DOI: 10.1002/elan.201400086
Development of TiO2 and Au Nanocomposite Electrode as CEA Immunosensor Transducer Sema Aslan[a] and lk Anik*[a]
Abstract: A carbon paste electrode (CPE) was modified by titanium(IV) oxide microparticles and then gold nanoparticles were dispersed in chitosan and immobilized on the CPE surface. As result a nanostructure modified composite electrode was obtained. Carcinoembriyonic antigen (CEA) was chosen as model analyte and the developed electrode was used as CEA immunosensor transducer for the first time. Two linear ranges of 0.01–1 ng/mL and 1–
20 ng/mL with corresponding correlation coefficients R2 = 0.986 and R2 = 0.990, were found respectively. The limit of detection of the developed immunosensor was calculated as 0.01 ng/mL with 2.8 % relative standard deviation (n = 5). The developed system was applied for CEA detection in synthetically prepared serum sample and very promising recovery values were obtained.
Keywords: Immunoassay · Nanostructures · Titanium(IV) oxide microparticles · Gold nanoparticles · Carcinoembriyonic antigen
1 Introduction Nanocomposite materials exhibit novel properties including quantum size effect, optical, catalytical and electrochemical properties which largely differ from the bulk materials due to their small size [1–5]. For these reasons, they have been extensively used in many areas including electrochemical biosensor applications [3, 6–11]. As a type of nanomaterials, gold nanoparticles (Au-np) are one of the most utilized metallic nanoparticles in electroanalytical applications. Au-np provides a suitable microenvironment similar to that of redox protein with more freedom in orientation. Moreover, it is claimed that this nanoparticle reduces the insulating effect of the protein shell by providing direct electron transfer through the conducting tunnels of gold nanocrystals. It is stated that the nanomeric edges of gold particles penetrate the insulating shell of enzyme. By this way, the distance between the electrode and biomolecular redox sites for electron transfer is decreased [4, 5, 12–18]. Titanium (IV) oxide (TiO2) nanocomposites are well known with their unique properties as being one of the typical biocompatible materials. They have been widely applied in biomedical and bioengineering fields due to their strong oxidizing properties, chemical inertness and nontoxicity [19–21]. These particles are also environmentally-friendly and because of this, they have been frequently proposed as a prospective interface for the immobilization of biomolecules [22, 23] and widely applied in photochemistry [24–26] and electrochemistry [10, 20, 27]. Immobilization of immunoreagent or enzyme onto the electrode surface is the key factor for the preparation of an effective electrochemical immunosensor/biosensor. Recently organic-inorganic composite materials have gained much attention and found a widespread application area in biosensor researches [28–35]. For example, Liu and Ma used Au–ionic liquid functionalized reduced www.electroanalysis.wiley-vch.de
graphene oxide nanocomposite for a sandwich-type electrochemical immunosensor design [36, 37]. Peng et al. reported a novel biomolecular immobilization strategy based on gold nanoparticles-polydopamine-thionine-graphene oxide nanocomposites [38, 39]. Jeong et al. examined increased electrocatalyzed performance through dendrimer-encapsulated gold nanoparticles and carbon nanotube-assisted multiple bienzymatic labels for highly sensitive protein detection with electrochemical immunosensor [40]. Also Jie et al. used dendrimer/CdSeZnS quantum dot nanoclusters for cancer cell detection [41, 42]. The usage of these materials provides the combination of unique physicochemical properties of compounds and as a result, better signals have been obtained. Chitosan (CS) is a kind of polysaccharide that can be produced by the deacetylation of chitin monomer. It has been preferred as immobilization material because of its membrane properties, permeability resistance in aqueous media, high adhesion and mechanical strength [1, 43]. On the other hand, carbon electrodes are widely used in electroanalysis due to their low background current, wide potential window, chemical inertness, low cost, and suitability for various sensing and detection applications [44–46]. Several forms of carbon, suitable for electroanalytical applications, are available [3, 6]. Also carbon paste electrodes (CPE)s are one of the most commonly used carbon type electrodes which have high advantages like renewa-
[a] S. Aslan, . Anik Mug˘la Sıtkı KoÅman University, Faculty of Science, Chemistry Department 48000 Kçtekli/Mug˘la Turkey *e-mail:
[email protected] Supporting Information for this article is available on the WWW under http://dx.doi.org/10.1002/elan.201400086.
