxation, through differential pulse volt-ammetry (DPV) [35]. Although the promising system can beused for assessing blood pressure-controlling drugs by regu-lating NO release, such as acetylcholine chloride (AcChCl),

Fig. (2). Scheme of cell-based electrochemical biosensors based on cellular functions. A1: amperometric biosensor based on detection ofoxygen; A2: amperometric biosensor based on detection of electroactive species produced by enzymatic reaction.




NOC 7 (a NO donor) and N G-monomethyl-L-arginine (L- NMMA), i t has not taken into account the diffusion of traceanalyte from cells to sensor surface. Furthermore, the mate-

rials modified on electrodes must be configured specificallyto ensure both molecular selectivity and molecular trans-duceability for the given mammalian cell culture media.

A common disadvantage of amperometric cell biosensorsis the lack of selectivity. This problem can be solved byinhibition or suppression of undesired transport mechanismsor metabolic pathways and coupling of enzymes with immo-

bilized microbial cells to form a hybrid biosensor. For example, alcohol oxidase can be introduced to C. tropicalis yeastcells by culturing in a culture medium containing high con-centration of ethanol [42].

Another shortcoming of cell-based sensors is long re-sponse time. By virtue of the development of genetic engi-neering, the dynamics of the microbial biosensing can be

significantly improved by engineering the cells to anchor thedesired enzyme into the periplasmic space [51]. However,when novel receptors are introduced into cells, it is importantto ensure that the receptor does not interfere with normalcellular processes. Genetic engineering also offers an effec-tive way to adjust the interface between cells and electrodes.A strategy to control the interface between Chinese hamsterovary (CHO) cells and the electrode by genetic engineeringfor converting activity of the expressed enzyme on cell sur-face into an electrical signal has been reported [34]. Con-struction of such electrochemically detectable interface can

be extended for responding to certain factors without the

Table 1. Amperometric Microbial Biosensors

Target Microbial Limit of detection Type Ref.

Fe 2+ Acidithiobacillus ferrooxidans 900 nM A1 [39]

p -nitrophenol Moraxella sp. 0.1 μ M A1 [40]

p -nitrophenol Moraxella sp. 20 nM A2 [41]

ethanolCandida

tropicalis0.5 mM A1 [42]

choline Arthrobacter

globiformis80 nM A1 [43]

L-lysineSaccharomyces

cerevisiae1.0 μ M A1 [44]

1,3-propanediolGluconobacter

oxydans0.15-0.42 mg/L A2 [45]

p -nitrophenyl-substitutedorganophosphates

Moraxella sp. 0.1 μ M A1 [46]

NH 2

OH

OH

NH 2

OH

OH

HO

HN

OH

OH

HO

CH 3

N

NH

NH 2

N

N N

N

NH 2

O

OHOH

OPO

O-

O

PO

O-

O

P-O

O-

O

1 , Adenosine triphosphate (ATP) 2 , Histamine

3 , Dopamine 4 , Norepinephrine 5 , Epinephrine Scheme (1). Adenosine triphosphate ( 1), glutamate, histamine ( 2), and catecholamine family, including dopamine ( 3), norepinephrine ( 4)and epinephrine ( 5 ).




necessity of the optical system, and can serve as the basis ofa compact biosensor.

The dissolved oxygen (DOX) sensor (see Glossary) cansimultaneously determine dissolved oxygen with 96 sets ofelectrodes for studying bacteria, cells and their interactionwith analytes and continuous real-time monitoring of bacte-ria activity [14,37,38]. Compared with methods based onfluorescent microscopy, flow cytometry and spectropho-tometry, the major advantage of DOX is that it is suitable forcontinuous monitoring. By integration with pattern recogni-tion techniques, this system can be used for the monitoringof bacteria activity upon exposure to antibiotics [38]. It pro-vides a simple and high throughput system in a noninvasivemanner without any preparation step for development of thenext generation of analytical devices.

Nowadays, the use of the miniaturized sensor is of greatdemand to minimize the size of the measurement chamber,thus one can work with small numbers of cells. An integrateddistance-control NO sensor has been constructed to performon-line Ca 2+ channel-related drug screening with transformedhuman umbilical vein endothelial cells (T-HUVEC) placed

at a known distance from the electrodes [48]. This automatedsystem features a unique design of an electrode set for dis-tance control, and increases sample throughput. However,the transfer of the electrode set among wells prolongs theassay time, in this regard, microelectrode arrays exhibit in-comparable superiority. A microelectrode array biochip fordrug screening purposes has been developed to study theinfluence of reserpine, nomifensine and L-3,4-dihydroxyphenylalanine (L-dopa) on quantitative catechola-mine release from PC12 cells [36]. This biochip focuses onquantifying release from multiple cells rather than resolvingindividual events. It can obtain high sensitivity throughminimization of the distance, and consequently the solutionvolume. Meanwhile, nanotechnology has also enormously

contributed to the development of miniaturized amperomet-ric cell-based biosensor, in which cells are integrated into ananosystem to perform sensing function. An innovativenano-biochip, which contained an array of nanovolume elec-trochemical cells, was presented [7]. In the presence of atoxin, the E. coli integrated into the chip produced -galactosidase, leading to an enzymatic reaction upon addi-tion of a substrate. The product of the enzymatic reactionwas then oxidized at an electrode. The chamber array withnL volume enabled simultaneous measurement of eight sam-

ples, allowed minimum interaction with the water flow andresulted in increased speed and sensitivity.

