1.Biosensor and Bioelectronic

This article appeared in a journal published by Elsevier.The attached copy is furnished to the author for internal non-commercial research and education use,including for instruction at the authors institutionand sharing with colleagues.Other uses,including reproduction and distribution,or selling or licensing copies,or posting to personal,institutional or third partywebsites are prohibited.In most cases authors are permitted to post their version of thearticle(e.g.in Word or Tex form)to their personal website orinstitutional repository.Authors requiring further informationregarding Elsevier’s archiving and manuscript policies areencouraged to visit:/copyrightBiosensors and Bioelectronics 26 (2011) 1866–1870Contents lists available at ScienceDirectBiosensors andBioelectronicsj o u r n a l h o m e p a g e :w w w.e l s e v i e r.c o m /l o c a t e /b i osThionin attached to a gold electrode modified with self-assembly of Mo 6S 9−X I X nanowires for amplified electrochemical detection of natural DNAHong Lin a ,b ,Huiming Cheng a ,Lu Liu a ,Zhiwei Zhu a ,Yuanhua Shao a ,Pagona Papakonstantinou c ,Dragan Mihailoviˇc d ,Meixian Li a ,∗aCollege of Chemistry and Molecular Engineering,Peking University,Beijing 100871,PR ChinabDepartment of Chemistry,College of Resources and Environment,Yuxi Normal University,Yuxi 653100,PR China cNanotechnology and Integrated Bioengineering Centre,University of Ulster,Newtownabbey,Co.Antrim BT370QB,Northern Ireland,UK dJozef Stefan Institute,Jamova 39,SI-1000Ljubljana,Sloveniaa r t i c l e i n f o Article history:Received 20November 2009Received in revised form 26January 2010Accepted 27January 2010Available online 4 February 2010Keywords:dsDNAMoSI nanowires ThioninSelf-assemblyElectrochemical sensora b s t r a c tWe demonstrate a novel electrochemical sensor for highly sensitive detection of natural double-stranded deoxyribonucleic acid (dsDNA)based on thionin (Th)attached to Mo 6S 9−X I X nanowires (MoSI NWs)self-assembled on a gold electrode.The sensing detection is based on a decrease of the voltammetric response of the immobilized Th due to the binding of Th with dsDNA through intercalation.MoSI NWs act as molecular connectors to provide an amplification and conductive sensing platform for the electrochem-ical detection of dsDNA,because many sulfur atoms at the ends and sides of MoSI NWs permit covalent bonds to be formed with gold as well as MoSI NWs with negative charges allow electrostatic binding with Th.A determination limit of 0.62ng/mL for dsDNA with this novel sensor is achieved,which is three orders of magnitude lower than that in the absence of MoSI NWs.The operation is simple and label-free.© 2010 Elsevier B.V. All rights reserved.1.IntroductionIt is well-known that the detection of DNA is important in genet-ics,pathology,criminology,pharmacogenetics,food safety and many other fields.Due to electrochemistry-based sensors offer-ing high sensitivity and low cost (Batchelor-McAuley et al.,2009;Drummond et al.,2003;Erdem,2007;Teles and Fonseca,2008),various approaches of electrochemical detection for DNA have been developed,including direct electron transfer of DNA at a mercury electrode and modified electrodes (Erdem et al.,2006;Johnston et al.,1995;Palecek,1960,1988;Wang et al.,1998),electrochemistry of DNA-specific redox reporters (Fojta et al.,2003;Yu et al.,2001),and electrochemical amplifications with nanomaterials (Lapierre-Devlin et al.,2005;Li et al.,2004).In recent years,nanomaterial-based electrochemical DNA sen-sors have attracted much attention due to their high sensitivity and selectivity in DNA detection.Various