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bility and easy modification nature of composite structure. In this study, CPE was modified with TiO2 mps, Au nps via CS and a renewable and sensitive immunosensor (CPE/TiO2-mp/(CS+Au-np)) was fabricated to detect the model reagent CEA. Detection of CEA levels in adults is very important in terms of early cancer diagnosis [15, 20, 47, 48]. For this reason, it is important to get promising results with developed CEA immunosensor system. Immunoassay techniques based on the highly specific molecular recognition reaction of antigens by antibodies, have become main analytical methods for biochemical analysis [49–51]. In the developed system, formation of CEA antibodyantigen complexes on the resulting electrode was probed by [Fe(CN)6]3/4 redox pair and monitored by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). These two techniques are preferred to observe the changes onto electrode surface. The experimental factors like working pH, incubation time and incubation temperature were investigated in detail. Then, proposed immunosensor was applied to CEA detection in synthetic serum samples.
2 Experimental 2.1 Materials and Reagents All chemicals used in experiments were pure of analytical grade. KH2PO4 (pH 7) buffer solution (PBS) was used as supporting electrolyte through all measurements, for pH adjustment 1 M NaOH(aq) was used and both of these chemicals were purchased from Merck (Germany). K3Fe(CN)6(aq) was used as probe for all electrochemical measurements and purchased from Sigma (USA). Graphite powder was purchased from Merck (Germany) and combined with mineral oil to prepare carbon paste electrodes. Mineral oil, CS monomer (middle molecular weight), acetic acid, commercial Au np (10 nm sized), for the preparation of TiO2-mps, Ti(OCH(CH3)2)4 and absolute ethanol, for synthetic serum preparation NaCl(s), CaCl2(s), KCl(s), urea(s) were all purchased from Sigma
(USA), HCl(aq) was from Merck (Germany). CEA antigen and antibody and all other interferences were purchased from Sigma-Aldrich (USA). 2.2 Instruments m-AUTOLAB TYPE III electrochemical analyzer was equipped with GPES/FRA and used for electrochemical measurements. A standard three-electrode cell contained a platinum wire auxiliary electrode, an Ag j AgCl ((Ag/ AgCl/KCl (1 M)) (filled with 1 M KCl, Metrohm) reference elelctrode and CPE/TiO2-mp/(CS+Au-np) electrode as working electrode. 2.3 Preparation of the Electrode A CPE/TiO2-mp/(CS+Au-np) electrode was prepared as reported before [13] as shown in Figure 1. Firstly, CPE/ TiO2-mp composite electrode was prepared by mixing 50 % graphite powder with 20 % TiO2-mp and 30 % mineral oil. Obtained paste was filled into 2 mm radius sized hole of Teflon body where copper wire provided as electrical contact for the electrode. On the other hand, CS flakes were dissolved in 50 mL of 2 M acetic acid solution. 2 mL of commercial Au nps were added to this solution and stirred for 30 min at room temperature. Then the solution was sonicated for 15 min. for providing uniform dispersion of Au-nps into CS. After that 25 mL of CS/Au-np solution was spread uniformly onto CPE/TiO2mp electrode, and subsequently left to dry in a dessicator for 45 min. at room temperature. As a result, CPE/TiO2mp/(CS+Au-np) electrode was obtained. 2.4 Fabrication of the Immunosensor Preparation steps of immunosensor were illustrated in Figure 1. CPE/TiO2-mp/(CS+Au-np) modified electrode was immersed into anti-CEA solution at 4 8C for 1 hour. Then remaining active sides on the surface of obtained electrode were blocked by BSA (0.25 %, w/w) for 20 minutes to avoid the unspecific bindings. After that, surface
Fig. 1. Illustration of the fabrication steps of amperometric immunosensor a) CPE/TiO2-mp/(CS+Au-np), b) CPE/TiO2-mp/(CS+Aunp)/anti-CEA, c) CPE/TiO2-mp/(CS+Au-np)/anti-CEA/BSA, d) CPE/TiO2-mp/(CS+Au-np)/anti-CEA/BSA/CEA. www.electroanalysis.wiley-vch.de
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were given in Figure 2. The electrode is renewed before all electrochemical measurements.