C. Conductometric Cell-Based Biosensors

Conductometric cell-based biosensors are based on detec-tion of changes in ionic species due to cell-catalyzed reac-tions. Even though the detection of solution conductance isnon-specific, conductance measurements are extremely sen-

sitive [8]. Microalgae were used as bi-enzymatic receptor todevelop a conductometric biosensor for monitoring the ac-tivities of two kinds of enzymes [49]. This biosensor over-came the disadvantage of using two kinds of pure enzymes,which faced the stability and commercial problems, and thus

provided a convenient way for multi-detection array.

4. BIOSENSORS BASED ON BARRIER BEHAVIOROF ATTACHED CELLS

The cell-to-electrode interactions have showed importantperspective in the development of biosensors and prostheticdevices [4]. Cells have excellent insulating properties and canaffect the local ionic environment at electrode/solution inter-

face, which leads to change of impedance [11,52-58]. Thuselectrochemical impedance spectroscopic (EIS) technique (seeGlossary) can be used to monitor the biological status ofcells, including cellular viability and morphology, cell num-

ber, and adhesion, proliferation and apoptosis of cells onsurface, and has shown great advantages for cell-based gen-eral screening of medicinal compounds in drug development[53,56-58]. This technique is inexpensive and relatively easyto perform in comparison with surface plasmon resonanceand quartz crystal microbalance techniques. Theoretically, itcan dynamically measure cellular movement in nanoscalewith better resolution than conventional optical methods [5].Two categories of EIS have been used for detection of cell-to-electrode interactions (Fig. ( 3)), leading to two kinds of cell-

based EIS b iosensors. A common disadvantage of cell-basedimpedance techniques is that the response measured is thetotal change produced by a number of cells.

4.1. Non-Faradaic EIS Technique

Different set-ups have been designed for impedancemeasurement, in all of which impedance is measured as afunction of time and changed environment [10]. Electric cell-substrate impedance sensing (ECIS) is the most popular non-

Fig. (3). Schemes of cell-based non-Faradaic ( a ) and Faradaic ( b ) impedance spectroscopic (non-Faradaic EIS and Faradaic EIS) biosensors based on detection of barrier behavior of attached cells.




Faradaic EIS technique [55,56]. Their applications includestudy of cell-cell contact, cytotoxic effect of toxin, and drugdiscovery. However, ECIS suffers from issues of insufficientcell coverage, leading to low sensitivity and uneven distribu-tion of cells between wells, which hampers their usage inhigh-throughput screening in drug discovery. Many groupshave been striving to improve the performance of ECIS. Animportant achievement is the invent of real-time cell elec-

tronic sensing (RT-CES), which can improve the sensitivityand the well-to-well distribution by increasing the coveringarea [53,57]. By constructing a set of “circle-on-line” micro-electronic sensors on the bottom of wells, 80% of the surfacearea can be covered, which allows more sensitive quantita-tive detection [53]. Furthermore, the RT-CES system hasraised cell impedance measurements to the heights nowserving as the basis of high throughput tools for investiga-tions of drug development and toxicity prediction. Severalresearches have demonstrated the capability of this system toobtain dynamic and concentration-dependent effects of toxin,drug and other chemicals on cellular status [53,57,59]. Forexample, the effects of antipyrene, trichlorfon, dimethylformamide, cadmium (II), p-phenylene diamine, propranolol,

ibuprofen, nalidixic acid, salicylic acid and glycerol onmouse fibroblasts have been studied using this system [59].

The second effort is to develop different set-ups that al-low the simultaneous visualization and measurement of

barrier function of cells. For this purpose, indium tin oxide(ITO), which combines electrical conductivity and opticaltransparency, is an attractive alternative to gold as a micro-impedance biosensor. Using an ITO–Si 3 N4 microelectrodesarray, the effect of cytochalasin D on porcine pulmonaryartery endothelial cell (PPAEC) has been monitored, thus

providing a more reliable method for measurement of cellu-lar response to drugs simultaneously by electrochemical andoptical approaches [58].

With the ability to manipulate cells and control mechani-cal, electrical and biochemical parameters down to thenanometer scale, micro-fabrication instrumentation has al-

lowed electrical impedance techniques to be used for study-ing the anchorage-dependent cells cultured on microelec-trode arrays (MEAs) [53,60]. The disadvantage is the in-creased electrode impedance caused by the electrode polari-zation as the electrode size decreases [61]. To avoid polari-zation, a microhole-based chip can be utilized to realize highcurrent density nearby the hole but no electric double layerfor sensitive impedance measurement, which can examine

the cytotoxicity of dimethlysulfoxide on L929 cells [61].This approach provides an effective way for on-line record-ing of the impedance of even a single cell under specific

physical/chemical environments in real time without chemi-cal markers.