electrochemical sensing plat-forms based on namomaterials such as carbon nanotubes (Zhang et al.,2009a,b;Zhu et al.,2009),nanoparticles (Das and Yang,2009;Du et al.,2009;Li et al.,2009)and nanowires (Chen et al.,2009;Wang and Ozkan,2008)have been reported.These studies indi-∗Corresponding author.Tel.:+861062757953;fax:+861062751708.E-mail address:lmwx@ (M.Li).cate that nanomaterials are making a big impact on development of electrochemical biosensors and nanotechnology is bringing new possibilities for improving both the sensitivity and selectivity of biosensing devices.The rapid development of nanotechnology has made demands on new nanomaterials with new functional proper-ties.Molybdenum–chalcogenide–halide NWs which are composed of molybdenum (Mo),sulfur (S)and iodine (I)in the form of Mo 6S 9−X I X (MoSI)are a new class of quasi-one-dimensional objects,having two different stoichiometries Mo 6S 3I 6and Mo 6S 4.5I 4.5(Berˇc iˇc et al.,2006;Mihailovic,2009;Vrbanic et al.,2004).An identical skeletal structure composed of indistinguishable one-dimensional polymer chains of molybdenum–sulfur–iodine clusters,strongly joined together by anions (either S or I).The indi-vidual nanowires are joined together into bundles by weak Van der Waals forces.The materials have many remarkable properties including strong anisotropy,large Young moduli along the wires,very small shear moduli and controllable electronic pared to CNTs,MoSI NWs have some significant advantages such as straightforward synthesis,monodisperse diameters and metallic properties (Nicolosi et al.,2005;Uplaznik et al.,2006).They have demonstrated their potential to become unique building ele-ments in biosensors because sulfur atoms at the ends of MoSI NWs are capable of forming covalent bonds with a gold surface or thi-olated proteins (Ploscaru et al.,2007).Recently,we showed that bioassembled nanocircuits of MoSI NWs and anti-estrone antibody0956-5663/$–see front matter © 2010 Elsevier B.V. All rights reserved.doi:10.1016/j.bios.2010.01.035H.Lin et al./Biosensors and Bioelectronics26 (2011) 1866–18701867Fig.1.Schematic representation of the fabrication of the modified electrode and application in dsDNA assay:(A)a bare AuE;(B)the AuE was immersed in the MoSI NWs suspension for12h to form a self-assembled NW layer on the gold surface via the formation of Au–S bonds(NW/AuE);(C)the NWs/AuE was immersed in0.5mM Th solution overnight for immobilization of Th(Th-NW/AuE);(D)application of the Th-NW/AuE in dsDNA assay.on glassy carbon electrodes provide an amplification and conduc-tive pathway for the specific electrochemical sensing of estrone hapten(Sun et al.,2008).Electroactive Th is an easy available cationic phenothiazine dye, which can bind strongly to dsDNA through intercalation(Du et al., 2005;Gong et al.,2008;Hecht et al.,2004;Ye et al.,2003),so it can be used as an intercalator to design electrochemical biosensors for the detection of DNA.In order to further explore the application of MoSI NWs in elec-trochemical DNA sensing platform,in this work,Th attached to a gold electrode modified with self-assembly of MoSI NWs(denoted as Th-NWs/AuE)was prepared for the amplified electrochemical detection of dsDNA.The immobilization of Th was based on strongly electrostatic interaction of negatively charged NWs and positively charged Th.A decrease in the voltammetric response current of Th-NWs/AuE was induced by dsDNA due to the combination of dsDNA with Th blocking the redox reaction of Th(Fig.1).Thus,a simple and sensitive sensing platform was constructed for the detection of dsDNA without requirement of hybridization and any external