2.5 Electrochemical Measurements EIS measurements were performed in 2 mM [Fe(CN)6]3/4 solution in 50 mM PBS (pH 7.0) at a frequency range from 105 to 10–1 Hz and a formal potential of 250 mV, using an alternating voltage of 10 mV. CV and DPV measurements of differently modified electrodes were performed in 1 mM [Fe(CN)6]3/4 solution in 50 mM PBS (pH 7.0). Potential ranges were from 0.8 to 0.6 V for CV at a scan rate of 100 mV s1, from 0.3 V to 0.7 V for differential pulse voltammetry (DPV) at a scan rate of 50 mV s1(vs. Ag/AgCl/KCl (1 M)).
3 Results and Discussions In our previous study, we developed and optimized the electrochemical behavior of TiO2-mp incorporated CPE where Au-np was attached with CS onto the electrode surface [13]. In this study, we used this optimized electrode system as an immunosensor transducer. Firstly electrochemical performance of the developed immunosensor was investigated. Then experimental parameters were optimized and analytical characteristics were examined. After the selectivity and stability of CPE/TiO2-mp/ (CS+Au np)/anti-CEA/BSA were observed, developed immunosensor system was applied for CEA detection in synthetically prepared serum samples.
3.1 Surface Imaging of the Developed Nanocomposite Immunosensor
Fig. 2. AFM images of (A) SPE/TiO2-mp/(CS+Au np), (B) SPE/TiO2-mp/(CS+Au np)/anti-CEA, (C) SPE/TiO2-mp/(CS+Au np)/anti-CEA/BSA/CEA.
of electrode was washed with distilled water. CPE/TiO2mp/(CS+Au np)/anti-CEA/BSA immunosensor was stored at 4 8C in PBS (pH 7) when not in use. These steps can also be observed from AFM images of developed immunosensor with screen printed electrodes (SPE) that www.electroanalysis.wiley-vch.de
AFM is a powerful tool to measure topography and properties of surfaces [51–56]. Here, the AFM technique was employed to illustrate the fabrication process of the immunosensor. The corresponding topographic images of stepwise self-assembly films on the electrode surface were presented in the Figure 2. Figure 2A showed that SPE/ TiO2-mp/(CS+Au np) electrode exhibited a surface with many globular nanoparticles dispersed over the entire substrate surface with grain diameter of 0.5 mm. After the modification of electrode surface with anti-CEA, Figure 2B showed an island like but larger sized grains about 1.5 mm. Also the little particles at the edges of these big grains showed that the SPE/TiO2-mp/(CS+Au np)/antiCEA modification was successful. After blocking with BSA and incubating with CEA, SPE/TiO2-mp/(CS+Au np)/anti-CEA/BSA/CEA surface film in Figure 2C became more flat because the antigen would fill up the interstice of the anti-CEA film. We can observe the homogenous cloud-like protein enwrapped and spread all over the surface with the grain size of 0.5–1 mm.