4.2. Faradaic EIS Technique

Faradaic EIS is a powerful tool for analyzing interfacial properties of modified electrodes during biorecognitionevents. The cell membranes (thickness 5-10 nm) show thecapacitance and resistance of 0.5-1.3 μ F cm 2 and 10 2-10 5 cm 2, respectively [62]. As a result, when cells attach to anelectrode, a barrier hindering the access of the redox probe to

the electrode surface is produced, thereby, resulting in anincrease in the electron transfer resistance ( Ret). The increasemagnitude of electron-transfer resistance is related to thenumber of immobilized cells. This behavior has been used todetect Escherichia coli O157:H7 concentration [52]. Uponcell adhesion the Ret value is proportional to the logarithmicvalue of the cell concentration ranging from 4.36 10 5 to4.36 10 8 cfu/ml, which is comparable to other label-freeimmunosensors. Based on this detection principle, severalFaradaic EIS sensors for cell number have been developed[3,28,30,52,63-68], and the fabrication characteristics aresummarized in Table 2. Some of these cell-based sensors are

prepared by immobilizing antibody on electrode surface tocapture cells [52,63-65]. A major problem associated with

anchoring cells by immunoreaction is the low capture effi-ciency (CE). As a result, optimal utilization of the functionalsurface area (where attached cells are detected) of an elec-

Table 2. Characteristics of Cell-Based Faradaic EIS Biosensors

Modified electrode Cells Linear range Ref.

Ab/Au IDAM a Escherichia coli O157:H7 10 4-10 7 cfu/ml [59]

Ab/epoxysilane/ITO a Escherichia coli O157:H7 6 10 4-6 10 7 cells/ml [60]

[61]

SAM/Au b S. cerevisiae 10 2-10 8 cfu/ml [62]

Ab/ITO IDAM a Escherichia coli O157:H7 4.36 10 5-4.36 108 cfu/ml [49]

Ab-Magnetic NPs/Au IDAM a,c Escherichia coli O157:H7 7.4 10 4-7.4 10 7 cfu/ml [63]

Ab-biotin/neutravidin/

biotin-SAM/Au a,b Escherichia coli 10-10 3 cfu/mL [3]

Ab/epoxysilane/GCE a K562A 5 10 4-1 10 7 cells/ml [29]

Polyaniline/SPCE K562 10 4-10 7 cells/ml [64]

GNPs embedded chitosan gel/GCE K562 1.34 104-1.34 10 8 cells/ml [28]

CNF-chitosan/GCE K562 5 10 3-5 10 7 cells/ml [31]

aAb refers to antibody. bSAM refers to self-assembly monolayer.c NPs refers to nanoparticles.




trode is of vital importance for detection [67]. Magneticnanoparticle-antibody conjugates (MNAC) can be utilized toconcentrate separated cells into a small volume, and furtherin the active layer of interdigitated array microelectrode(IDAM), at which cells are then detected [67]. Another prob-lem of Faradaic impedance technique is that the recognitioncomplex often leads to minute and usually undetectablechanges in the interface impedance, thus reducing the repro-

ducibility. In the aspect of data processing, it is unclear whythe value of R et is proportional to the logarithmic value of thecell concentration, rather than linear relationship, which canalso be observed in protein and DNA impedance sensor [69].

Faradaic EIS measurement can be used to detect bothcells in suspensions and the changes on the electrode surfaceupon the proliferation of living tumor cells [23,28,68]. Asshown in Fig. ( 4) [23], a significant change of the diameterof the semicircular part of spectra, which represents the elec-tron transfer resistance, can be observed when cells are pro-liferating on the modified surface. With the increasing ofculture time, the electron transfer resistance increases. Aftera culture time of 120 hours, the drastically increasing resis-tance is observed, which is related to the apoptosis of cells(inset in Fig. ( 4)). This electrochemical strategy is an effec-tive and simple way for continuous online monitoring of cell

proliferation and apoptosis when disposable electrodes areused [68]. Further development of this method may be com-

bined with novel type of disposable electrodes, for example ,ITO chips, and more suitable biomaterials for cell adhesion,and it can also be extended for studying drug effects and

biocompatibility of materials on elec trodes.

Fig. (4). Electrochemical impedance spectrum measurements ofcells adhered at ( a ) bare and ( b ) zwi-film modified electrodes, andK562 cells proliferated on zwi-film modified electrode after cellincubation for ( c) 24, ( d ) 48, ( e) 72, ( f ) 96, ( g) 120 and ( h ) 140 hrecorded using a conventional three-electrode system with modifiedelectrode as working, platinum wire as auxiliary, and saturatedcalomel electrode as reference electrodes with 5 mV amplitude ofthe applied sine wave potential in the frequency range of 10 -2-10 6 Hz,. Inset: plot of electron-transfer resistance vs proliferation timeof K562 on zwi-film modified electrodes.

5. BIOSENSORS BASED ON THE RECORD-ING/STIMULATION OF CELLULAR ELECTRICPOTENTIAL

Electrogenic cells (neural cells, heart muscle cells, pan-creas beta cells) or a neural cell network grown in culture onmicroelectrode arrays or on field effect transistors are verysensitive to neuroactive compounds added to the culture

medium, which can be measured by the change of membrane potential during an action potential (see G lossary).