labeling.2.Experimental2.1.Materials and reagentsMo6S9−X I X nanowires were fabricated by direct synthesis from elemental material that had been mixed in the desired stoi-chiometries,as described elsewhere(Vrbanic et al.,2004).Powders composed of aggregates of individual nanowires were obtained with two different stoichiometries Mo6S3I6and Mo6S4.5I4.5.The stoichiometries were determined by a combination of techniques such as chemical analysis,high-resolution transmission elec-tron microscopy,and X-ray photoelectron spectroscopy(XPS),as reported in the literature(Mihailovic,2009).In this work,all stud-ies were carried out on Mo6S3I6NWs.Fish sperm DNA was obtained from Acros;solution offish sperm DNA in0.2M NaCl,gave a ratio of UV absorbance at260nm and280nm,A260/A280,of ca.1.85,indi-cating that DNA was sufficiently free of protein and RNA(Marmur, 1961).The concentration offish sperm DNA standard stock solution (1.223mg/mL)was spectrophotometrically determined at260nm. Thionin chloride of analytical grade was from Aldrich.0.1M,pH 6.9phosphate buffer solution(KH2PO4–Na2HPO4)(PBS)was used as the supporting electrolyte.All other reagents were of analytical grade.Water was triply distilled with a quartz apparatus.In2-propanol,1mg/mL MoSI NW dispersions were prepared. The dispersions were initially sonicated for2min using a high-power ultrasonic tip(120W,60kHz)followed by a mild sonication for2h using a low power ultrasonic bath.Prior to use,the disper-sions were resonicated for20min to obtain uniform suspension.2.2.ApparatusCyclic voltammetric(CV)and differential pulse voltammetric (DPV)measurements were performed with a model CHI660A elec-trochemical workstation(Chenhua Instrument Co.Ltd.,Shanghai, PR China).A three-electrode system was employed with Pt wire as the auxiliary electrode,a KCl saturated calomel electrode(SCE)as the reference electrode,and a modified gold electrode(AuE)or bare AuE with a diameter of4mm as the working electrode.Prior to elec-trochemical experiments,the solutions were routinely deaerated by purging with high purity nitrogen.XPS analysis was performed with an Axis Ultra spectrometer(Kratos,UK).Using Mono Al K␣(1486.7eV)radiation at a power of225W(15mA,15kV).To com-pensate for surface charge effects,binding energies were calibrated using C1s hydrocarbon peak at284.8eV.UV–vis absorption spec-tra were recorded with a UV–vis spectrophotometer(PerkinElmer Lambda11)in a1cm quartz cuvette within a wavelength range from200nm to800nm.Atomic force microscopic(AFM)char-acterizations were performed in the tapping mode in air with a Nanoscope IIIa microscope(Digital instruments).All experiments were conducted at the ambient temperature(approx.20±2◦C).2.3.Fabrication of the modified electrodePrior to modification,an AuE was polished with1␮m alu-mina slurry followed by0.3␮m and0.05␮m alumina slurries (Buehler),respectively.The polished electrode was sonicated for 5min in deionized water to remove any remaining polishing agent.A smooth gold surface is important to support efficient NW bind-ing.The electrode was then scanned under basic conditions(0.5M NaOH)with a scan rate of2V/s over the potential range from −0.35V to−1.35V to clean the electrode till constant cyclic voltam-mograms were obtained(typically500scans).Then oxidation and reduction scans under acidic conditions(0.5M H2SO4)over the potential range from−0.1V to1.7V(60scans at a scan rate of 0.5V/s)were conducted.Sequently,the electrode was placed in 1mg/mL MoSI NW suspension for12h to form a self-assembled NW layer on the gold surface,then washed with2-propanol and dried(this modified electrode is denoted as NW/AuE).Finally, the NWs/AuE