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3.2 Electrochemical Characteristics of the Immunosensor CV and EIS are effective and convenient techniques for probing the feature of the modified electrode surface. Here, both techniques were used to investigate electrochemical behavior of each assembly step. CVs of differently modified electrodes for 1 mM [Fe(CN)6]3/4 solution were presented in Figure 3A, while EIS results were given in Figure 3B. As can be seen from the Figure 3A, the maximum peak current at CV voltammograms was obtained with plain CPE/TiO2-mp/(CS+Au np). After the immobilization of anti-CEA on the electrode surface, the peak current clearly decreases (Figure 3Ab) suggesting the reduced effective area and active sites for electron transfer of the electrode. Then remaining active sites of electrode surface were blocked with BSA and obtained immunosensor was immersed into CEA solution to complete antigen-antibody interaction. As a result, a significant decrease at peak current was obtained due to immunocomplex reaction of the system (Figure 3Ac). EIS is an effective technique for probing the features of surface modified electrodes. The impedance spectra include a semicircle portion and a linear portion. The semicircle diameter at higher frequencies corresponds to the electron-transfer resistance (Ret), and the linear part at
lower frequencies corresponds to the diffusion process. The stepwise construction process of the immunosensor was examined by EIS. Obtained semicircles were according to Randles cirquit and phase angle of linear portion fits well to Warburg model. Figure 3B shows the EIS characterization of the modified electrodes at different stages and the best fitting circuit model was represented inset. In this [Rs(RQ)1(RQ)2] circuit Rs refers to solution resistance, (RQ)1 and (RQ)2 refer to phase layer and diffusion process between solution media and Au-np dispersed CS/CEA antibody layers on the electrode surface. From the Nyquistic diagram (Figure 3B), it is observed that the EIS of the CPE/TiO2-mp/(CS+Au np) modified electrode gives a small semicircle at high frequencies and linear part at low frequencies (curve a), suggesting very low Ret to redox probe [Fe(CN)6]3/4. After the immobilization of anti-CEA, resistance for the redox probe obviously increases (curve b), implying that anti-CEA is excellent electric insulating material and form a barrier for the electron transfer on the electrode surface. This also means that CPE/TiO2-mp/(CS+Au np)/anti-CEA modification step was accomplished. Subsequently, after blocking with BSA the CPE/TiO2-mp/(CS+Au np)/anti-CEA/ BSA modified electrode was immersed into CEA solution. Because of hydrophobic immunocomplex layer formation, the interfacial electron transfer is hindered
Fig. 3. A) Cyclic voltammograms, B) EIS of the modified electrodes at different stages (a) CPE/TiO2-mp/(CS+Au np), (b) CPE/ TiO2-mp/(CS+Au np)/anti-CEA, (c) CPE/TiO2-mp/(CS+Au np)/anti-CEA/BSA/CEA. www.electroanalysis.wiley-vch.de
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(curve c). As a result, CPE/TiO2-mp/(CS+Au np)/antiCEA/BSA/CEA electrode shows the maximum Ret value which indicates the reaction between antibody and antigen is satisfactory. Both CV and EIS provided the information about modification steps of electrode. On the other hand if these two methods were compared, CV gave more significant decrease compared to EIS. As a result, it can be concluded that for our developed electrode, CV is better than EIS for monitoring the modification steps.
surement we renewed the immunosensor. The current responses increase with increasing temperature up to 34 8C. The current values decrease harshly when temperatures are over 34 8C (Figure 5). This was attributed to the denaturation of protein at higher temperatures [49]. Thus, the temperature of 34 8C was selected as optimum working temperature for this study.
3.3 Optimization of Immunoassay Procedure
At 34 8C, the influence of the immunochemical incubation time was also investigated for 10 ng/mL CEA solution. As shown in Figure 6, the effect of incubation time on the current of immunosensor was investigated in the range of 2–28 min and current differences were calculated. We renewed the immunosensor before every measurement. The current difference of the amperometric response increases rapidly with the incubation time up to 12 min and reaches to steady state at 24 min. Because of minor increases at current values after 12 min, we preferred to use 12 min as optimal incubation time considering the practicality of immunosensor.