The voltage clamp and patch clamp techniques are domi-nant techniques for monitoring cellular electrophysiology[70-76], particularly the study of nicotinic cholinergic recep-tor, which is a focus of interest for drug discovery [71,72].However, these techniques are basically invasive methods,and the manipulation procedure needs high skill. In contrast,the cell-based biosensors using a micro-sensor array providea non-invasive way for recording the action potential in realtime, which can reach a single cell and cell network. In thisfield, microelectrode array (MEA) cell-based biosensor[77,78], and the light-addressable potentiometric sensor(LAPS) [79,80] are the most popular detection methods.

5.1. MEA-Based Electrochemical Biosensor

The integration of a fixed microelectrode array with theelectrodes on a silicon or glass substrate and a cell culturechamber has been demonstrated to allow for long-term, non-invasive and multisite extracellular potential recording fromelectrogenic cells, and is used for biosensing [77] (Fig. ( 5a ))However, this technique suffers from the difficulty to pre-cisely control the cell density and the interaction betweencells and electrodes, which reduces the reliability of meas-urement. Microfabricated systems create new opportunitiesto solve the problem by integration of extracellular matrix(ECM) surface patterning technique with microfluidic chan-nels (see Glossary). The control of surface makes it possibleto reproducibly place the cells at precise points [13], andmicrofluidics can not only deliver soluble factors, but alsohelp to control neuronal connectivity. More importantly,compared with cells immobilized on electrodes as sensoryelements, cells in microfluidics avoid the impact to cellular

physiological activity resulted from the immobilization ofcells [81]. By combination of planar microelectrode arrayswith microfluidic devices, extracellular electrical signals ofvarious type of primary neural can successfully be recorded,and this bioelectrical activity can be presented for severalweeks [78]. In addition, the MEA-based cell biosensor canonly detect a limit number of discrete active sites, ascribingto the fabrication of electrode arrays. This shortcoming has

been solved by utilization of light-addressable potentiometricsensor [79].

5.2. Light-Addressable Potentiometric Sensor-BasedBiosensor

Cells are active and dynamic entities, not confined to permanent localization on a device to allow easy interroga-tion. Light-addressable potentiometric sensor is a commonlyused semiconductor chip for developing cell-semiconductorhybrid system, by which cells can be randomly cultured andthe change of cellular electric potential can be addressably




(90713015) from the National Natural Science Foundation ofChina for related research.

ABBREVIATIONS

AcChCl = Acetylcholine chloride

CE = Capture efficiency

CHO = Chinese hamster ovaryCNF = Carbon nanofiber

DOX = Dissolved oxygen

DPV = Differential pulse voltammetry

ECIS = Electric cell-substrate impedance sensing

ECM = Extracellular matrix

EIS = Electrochemical impedance spectroscopic

GSE = Gas-sensing electrode

HUVEC = Human umbilical vein endothelial cells

IDAM = Interdigitated array microelectrode

ISE = Ion-selective electrode

ITO = Indium tin oxide

LAPS = Light-addressable potentiometric sensor

L-dopa = L-3,4-dihydroxyphenylalanine

L-NMMA = N G-monomethyl-L-arginine

MEAs = Microelectrode arrays

MNAC = Magnetic nanoparticle-antibody conjugates

MTT = 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NEAs = Nanoelectrode arrays

RT-CES = Real-time cell electronic sensing

PPAEC = Porcine pulmonary artery endothelial cell

Ret = Electron-transfer resistance

T-HUVEC = Transformed human umbilical vein endo-thelial cells

REFERENCES

[1] Wilson, G.S.; Gifford, R. Biosensors for real-time in vivo meas-urements. Biosens. Bioelectron. , 2005 , 20 , 2388-403.

[2] Bakker, E.; Qin, Y. Electrochemical sensors. Anal. Chem. , 2006 ,78 , 3965-84.

[3] Maalouf, R.; Fournier-Wirth, C.; Coste, J.; Chebib, H.; Saïkali, Y.;Vittori, L.; Errachid, A.; Cloarec, J.-P.; Martelet, C.; Jaffrezic-Renault, N. Label-free detection of bacteria by electrochemical im-

pedance spectroscopy: comparison to surface plasmon resonance. Anal. Chem. , 2007 , 79 , 4879-86.

[4] Slaughter, G.E.; Bieberich, E.; Wnek, G.E.; Wynne, K.J.;Guiseppi-Elie, A. Improving neuron-to-electrode surface attach-ment via alkanethiol self-assembly: an alternating current imped-ance study. Langmuir , 2004 , 20 , 7189-200.

[5] Pancrazio, J.J.; Whelan, J.P.; Borkholder, D.A.; Ma, W.; Stenger,D.A. Development and application of cell-based biosensors. Ann.

Biomed. Eng. , 1999 , 27 , 697-711.[6] Wang, P.; Xu, G.X.; Qin, L.F.; Xu, Y.; Li, Y.; Li, R. Cell-based

biosensors and its application in biomedicine. Sensor. Actuat. B-Chem. , 2005 , 108 , 576-84.