was immersed in ethanol solution of0.5mM Th overnight,resulting in the incorporation of Th,thoroughly rinsed with water to remove loosely bound Th,and subsequently air-dried (this modified electrode is denoted as Th-NWs/AuE).1868H.Lin et al./Biosensors and Bioelectronics26 (2011) 1866–1870 All the steps of the preparation of the samples for AFM charac-terization were repeated on smooth gold sheets.3.Results and discussion3.1.Characterization of the modified electrodeIn order to verify that MoSI NWs were covalently attached toan AuE through forming the Au–S bonds,XPS measurement hasbeen performed.XPS analysis clearly shows that the sulfur groupsin MoSI molecules with an electron binding energy of162.3eV and163.5eV corresponding to S2p3/2and S2p1/2signals(Nicolosi et al.,2007),overlay precisely with the characteristic binding energies ofsulfur atoms bound to the gold surface(Laibinis et al.,1991;Tarlovet al.,1993).Thus,the immobilization of MoSI NWs on the AuE wasvalidated.In order to attest the immobilization of Th on the NWs/AuE,elec-trochemical behaviors of the bare AuE,NWs/AuE,and Th-NWs/AuEin0.1M,pH6.9PBS were investigated.None redox peaks could beobserved on the bare AuE(Fig.2A,curve a)and NWs/AuE(Fig.2A,curve b)in the potential range from0.20V to−0.60V,whereasthe Th-NWs/AuE exhibited a pair of well-defined redox peaks atE pa=−0.249V and E pc=−0.300V,respectively(Fig.2A,curve c),which was attributed to two-electron-transfer reversible reductionof the immobilized Th on the basis of the reported literatures(LoetFig.2.(A)CVs of the bare AuE(a),NWs/AuE(b),Th-NWs/AuE(c)and Th/AuE(d)in 0.1M pH6.9PBS at a scan rate of0.1V/s;(B)CVs of the Th-NWs/AuE in0.1M pH6.9 PBS at different scan rates of0.05V/s(a),0.1V/s(b),0.3V/s(c),0.5V/s(d),0.7V/s(e), and1.0V/s(f).Inset:plot of the anodic(P a)and cathodic(P c)peak currents versus the scanrate.Fig.3.CVs of the NWs/AuE(a)and Th-NWs/AuE(b and c)in the absence(b)and presence of0.2446␮g/mL dsDNA(a and c)in0.1M pH6.9PBS at a scan rate of 0.1V/s.al.,2008;Salimi et al.,2006;Xu et al.,1998).Furthermore,the redox peak currents showed a linear dependence on the potential scan rate in the range from0.05V/s to1.0V/s(Fig.2B),which manifested the characteristic of surface-confined waves.The peak-to-peak sep-aration of around50mV is the most likely due to large IR drop.These results suggest that Th has been successfully incorporated onto the NWs/AuE.Since Th is a heteroaromatic sulfur-containing molecule and there are many sulfur atoms at the ends and sides of MoSI NWs, Raman and XPS spectroscopies were employed to test whether immobilization of Th on the NWs/GCE is accompanied by formation of disulfide bonds between MoSI NWs and Th.However,the for-mation of disulfide bonds is inconclusive.Therefore,we conclude that the immobilization of Th is mainly based on the strongly elec-trostatic interaction between negatively charged MoSI NWs and positively charged Th.Compared to the voltammetric response of the Th-NWs/AuE, the bare AuE without MoSI NWs immersed in0.5mM Th solution overnight(denoted as Th/AuE)showed a pair of small redox peaks, as shown in Fig.2A,curve d,which elucidates that the existence of MoSI NWs provides negative surface charge,large surface area and good conductivity,being beneficial for the immobilization of Th and enhancement of the voltammetric responses.3.2.Interaction of dsDNA with the immobilized ThBased on the reported literatures(Gong et al.,2008;Pang and Abruna,1998;Wang et al.,2008),electroactive Th could be utilized as an intercalator to design the electrochemical biosensor for the detection of DNA,therefore,the fabricated Th-NWs/AuE was used