Experimental conditions like working pH, incubation time and incubation temperature were investigated with DPV method in order to compose the best analytical conditions for the developed immunosensor. 3.3.1 Optimization of pH The effect of pH of the buffer solution on the immunosensor behavior was investigated between pH 5.5 and 8.0. As shown in Figure 4, the peak current increases with increasing pH value from 5.5 to 7.0 and decreases with further increase at pH values. The results show that the maximum current response obtained at pH 7.0. It is known that the immunoreaction generally exhibits optimal binding at pH 7.4 [57, 58]. Since, pH 7 is very close to this completion pH, this value was chosen as working pH with PBS buffer and used for further studies. 3.3.2 Optimization of Incubation Temperature The effect of incubation temperature on the developed immunosensor was studied in the range of 15–40 8C with 10 ng/mL CEA. Before any temperature value was mea-
3.3.3 Optimization of Incubation Time
3.4 Analytical Characteristics of the Immunosensor After optimization of the experimental conditions, analytical characteristics of the developed immunosensor were examined (Figure 7). Two linear ranges were obtained. In the range of 0.01–1 ng/mL with a regression equation of y = 2.146x + 6.644, and correlation coefficient of 0.986, the LOD value was 0.01 ng/mL. In the range of 1–20 ng/mL the regression equation was y = 0.120x + 25.35 with a correlation coefficient of 0.990. The analytical performance of the proposed immunoassay has been compared with those of other CEA immu-
Fig. 4. Influence of the pH of the solution on the response of the immunosensor. www.electroanalysis.wiley-vch.de
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Fig. 5. The effect of incubation temperature (15–40 8C) on the response of the immunosensor.
Fig. 6. The effect of incubation time (2–30 min) on the response of the immunosensor.
noassays reported before (Table 1). The comparative data suggested that the present immunosensor is preferable over some earlier reported methods, especially considering the detection limit. 3.5 Selectivity and Stability of the Immunosensor In order to investigate the selectivity of CPE/TiO2-mp/( CS+Au np)/anti-CEA/BSA immunosensor, the effect of possible interferences like VEGF, IgG, BSA, dopamine, ascorbic acid, l-cysteine reagents were investigated. For this purpose three parallel studies were carried out and obtained current values were compared to 10 ng/mL CEA containing solutions incubation results. We renewed our electrode before every measurement. Firstly CPE/TiO2-mp/(CS+Au np)/anti-CEA/BSA was incubated into 10 ng/mLs of VEGF, IgG, BSA, dopamine, ascorbic acid, l-cysteine and CEA containing matrix and the obtained current values showed difference lower than 1 %. www.electroanalysis.wiley-vch.de
Secondly IgG was added to the 10 ng/mL CEA containing matrixes as increasing concentrations of 10, 20, 50 and 100 ng/mL. After that immunosensor was incubated into each of these solutions and obtained current results showed about 1–2 % difference in current values. Finally the same procedure with IgG was followed for BSA and no significant change was observed in current values. Only, 3.7 % difference was obtained when 10 fold more BSA (100 ng/mL) presents in the same medium with CEA. Difference on amperometric responses can be followed as % current difference and RSD values from the Table S1 (Supporting Information). As a result, it can be concluded that the obtained results show that the selectivity of the immunoreaction between antibody and antigen displays good specificity for the determination of CEA. For testing the storage stability of developed CEA immunosensor, the measurements were taken for 10 ng/mL CEA after 1, 3, 6, 12, 24 and 36 hour. Obtained current
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Fig. 7. Calibration curves for CEA determination, Inset: Linear relationship between the amperometric current response and CEA concentration in ranges of (A) 0.01–1 ng/mL and (B) 1–20.0 ng/mL.
Table 1. Comparison of different electrochemical immunosensors for the determination of CEA. Immunosensors
Linear range (ng/mL)
LOD (ng/mL)
References
Anti CEA/nano Au/poly o aminophenol/Au electrode Anti-CEA/colloid Au/SPCE (screen-printed carbon electrode Anti-CEA/core-shell Fe3O4/SiO2 nanoparticle Anti-CEA/HRP/solgel, reagentless Anti-CEA/thionine/HRP/Au electrode Anti-CEA/nanogold/(3-mercaptopropyl) trimethoxysilane/nano-Fe3O4/carbon paste electrode GCE/[AuCl4]/prussian blue/nanogold/anti-CEA anti-CEA/FCA-MPS/PVA/ITO (MWNTPEIAu/PB)5/(CSAu)/Ab1 [Ru(bpy)3]2 + -Ab2 as probes CGS-TB/PB-Ab2 as probes CPE/TiO2mp/(CS+Au np)/anti-CEA
0.5–20 0.50–25 1.6–60 0.5–120 0.6–200 1.0–55 3–80 0.5–45 0.5–160 1.0–100 0.5–60 0.01–20
0.1 0.22 0.5 0.4 0.2 0.13 0.9 0.2 0.08 0.5 0.1 0.01
[17] [50] [59] [60] [61] [62] [63] [64] [65] [66] [67] Present work
values showed no difference with other times besides 36 h. After 36 hours 8.4 % current decrease is observed. Though this difference can be seem as a big difference, since the system is composite in nature, a new electrode can be formed easily. Testing stability of developed CEA www.electroanalysis.wiley-vch.de
immunosensor was also checked for 50 cycles. As a result, 4.3 % current difference was obtained between 1st and 50th cycles when same electrode was used during the measurement.