[7] Popovtzer, R.; Neufeld, T.; Biran, D.; Ron, E.Z.; Rishpon, J.;Shacham-Diamand, Y. Novel integrated electrochemical nano-

biochip for toxicity detection in water. Nano Lett. , 2005 , 5, 1023-7.[8] Lei, Y.; Chen, W.; Mulchandani, A. Microbial biosensors. Anal

Chim. Acta , 2006 , 568 , 200-10.[9] Daunert, S.; Barrett, G.; Feliciano, J.S.; Shetty, R.S.; Shrestha, S.;

Smith-Spencer, W. Genetically engineered whole-cell sensing sys-tems: coupling biological recognition with reporter genes. Chem

Rev. , 2000 , 100 , 2705-38.[10] Ziegler, C. Cell-based biosensors. Fresenius J. Anal. Chem. , 2000

366 , 552-9.[11] Chen, Y.; Zhang, J.; Wang, Y.; Zhang, L.; Julien, R.; Tang, K.;

Balasubramanian, N. Real-time monitoring approach: assessmentof effects of antibodies on the adhesion of NCI-H460 cancer cellsto the extracellular matrix. Biosens. Bioelectron. , 2008 , 23 , 1390-6.

[12] Bashir, R. BioMEMS: state-of-the-art in detection, opportunitiesand prospects. Adv. Drug Deliver. Rev. , 2004 , 56 , 1565-86.

[13] El-Ali, J.; Sorger, P.K.; Jensen, K.F. Cells on chips. Nature , 2006

442 , 403-11.[14] Andreescu, S.; Sadik, O.A. Advanced electrochemical sensors for

cell cancer monitoring. Methods , 2005 , 37 , 84-93.[15] Haque, A.U.; Chatni, M.R.; Li, G.; Porterfield, D.M. Biochips and

other microtechnologies for physiomics. Expert Rev. Proteomics2007 , 4, 553-63.

[16] Rodriguez-Mozaz, S.; de Alda, M.J.L.; Barceló, D. Biosensor asuseful tools for environmental analysis and monitoring. Anal

Bioanal. Chem. , 2006 , 386 , 1025-41.[17] Porterfield, D.M. Measuring metabolism and biophysical flux in

the tissue, cellular and sub-cellular domains: recent developmentsin self-referencing amperometry for physiological sensing. Biosens

Bioelectron. , 2007 , 22 , 1186-96.[18] Xu, Z.A.; Chen, X.; Dong, S.J. Electrochemical biosensors based

on advanced bioimmobilization matrices. Trends Anal. Chem.2006 , 25 , 899-908.

[19] Lee, S.J.; Choi, J.S.; Park, K.S.; Khang, G.; Lee, Y.M.; Lee, H.B.Response of MG63 osteoblast-like cells onto polycarbonate mem-

brane surfaces with different micropore sizes. Biomaterials , 2004

25 , 4699-707.[20] Claase, M.B.; Riekerink, M.B.O.; de Bruijn, J.D.; Grijpma, D.W.;

Engbers, G.H.M.; Feijen, J. Enhanced bone marrow stromal celladhesion and growth on segmented poly(ether ester)s based on

poly(ethylene oxide) and poly(butylenes terephthalate). Biomacro-molecules , 2003 , 4, 57-63.

[21] Gu, H.Y.; Chen, Z.; Sa, R.X.; Yuan, S.S.; Chen, H.Y.; Ding, Y.T.;Yu, A.M. The immobilization of hepatocytes on 24 nm-sized goldcolloid for enhanced hepatocytes proliferation. Biomaterials , 2004

25 , 3445-51.[22] Du, D.; Ju, H.X.; Zhang, X.J.; Chen, J.; Cai, J.; Chen, H.Y. Electro-

chemical immunoassay of p-glycoprotein on cell membrane by cellimmobilization on gold nanoparticles modified methoxysilyl-terminated butyrylchitosan matrix. Biochemistry , 2005 , 44 , 11539-45.

[23] Du, D.; Cai, J.; Ju, H.X.; Yan, F.; Chen, J.; Jiang, X.Q.; Chen, H.Y.Construction of a biomimetic zwitterionic interface for monitoringcell proliferation. Langmuir , 2005 , 21 , 8394-9.

[24] Curran, J.M.; Chen, R.; Hunt, J.A. The guidance of human mesen-chymal stem cell differentiation in vitro by controlled modifica-tions to the cell substrate. Biomaterials , 2006 , 27 , 4783-93.

[25] Bery, M.N.; Grivell, M.B. In Bioelectrochemistry of Cells andTissues ; Walz, D.; Berg, H.; Milazzo G., Ed.; Birkhauser Verlag:Basel, Switzerland, 1995 , pp. 134-58.

[26] Nonner, W.; Eisenberg, B. Electrodiffusion in ionic channels of biological membranes. J. Mol. Liq. , 2000 , 87 , 149-62.

[27] Chen, J.; Du, D.; Yan, F.; Ju, H.X.; Lian, H.Z. Electrochemicalanti-tumor drug sensitivity test for leukemia K562 cells at a carbonnanotubes modified electrode. Chem-Eur. J. , 2005 , 11 , 1467-72.

[28] Ding, L.; Hao, C.; Xue, Y.D.; Ju, H.X. A bio-inspired support ofgold nanoparticles-chitosan nanocomposites gel for immobilizationand electrochemical study of K562 Leukemia cells. Biomacro-molecules , 2007 , 8 , 1341-6.

[29] Yan, F.; Chen, J.; Ju, H.X. Immobilization and electrochemical behavior of gold nanoparticles modified leukemia K562 cells andapplication in drug sensitivity test. Electrochem. Commun. , 2007

9, 293-8.