for the sensing detection of dsDNA.Fig.3shows CVs of the Th-NWs/AuE in0.1M pH6.9PBS in the absence(curve b)and presence of dsDNA(curve c).An obvious decrease in the redox peak currents for the Th-NWs/AuE with the addition of dsDNA was observed,and the redox peak potentials shifted to more positive positions.These indicated that free Th molecules on the surface of electrode were embedded by dsDNA and dsDNA inhibited the redox reaction of Th due to the intercalation of Th between two successive base pairs of dsDNA as the predominant interaction mode(Hecht et al.,2004; Rohs et al.,2000),resulting in the decrease of the redox peak cur-rents.In order to verify the interaction between Th and dsDNA,AFM characterization was performed.Fig.4shows AFM images of MoSI NWs self-assembled on gold sheets with and without dsDNA.The appearance of MoSI NWs is needle-shaped on the gold sheet(Fig.4A),whereas,it became curv-ing after Th-NWs/Au was immersed in dsDNA solution(Fig.4B).H.Lin et al./Biosensors and Bioelectronics26 (2011) 1866–18701869Fig.4.AFM tapping mode images of MoSI NWs self-assembled on gold sheets without(A)and with(B)dsDNA.The height profiles(a)and(b)correspond to the positions marked by white lines in A and B,respectively.Moreover,the height profiles revealed an increase in the diameter of nanowire bundle from12nm(Fig.4a)to22nm(Fig.4b),attesting the immobilization of DNA on MoSI NWs.Existence of the interac-tion between Th and DNA on MoSI NWs leads toflexible connection of different MoSI NW pared to the AFM image of MoSI NWs self-assembled on the gold sheet,no significant difference can be found for the AFM image of MoSI NWs self-assembled on the gold sheet with Th,the reason for this is that the resolution of AFM is not enough to observe Th immobilized on MoSI NWs.In order to further verify the interaction between Th and DNA, UV–vis spectral measurements were also conducted.A maximum absorption at262nm for the mixture corresponding to overlapping absorption peak of Th and dsDNA was observed.The characteristic absorption peak of Th at599nm shifted to604nm for the mix-ture,and the absorbance distinctly decreased.The red shift and hypochromic effect of the characteristic absorption peak for Th illustrate that Th can intercalate into dsDNA-duplex structure sta-bilized by␲–␲stacking and electrostatic interactions,similar to those reported in the literatures(Du et al.,2005;Long and Barton, 1990).3.3.Detection of dsDNA based on the Th-NWs/AuEBased on above experimental results,a sensitive electrochemi-cal sensor for the detection of dsDNA based on the Th-NWs/AuEwas Fig.5.(A)DPVs of Th-NWs/AuE immersed in different concentrations of dsDNA (from0.62ng/mL to489ng/mL)in0.1M pH6.9PBS.Pulse amplitude and pulse width adopted were50mV and50ms,respectively.The scan rate is20mV/s.(B) Calibration curve for the peak current responses(i p)with different concentrations of dsDNA on the Th-NWs/AuE in0.1M pH6.9PBS.(The inset shows a linear part of the curve.)Every point corresponds to the average value of three independent measurements.constructed,and DPV was employed to record current responses of the Th-NWs/AuE in the absence and presence of dsDNA by using the same modified electrode and adding successively different con-centrations of DNA to the same solution.Fig.5A shows that the current response decreased with the increase in the dsDNA con-centration from0.62ng/mL to489ng/mL.A typical sigmoid curve presenting the current response versus dsDNA concentration in logarithmic scale was obtained,as shown in Fig.5B.The linear range was from4.89ng/mL to79.5ng/mL for dsDNA,and the regression equation was i p(nA)=560.0−1.61C