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3.6 Application of the Immunosensor to CEA Detection in Synthetic Serum Samples Sample pretreatment studies were carried out in synthetic serum solution, including different concentrations of CEA in the range of 1, 5, 10, 15, 20 ng/mL. Each measurement was taken for five times. Current differences on detection of added amounts of CEA were measured with 103 %; 104 %; 100 %; 97 %; 102 % recoveries, respectively. Synthetic serum was prepared as reported before [68]. 140 mM NaCl, 4.5 mM KCl, 2.5 mM CaCl2, 0.8 mM MgCl2, 2.5 mM urea and 4.7 mM glucose containing 10 mM 10 mL of Tris-HCl solution were mixed well and pH of obtained solution adjusted to 7.3 by 1 M HCl(aq). . Then stock solutions of CEA were prepared with this snythetic serum. Developed immunosensor was immersed into these solutions for 12 min. at 34 8C.
4 Conclusions In this work, a novel CPE/TiO2-mp/(CS+Au np) modified nanocomposite electrode was fabricated and used for CEA detection which was selected as model reagent. Experimental results showed that the robust, sensitive and practical immunosensor was developed by introducing nano- and micro-particles into the electrode structure. Overall it can be said that a simple and sensitive immunosensor was developed which has the potential to be used for clinical applications.
Acknowledgements The grant from Mug˘la Sıtkı KoÅman University BAP Project Number 2011/10 is greatly acknowledged.
References [1] M. Zhang, W. Gorski, Anal. Chem. 2005, 77, 3960 – 3965. [2] W. Zhang, X. Qiao, J. Chen, Mater. Sci. Eng. B 2007, 142, 1 – 15. [3] U. Anik, S. Cevik, M. Pumera, Nanoscale Res. Lett. 2010, 5, 846 – 52. [4] S. Cevik, U. Anik, Sens. Lett. 2010, 8, 667 – 671. [5] M. Cubukcu, F. N. Ertas¸, U. Anik, Curr. Anal. Chem. 2012, 8, 351 – 357. [6] . Anık, M. C¸ubukÅu, Turk. J. Chem. 2008, 32, 711 – 719. [7] . Anik, S. C¸evik, Microchim. Acta 2009, 166, 209 – 213. [8] A. MerkoÅi, U. Anik, S. C¸evik, M. C¸ubukÅu, M. Guix, Electroanalysis 2010, 22, 1429 – 1436. [9] U. Anik, S. C¸evik, Microchim. Acta 2011, 174, 207 – 212. [10] M. C¸ubukÅu, F. N. Ertas¸, . Anık, Microchim. Acta 2013, 180, 93 – 100. [11] . Anik, Microchim. Acta 2013, 180, 741 – 749. [12] U. Anik, M. Cubukcu, Y. Yavuz, Artif. Cell. Nanomed. Biotechnol. 2013, 41, 8 – 12. [13] S. Aslan, Y. Yavuz, . Anık, Curr. Anal. Chem., 2012, DOI: 10.2174/15734110113090990015, in press. [14] A. Gole, C. Dash, C. Soman, S. R. Sainkar, M. Rao, M. Sastry, Bioconj. Chem. 2001, 12, 684 – 690. [15] Y. Liu, H. Jiang, Electroanalysis 2006, 18, 1007 – 1013. www.electroanalysis.wiley-vch.de