[30] Hao, C.; Ding, L.; Zhang, X.J.; Ju, H.X. Biocompatible conductivearchitecture of carbon nanofiber-doped chitosan prepared with con-trollable electrodeposition for cytosensing. Anal. Chem. , 2007 , 79 ,4442-7.

[31] Ivnitski, D.; Abdel-Hamid, I.; Atanasov, P.; Wilkins, E.; Stricker,S. Application of electrochemical biosensors for detection of food pathogenic bacteria. Electroanalysis , 2000 , 12 , 317-25.

[32] May, K.M.L.; Wang, Y.; Bachas, L.G.; Anderson, K.W. Develop-ment of a whole-cell-based biosensor for detecting histamine as amodel toxin. Anal. Chem. , 2004 , 76 , 4156-61.

[33] Neufeld, T.; Biran, D.; Popovtzer, R.; Erez, T.; Ron, E.Z.; Rishpon,J. Genetically engineered pfabA pfabR bacteria: an electrochemicalwhole cell biosensor for detection of water toxicity. Anal. Chem. ,2006 , 78 4952-6.

[34] Collier, J.H.; Mrksich, M. Engineering a biospecific communica-tion pathway between cells and electrodes.Proc. Natl. Acad. Sci. U.S.A. , 2006 , 103 , 2021-5.

[35] Kamei, K.; Haruyama, T.; Mie, M.; Yanagida, Y.; Aizawa, M.;Kobatake, E. The construction of endothelial cellular biosensingsystem for the control of blood pressure drugs. Biosens. Bioelec-tron. , 2004 , 19 , 1121-4.

[36] Cui, H.F.; Ye, J.-S.; Chen, Y.; Chong, S.-C.; Sheu, F.-S. Microelec-trode array biochip: tool forin vitro drug screening based on thedetection of a drug effect on dopamine release from PC12 cells.

Anal. Chem. , 2006 , 78 , 6347-55.[37] Andreescu, S.; Sadik, O.A.; McGee, D.W. Autonomous multielec-

trode system for monitoring the interactions of isoflavonoids withlung cancer cells. Anal. Chem. , 2004 , 76 , 2321-30.

[38] Karasinski, J.; Andreescu, S.; Sadik, O.A. Multiarray sensors with pattern recognition for the detection, classification, and differentia-tion of bacteria at subspecies and strain levels. Anal. Chem. , 2005 ,77 , 7941-9.

[39] Zlatev, R.; Magnin, J.-P.; Ozil, P.; Stoytcheva, M. Bacterial sensors base on acidithiobacillus ferrooxidans Part I. Fe2+ and S2O3

2- de-termination. Biosens. Bioelectron. , 2006 , 21 , 1493-500.

[40] Mulchandani, P.; Lei, Y.; Chen, W.; Wang, J.; Mulchandani, A.Microbial biosensor for p-nitrophenol using Moraxella sp. Anal. Chim. Acta , 2002 , 470 , 79-86.

[41] Mulchandani, P.; Hangarter, C.M.; Lei, Y.; Chen, W.; Mulchan-dani, A. Amperometric microbial biosensor for p-nitrophenol using

Moraxella sp.-modified carbon paste electrode. Biosens. Bioelec-tron. , 2005 , 21 , 523-7.

[42] Akyilmaz, E.; Dinckaya, E. An amperometric microbial biosensor

development based onCandida tropicalis yeast cells for sensitivedetermination of ethanol. Biosens. Bioelectron. , 2005 , 20 , 1263-9.[43] Stoytcheva, M.; Zlatev, R.; Valdez, B.; Magnin, J.-P.; Velkova, Z.

Electrochemical sensor based on Arthrobacter globiformis for cho-linesterase activity determination. Biosens. Bioelectron. , 2006 , 22 ,1-9.

[44] Akyilmaz, E.; Erdoan, A.; Öztürk, R.; Ya a, . Sensitive determi-nation of l-lysine with a new amperometric microbial biosensor based on Saccharomyces cerevisia e yeast cells. Biosens. Bioelec-tron. , 2007 , 22 , 1055-60.

[45] Katrlík, J.; Vo tiar, I.; ef ovi ová, J.; Tká , J.; Mastihuba, V.;Valach, M.; tefuca, V.; Gemeiner, P. A novel microbial biosensor based on cells ofGluconobacter oxydans for the selective determi-nation of 1,3-propanediol in the presence of glycerol and its appli-cation to bioprocess monitoring. Anal. Bioanal. Chem. , 2007 , 388 ,287-95.

[46] Mulchandani, P.; Chen, W.; Mulchandani, A. Microbial biosensorfor direct determination of nitrophenyl-substituted organophos- phate nerve agents using genetically engineered Moraxella sp. Anal. Chim. Acta , 2006 , 568 , 217-21.

[47] Motrescu, E.R.; Otto, A.M.; Brischwein, M.; Zahler, S.; Wolf, B.Dynamic analysis of metabolic effects of chloroacetaldehyde andcytochalasin B on tumor cells using bioelectronic sensor chips. J. Cancer Res. Clin. Oncol. , 2005 , 131 , 683-91.