DNA(ng/mL)(R=0.995,n=8) (Fig.5B,inset).The determination limit was0.62ng/mL dsDNA, and it represents the minimum concentration of DNA at which the decrease of the current response for the Th-NWs/AuE could be observed.The relative standard deviation for thefixed concentra-tion of9.75ng/mL dsDNA withfive-time replicative preparations of the electrochemical sensor was found to be5.6%.These results indicate that the electrochemical sensor has high sensitivity and repeatability for the detection of dsDNA.In order to clarify the specificity of the sensor,BSA was used to perform the following control experiments:firstly,different con-centrations of BSA were determined by using the sensor,and the decrease of the peak current was also observed due to the electro-static interaction of negatively charged BSA and positively charged Th,but the resulting current responses were irregular and insignif-icant,as shown in Fig.6,curve a.Secondly,different concentrations of dsDNA in the presence of afixed concentration of1␮g/mL BSA were detected,and the similar sigmoid curve was achieved.A lin-ear dependence of the current response on the concentration of dsDNA over the range from6.12ng/mL to73.4ng/mL was obtained1870H.Lin et al./Biosensors and Bioelectronics26 (2011) 1866–1870Fig.6.Calibration curves for the current responses(i p)of different concentrations of BSA(a)and different concentrations of dsDNA at afixed concentration of1␮g/mL BSA(b)(the inset shows a linear part of curve b)on the Th-NWs/AuE in0.1M pH6.9PBS.Every point corresponds to the average value of three independent measurements.with a regression equation of i p(nA)=548.1−4.67C DNA(ng/mL) (R=0.998,n=7).In addition,the effect of other negatively charged molecules such as KCl and myoglobin on the detection of DNA was also studied,and no obvious interference was observed.Above results confirmed that the sensing measurement of dsDNA was mainly a consequence of a specific interaction rather than elec-trostatic or other nonspecific interactions.In order to elucidate the role of MoSI NWs on the specific inter-action between Th and dsDNA,a control experiment was designed involving all the steps in the construction process except the use of MoSI NWs.The resulting Th/AuE was also capable of detect-ing dsDNA.However,the determination limit for the Th/AuE was 1.22␮g/mL DNA,which is three orders of magnitude higher than that for the Th-NWs/Au.The control experiments indicate that the presence of MoSI NWs provides significant amplification of the electrochemical signal when coupled with the described scheme. Importantly,MoSI NWs act as molecular connectors because of many sulfur atoms at the ends and sides of MoSI NWs permitting covalent bonds to be formed with gold as well as MoSI NWs with negative charges allowing electrostatic binding with Th.In addition, MoSI NWs possess some appealing properties,such as high surface to volume ratio and metallic behavior,providing a sensitive and effective sensing interface.4.ConclusionsIn conclusion,we have successfully constructed a novel electro-chemical sensor based on Th attached to MoSI NWs self-assembled on the AuE for highly sensitive detection of natural DNA.The sens-ing detection is based on a decrease of the voltammetric response of the immobilized Th due to the binding of Th with dsDNA through intercalation.The operation is simple and label-free,yet a low determination limit of0.62ng/mL is achieved,which illustrates that MoSI NWs can act as a good sensing platform to construct highly sensitive and effective sensing devices.AcknowledgmentsThis project was supported by the National Natural Science Foundation of