[16] W. J. Parak, T. Pellegrino, C. M. Micheel, D. Gerion, S. C. Williams, A. Paul, Nano Lett. 2003, 3, 33 – 36. [17] D. P. Tang, R. Yuan, Y. Q. Chai, X. Zhong, Y. Liu, J. Y. Dai, Biochem. Eng. J. 2004, 22, 43 – 49. [18] S. Xu, X. Han, Biosens. Bioelectron. 2004, 19, 1117 – 1120. [19] N. M. Dimitrijevic, Z. V. Saponjic, B. M. Rabatic, T. Rajh, J. Am. Chem. Soc. 2005, 127, 1344 – 1345. [20] Q. Shen, S. You, S. Park, H. Jiang, D. Guo, B. Chen, Electroanalysis 2008, 20, 2526 – 2530. [21] M. Song, C. Pan, J. Li, X. Wang, Z. Gu, Electroanalysis 2006, 18, 1995 – 2000. [22] S. Mathur, A. Erdem, C. Cavelius, S. Barth, J. Altmayer, Sens. Actuators B, Chem. 2009, 136, 432 – 437. [23] E. Topoglidis, A. E. G. Cass, G. Gilardi, S. Sadeghi, N. Beaumont, J. R. Durrant, Anal. Chem. 1998, 70, 5111 – 5113. [24] L. Lucarelli, V. Nadtochenko, J. Kiwi, Langmuir 2000, 16, 1102 – 1108. [25] D-W. Seo, S. Sarker, N. C. Deb Nath, S-W. Choi, A. J. S. Ahammad, J-J. Lee, W-G. Kim, Electrochim. Acta 2010, 55, 1483 – 1488. [26] M. Tian, G. Wu, B. Adams, J. Wen, J. Chen, J. Phys. Chem. C 2008, 112, 825 – 831. [27] L. Kavan, J. Rathousky, M. Graetzel, V. Shklover, A. Zukal, J. Phys. Chem. B 2000, 104, 12012 – 12020. [28] X. Jia, Z. Liu, N. Liu, Z. Ma, Biosens. Bioelectron. 2014, 53, 160 – 166. [29] W. Lu, C. Qian, L. Bi, L. Tao, J. Ge, J. Dong, W. Qian, Biosens. Bioelectron. 2014, 53, 346 – 354. [30] X. Miao, S. Zou, H. Zhang, L. Ling, Sens. Actuators B, Chem. 2014, 191, 396 – 400. [31] J. Zhou, J. Tang, G. Chen, D. Tang, Biosens. Bioelectron. 2014, 54, 323 – 328. [32] Z. Wang, N. Liu, Z. Ma, Biosens. Bioelectron. 2014, 53, 324 – 329. [33] R. Wang, X. Chen, J. Ma, Z. Ma, Sens. Actuators B, Chem. 2013, 176, 1044 – 1050. [34] Z. Liu, Z. Ma, Biosens. Bioelectron. 2013, 46, 1 – 7. [35] R. Akter, C. K. Rhee, Md. A. Rahman, Biosens. Bioelectron. 2013, 50, 118 – 124. [36] J. Yan, M. Yan, L. Gea, S. Geb, J. Yu, Sens. Actuators B, Chem. 2014, 193, 247 – 254. [37] N. Liu, Z. Ma, Biosens. Bioelectron. 2014, 51, 184 – 190. [38] H. P. Peng, Y. Hu, A. L. Liu, W. Chen, X. H. Lin, X. B. Yu, J. Electroanal. Chem. 2014, 712, 89 – 95. [39] B. Jin, P. Wang, H. Mao, H. Zhang, Z. Cheng, Z. Wu, B. Hu, X. Bian, C. Jia, F. Jing, Q. Jin, J. Zhao, Biosens. Bioelectron. 2013, DOI: http://dx.doi.org/10.1016/j.bios.2013.12.025, in press. [40] B. Jeong, R. Akter, O. H. Han, C. K. Rhee, M. D. A. Rahman, Anal. Chem. 2013, 85, 1784 – 1791. [41] M. Hasanzadeh, N. Shadjou, M. Eskandani, J. Soleymani, F. Jafari, M. Guardia, Trac – Trends Anal. Chem. 2014, 53, 137 – 149. [42] G. Jie, L. Wang, J. Yuan, S. Zhang, Anal. Chem. 2011, 83, 3873 – 3880. [43] Y. Liu, M. Wang, F. Zhao, Z. Xu, S. Dong, Biosens. Bioelectron. 2005, 21, 984 – 988. [44] P. T. Kissinger, W. R. Heineman, in Laboratory Techniques in Electroanalytical Chemistry, 2nd ed., Marcel Dekker, New York 1996. [45] R. L. McCreery, in: Electroanalytical Chemistry (Ed: A. J. Bard), Marcell Dekker, New York 1991, pp. 221 – 374. [46] J. Wang, Electroanalytical Chemistry, 2nd ed., Wiley, New York 2000. [47] N. Zamcheck, E. W. Martin, Am. Cancer Soc. 1981, 47, 1620 – 1627.