[48] Borgmann, S.; Radtke, I.; Erichsen, T.; Blöchl, A.; Heumann, R.;Schuhmann, W. Electrochemical high-content screening of nitricoxide release from endothelial cells.ChemBioChem , 2006 , 7 , 662-8.

[49] Chouteau, C.; Dzyadevych, S.; Durrieu, C.; Chovelon, J.-M. A bi-enzymatic whole cell conductometric biosensor for heavy metalions and pesticides detection in water samples. Biosens. Bioelec-

tron. , 2005 , 21 , 273-81.[50] Simonian, A.L.; Rainina, E.I.; Wild, J.R. In Enzyme and Microbial

Biosensors: Techniques and Protocols ; Mulchandani, A.; RogerK.R., Ed.; Humana Press, Totowa: New Jersey,1998 , pp. 237-48.

[51] Tká , J.; tefuca, V.; Gemeiner, P. In Applications of Cell Immobi-lization Biotechnology. Focus on Biotechnology ; Nedovi , V.; Willaert, R., Ed.; Springer, Dordrecht: the Netherlands,2005 , pp. 54966.

[52] Yang, L.J.; Li, Y.B.; Erf, G.F. Interdigitated array microelectro based electrochemical impedance immunosensor for detection Escherichia coli O157:H7. Anal. Chem. , 2004 , 76 , 1107-13.

[53] Xing, J.Z.; Zhu, L.J.; Jackson, J.A.; Gabos, S.; Sun, X.-J.; WaX.-B.; Xu, X. Dynamic monitoring of cytotoxicity on microetronic sensors.Chem. Res. Toxicol. , 2005 , 18 , 154-61.

[54] Tlili, C.; Reybier, K.; Géloën, A.; Ponsonnet, L.; Martelet, Ouada, H.B.; Lagarde, M.; Jaffrezic-Renault, N. Fibroblast celsensing bioelement for glucose detection by impedance spectcopy. Anal. Chem. , 2003 , 75 , 3340-4.

[55] Giaever, I.; Keese, C.R. A morphological biosensor for mammian-cells. Nature , 1993 , 366 , 591-2.

[56] Xiao, C.D.; Lachance, B.; Sunahara, G.; Luong, J.H.T. Assessmof cytotoxicity using electric cell-substrate impedance sensconcentration and time response function approach. Anal. Chem.2002 , 74 , 5748-53.

[57] Yu, N.C.; Atienza, J.M.; Bernard, J.; Blanc, S.; Zhu, J.; WaX.B.; Xu, X.; Abassi, Y.A. Real-time monitoring of morphologchanges in living cells by electronic cell sensor arrays: an approto study G protein-coupled receptors. Anal. Chem. , 2006 , 78 , 3543.

[58] Choi, C.K.; English, A.E.; Jun, S.-I.; Kihm, K.D.; Rack, P.D. endothelial cell compatible biosensor fabricated using optically indium tin oxide silicon nitride electrodes. Biosens. Bioelectron.2007 , 22 , 2585-90.

[59] Xing, J.Z.; Zhu, L.; Gabos, S.; Xie, L. Microelectronic cell seassay for detection of cytotoxicity and prediction of acute toxicToxicol. in Vitro , 2006 , 20 , 995-1004.

[60] Mishra, N.N.; Retterer, S.; Zieziulewicz, T.J.; Isaacson, Szarowski, D.; Mousseau, D.E.; Lawrence, D.A.; Turner, J.N. Ochip micro-biosensor for the detection of human CD4+ cells baseon AC impedance and optical analysis. Biosens. Bioelectron. , 2005

21 , 696-704.[61] Cho, S.; Thielecke, H. Micro hole-based cell chip with impeda

spectroscopy. Biosens. Bioelectron. , 2007 , 22 , 1764-8.

[62] Pethig, R. Dielectric and Electronic Properties of Biological Materials , J. Wiley& Sons, Chichester: New York,1979 .

[63] Radke, S.M.; Alocilja, E.C. A high density microelectrode a biosensor for detection of E. coli O157:H7. Biosens. Bioelectron.2005 , 20 , 1662-7.

[64] Ruan, C.M.; Yang, L.J.; Li, Y.B. Immunobiosensor chips detection of Escherichia coli O157:H7 using electrochemical i pedance spectroscopy. Anal. Chem. , 2002 , 74 , 4814-20.

[65] Yang, L.; Li, Y. AFM and impedance spectroscopy charactertion of the immobilization of antibodies on indium–tin oxide etrode through self-assembled monolayer of epoxysilane and thcapture of Escherichia coli O157:H7. Biosens. Bioelectron. , 2005

20 , 1407-16.[66] Chen, H.; Heng, C.K.; Puiu, P.D.; Zhou, X.D.; Lee, A.C.; L

T.M.; Tan, S.N. Detection ofsaccharomyces cerevisiae immoblized on self-assembled monolayer of alkanethiolate using elecchemical impedance spectroscopy. Anal. Chim. Acta , 2005 , 55452-9.

[67] Varshney, M.; Li, Y., Interdigitated array microelectrode baimpedance biosensor coupled with magnetic nanoparticle–antibconjugates for detection of Escherichia coli O157:H7 in food sam ples. Biosens. Bioelectron. , 2007 , 22 , 2408-14.