China(Grants20875005,20735001)and the Natural Science Foundation of Yunnan province(No.2008CD215). ReferencesBatchelor-McAuley,C.,Wildgoose,G.G.,Compton,R.G.,2009.Biosens.Bioelectron.24,3183–3190.Berˇc iˇc,B.,Pirnat,U.,Kusar,P.,Dvorsek,D.,Mihailovic,D.,Vengust,D.,Podobnik,B., 2006.Appl.Phys.Lett.88,173103.Chen,C.P.,Ganguly,A.,Wang,C.H.,Hsu,C.W.,Chattopadhyay,S.,Hsu,Y.K.,Chang, Y.C.,Chen,K.H.,Chen,L.C.,2009.Anal.Chem.81,36–42.Das,J.,Yang,H.,2009.J.Phys.Chem.C113,6093–6099.Drummond,T.G.,Hill,M.G.,Barton,J.K.,2003.Nat.Biotechnol.21,1192–1199. Du,J.Y.,Huang,X.H.,Xu,F.,Feng,Y.Y.,Xiang,W.,Lu,T.H.,2005.Spectro.Spec.Anal.25,1435–1438.Du,P.,Li,H.X.,Cao,W.,2009.Biosens.Bioelectron.24,3223–3228.Erdem,A.,2007.Talanta74,318–325.Erdem,A.,Papakonstantinou,P.,Murphy,H.,2006.Anal.Chem.78,6656–6659. Fojta,M.,Havran,L.,Billova,S.,Kostecka,P.,Masarik,M.,Kizek,R.,2003.Electroanal-ysis15,431–440.Gong,M.J.,Han,T.,Cai,C.X.,Lu,T.H.,Du,J.Y.,2008.J.Electroanal.Chem.623,8–14. Hecht,C.,Friedrich,J.,Chang,T.C.,2004.J.Phys.Chem.B108,10241–10244. Johnston,D.H.,Glasgow,K.,Thorp,H.H.,1995.J.Am.Chem.Soc.117,8933–8938. Laibinis,P.E.,Whitesides,G.M.,Allara,D.L.,Tao,Y.T.,Parikh,A.N.,Nuzzo,R.G.,1991.J.Am.Chem.Soc.113,7152–7167.Lapierre-Devlin,M.A.,Asher,C.L.,Taft,B.J.,Gasparac,R.,Roberts,M.A.,Kelley,S.O., 2005.Nano Lett.5,1051–1055.Li,G.J.,Li,X.L.,Wan,J.,Zhang,S.S.,2009.Biosens.Bioelectron.24,3281–3287.Li,Z.,Chen,Y.,Li,X.,Kamins,T.I.,Nauka,K.,Williams,R.S.,2004.Nano Lett.4, 245–247.Lo,P.H.,Kumar,S.A.,Chen,S.M.,2008.Colloids Surf.B:Biointerf.66,266–273. Long,E.C.,Barton,J.K.,1990.Acc.Chem.Res.23,271–273.Marmur,J.,1961.J.Mol.Biol.3,208–218.Mihailovic,D.,2009.Prog.Mater.Sci.54,309–350.Nicolosi,V.,Vrbanic,D.,Mrzel,A.,McCauley,J.,O’Flaherty,S.,McGuinness,C.,Com-pagnini,G.,Mihailovic,D.,Blau,W.J.,Coleman,J.N.,2005.J.Phys.Chem.B109, 7124–7133.Nicolosi,V.,Nellist,P.D.,Sanvito,S.,Cosgriff,E.C.,Krishnamurthy,S.,Blau,W.J.,Green, M.L.H.,Vengust,D.,Dvorsek,D.,Mihailovic,D.,Compagnini,G.,Sloan,J.,Stolojan, V.,Carey,J.D.,Pennycook,S.J.,Coleman,J.N.,2007.Adv.Mater.19,543–547. Palecek,E.,1960.Nature188,656–657.Palecek,E.,1988.Anal.Biochem.170,421–431.Pang,D.W.,Abruna,H.D.,1998.Anal.Chem.70,3162–3169.Ploscaru,M.I.,Kokalj,S.J.,Uplaznik,M.,Vengust,D.,Turk,D.,Mrzel,A.,Mihailovic,D.,2007.Nano Lett.7,1445–1448.Rohs,R.,Sklenar,H.,Lavery,R.,Ro1der,B.,2000.J.Am.Chem.Soc.122,2860–2866. Salimi,A.,Noorbakhsh,A.,Soltanian,S.,2006.Electroanalysis18,703–711.Sun,N.J.,McMullan,M.,Papakonstantinou,P.,Gao,H.,Zhang,X.X.,Mihailovic,D.,Li, M.X.,2008.Anal.Chem.80,3593–3597.Tarlov,M.J.,Burgess,D.R.F.,Gillen,G.,1993.J.Am.Chem.Soc.115,5305–5306. Teles,F.R.R.,Fonseca,L.P.,2008.Talanta77,606–623.Uplaznik,M.,Bercic, B.,Strle,J.,Ploscaru,M.I.,Dvorsek, D.,Kusar,P.,Deve-tak,M.,Vengust,D.,Podobnik,B.,Mihailovic,D.,2006.Nanotechnology17, 5142–5146.Vrbanic,D.,Remskar,M.,Jesih,A.,Mrzel,A.,Umek,P.,Ponikvar,M.,Jancar,B.,Meden,A.,Novosel,B.,Pejovnik,S.,Venturini,P.,Coleman,J.C.,Mihailovic,D.,2004.Nanotechnology15,635–638.Wang,J.,Rivas,G.,Fernandes,J.R.,Lopez Paz,J.L.,Jiang,M.,Waymire,R.,1998.Anal.Chim.Acta375,197–203.Wang,X.,Ozkan,C.S.,2008.Nano Lett.8,398–404.Wang,Z.J.,Li,M.Y.,Su,P.P.,Zhang,Y.J.,Shen,Y.F.,Han,D.X.,Ivaska,A.,Niu,L.,2008.mun.10,306–310.Xu,J.J.,Zhou,D.M.,Chen,H.Y.,1998.Electroanalysis10,713–716.Ye,B.X.,Liu,L.F.,Qi,X.H.,Wang,C.H.,2003.J.Anal.Sci.19,51–53.Yu,C.J.,Wan,Y.J.,Yowanto,H.,Li,J.,Tao,C.L.,Blackburn,G.F.,Meade,T.J.,2001.J.Am.Chem.Soc.123,11155–11161.Zhang,X.Z.,Jiao,K.,Liu,S.F.,Hu,Y.W.,2009a.Anal.Chem.81,6006–6012.Zhang,Y.Z.,Ma,H.Y.,Zhang,K.Y.,Zhang,S.J.,Wang,J.,2009b.Electrochim.Acta54, 2385–2391.Zhu,N.N.,Lin,Y.Q.,Yu,P.,Su,L.,Mao,L.Q.,2009.Anal.Chim.Acta650,44–48.。