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[48] M. Withofs, F. Offner, P. Paepe, M. Praet, Br. J. Haematol. 2000, 110, 743 – 744. [49] X. Li, R. Yuan, Y. Chai, L. Zhang, Y. Zhuo, Y. Zhang, J. Biotechnol. 2006, 123, 356 – 366. [50] J. Wu, J. Tang, Z. Dai, F. Yan, H. Ju, N. El Murr, Biosens. Bioelectron. 2006, 22, 102 – 108. [51] Y. Zhuo, R. Yuan, Y. Chai, A. Sun, Y. Zhang, J. Yang, J. Biomater. 2006, 27, 5420 – 5429. [52] L. Mao, R. Yuan, Y. Chai, Y. Zhuo, X. Yang, Sens. Actuators B, Chem. 2010, 149, 226 – 232. [53] Y. Liao, R. Yuan, Y. Chai, Y. Zhuo, X. Yang, Anal. Biochem. 2010, 402, 47 – 53. [54] H. Yang, R. Yuan, Y. Chai, L. Mao, H. Su, W. Jiang, M. Liang, Biochem. Eng. J. 2011, 56, 116 – 124. [55] H. Yang, R. Yuan, Y. Chai, H. Su, Y. Zhuo, W. Jiang, Z. Song, Electrochim. Acta 2011, 56, 1973 – 1980. [56] X. Sun, Z. Ma, Anal. Chim. Acta 2013, 780, 95 – 100. [57] J. Wang, R. Yuan, Y. Chai, B. Yin, Y. Xu, S. Guan, Electroanalysis 2009, 21, 707 – 714. [58] Z. J. Song, R. Yuan, Y. Q. Chai, X. Che, P. Lv, Sci. China Chem. 2011, 54, 536 – 544. [59] J. Pan, Q. Yang, Anal. Bioanal. Chem. 2007, 88, 279 – 286.
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[60] F. Tan, F. Yan, H. Ju, Electrochem. Commun. 2006, 8, 1835 – 1839. [61] Z. Dai, J. Chen, F. Yan, H. Ju, Cancer Detect. Prev. 2005, 29, 233 – 240. [62] D. Tang, J. Ren, Anal. Chem. 2008, 80, 8064 – 8070. [63] K. G. Liu, R. Yuan, Y. Q. Chai, D. P. Tang, Bioproc. Biosyst. Eng. 2010, 33, 179 – 185. [64] J. Lin, Z. Wei, C. Mao, Biosens. Bioelectron. 2011, 29, 40 – 45. [65] Y. Zhang, H. Chen, X. Gao, Z. Chen, X. Lin, Biosens. Bioelectron. 2012, 35, 277 – 283. [66] L. Ge, J. Yan, X. Song, M. Yan, S. Ge, J. Yu, Biomaterials 2012, 33, 1024 – 1031. [67] X. Chen, X. Jia, J. Han, J. Ma, Z. Ma, Biosens. Bioelectron. 2013, 50, 356 – 361. [68] F. Coldur, M. AndaÅ, I. Isildak, J. Solid State Electrochem. 2010, 14, 2241 – 2249.
2014 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Received: February 18, 2014 Accepted: March 10, 2014 Published online: April 15, 2014
Electroanalysis 2014, 26, 1373 – 1381
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