[68] Ding, L.; Du, D.; Wu, J.; Ju, H.X. A disposable impedance senfor electrochemical study and monitoring of adhesion and proliftion of K562 leukaemia cells. Electrochem. Commun. , 2007 , 9953-8.

[69] Daniels, J.S.; Pourmand, N. Label-free impedance biosensopportunities and challenges. Electroanalysis , 2007 , 19 , 1239-57.

[70] Pappas, T.C.; Wickramanyake, W.M.S.; Jan, E.; Motamedi, Brodwick, M.; Kotov, N.A. Nanoscale engineering of a cellularterface with semiconductor nanoparticle films for photoelec



Cell-Based Electrochemical Biosensors Current Medicinal Chemistry, 2008 Vol. 15, No. 14 1

stimulation of neurons. Nano Lett. , 2007 , 7 , 513-9.[71] Dunlop, J.; Roncarati, R.; Jow, B.; Bothmann, H.; Lock, T.; Kowal,

D.; Bowlby, M.; Terstappen, G.C. In v itro screening strategies fornicotinic receptor ligands. Biochem. Pharmacol. , 2007 , 74 , 1172-81.

[72] Arneric, S.P.; Holladay, M.; Williams, M. Neuronal nicotinicreceptors: a perspective on two decades of drug discovery research.

Biochem. Pharmacol. , 2007 , 74 , 1092-101.[73] Gordon, E.; Cohen, J.-L.; Engel, R.; Abbott, G.W. 1,4-

Diazabicyclo[2.2.2]octane derivatives: a novel class of voltage-gated potassium channel blockers. Mol. Pharmacol. , 2006 , 69 , 718-26.

[74] Theis, S.; Hartrodt, B.; Kottra, G.; Neubert, K.; Daniel, H. Definingminimal structural features in substrates of the H +/peptide cotrans-

porter PEPT2 using novel amino acid and dipeptide derivatives. Mol. Pharmacol. , 2002 , 61 , 214-21.

[75] Amiri, S.; Shimomura, M.; Vijayan, R.; Nishiwake, H.; Akamatsu,M.; Matsuda, K.; Jones, A.K.; Sansom, M.S.P.; Biggin, P.C.; Sat-telle, D.B. A role for Leu118 of loop E in agonist binding to the 7nicotinic acetylcholine receptor. Mol. Pharmacol. , 2008 , 73 , 1659-67.

[76] Mihalak, K.B.; Carroll, F.I.; Luetje, C.W. Varenicline is a partialagonist at 4 2 and a full agonist at 7 neuronal nicotinic recep-tors. Mol. Pharmacol. , 2006 , 70 , 801-5.

[77] Xiang, G.X.; Pan, L.B.; Huang, L.H.; Yu, Z.Y.; Song, X.D.;Cheng, J.; Xing, W.L.; Zhou, Y.X. Microelectrode array-based sys-

tem for neuropharmacological applications with cortical neuronscultured in vitro . Biosens. Bioelectron. , 2007 , 22 , 2478-84.[78] Morin, F.; Nishimura, N.; Griscom, L.; LePioufle, B.; Fujita, H.;

Takamura, Y.; Tamiya, E. Constraining the connectivity of neu-ronal networks cultured on microelectrode arrays with microfluidictechniques: a step towards neuron-based functional chips. Biosens.

Bioelectron. , 2006 , 21 , 1093-100.[79] Liu, Q.J.; Huang, H.R.; Cai, H.; Xu, Y.; Li, Y.; Li, R.; Wang, P.

Embryonic stem cells as a novel cell source of cell-based biosen-sors. Biosens. Bioelectron. , 2007 , 22 , 810-5.

[80] Liu, Q.J.; Cai, H.; Xu, Y.; Li, Y.; Li, R.; Wang, P. Olfactory cell- based biosensor: a first step towards a neurochip of bioelectronicnose. Biosens. Bioelectron. , 2006 , 22 , 318-22.

[81] Reininger-Mack, A.; Thielecke, H.; Robitzki, A.A. 3D-biohybridsystems: applications in drug screening. Trends Biotechnol., 2002

20 , 56-61.[82] Wang, K.; Fishman, H.A.; Dai, H.J.; Harris, J.S. Neural stimulation

with a carbon nanotube microelectrode array. Nano Lett. , 2006 , 62043-8.

[83] Keefer, E.W.; Botterman, B.R.; Romero, M.I.; Rossi, A.F.; Gross,G.W. Carbon nanotube coating improves neuronal recordings. Nat.

Nanotechno l. , 2008 , 3 , 434-9.[84] Nguyen-Vu, T.D.B.; Chen, H.; Cassell, A.M.; Andrews, R.;

Meyyappan, M.; Li, J. Vertically aligned carbon nanofiber arrays:an advance toward electrical-neural interfaces. Small , 2005 , 2 , 8994.

[85] Krommenhoek, E.E.; Gardeniers, J.G.E.; Bomer, J.G.; Li, X.;Ottens, M.; van Dedem, G.W.K.; van Leeuwen, M.; van Gulik,W.M.; van der Wielen, L.A.M.; Heijnen, J.J.; van den Berg, A. In-tegrated electrochemical sensor array for on-line monitoring ofyeast fermentations. Anal. Chem. , 2007 , 79 , 5567-73.

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