Electrochemical Thinning of Thicker Gold Film with Qualified

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Anal. Chem. 2005, 77, 5760-5765

Electrochemical Thinning of Thicker Gold Film with Qualified Thickness for Surface Plasmon Resonance Sensing Jianlong Wang, Yong Shao, Yongdong Jin, Fuan Wang, and Shaojun Dong*

State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Changchun Jilin 130022, China, and Graduate School of the Chinese Academy of Sciences, Beijing 100039, China

To meet the requirement of surface plasmon resonance (SPR) sensing, controlling the thickness of the gold film is very important. Here, we report an efficient and simple approach to prepare a SPR-active substrate when the thickness of the gold film is larger than the optimizing 50 nm and smaller than 100 nm. This method is based on anodic electrodissolution of gold in electrolyte containing chloride ions. Using this method, the thickness of gold films can be easily changed at a nanometer scale by controlling the number of potential scans and the concentrations of chloride ions in the electrolyte. At the same time, the influence of gold film thickness on the SPR signal is recorded by SPR in real time. To assess the change of the surface roughness and morphology of gold film through anodic electrodissolution, atomic force microscopy was used. The surface roughness of the same Au film before and after anodic electrodissolution is 1.179 and 2.767 nm, respectively. The change of the surface roughness of Au film brings out a slight angle shift of SPR. This indicates that surface electrodissolution of the gold does not affect the character of the original bulk film and this film can be used for SPR experiments. To confirm our expectation, a simple adsorption experiment of cytochrome c (Cyt c) on the gold film treated with anodic electrodissolution modified by 11-mercaptoundecanic acid was carried out. The angle shift of SPR confirmed the adsorption of Cyt c, and the cyclic voltammetry of Cyt c provided a complementary confirmation for the adsorption of Cyt c. These results show that this approach provides a good way to change the thicker gold film to an optimized thickness of SPR sensing. The great advantage brought by this approach is in that it can convert the waste gold films with greater thicknesses fabricated by the vacuum deposition method or other methods into useful materials as active SPR substrates. Gold has been widely used in different fields of science and technology because of its excellent stability. However, it becomes unstable and dissolves in the positive potential region,1,2 especially * Corresponding author. Tel.: +86 431 5262101. Fax: +86 431 5689711. E-mail: [email protected]. (1) Rand, D. A. J.; Woods, R. J. Electroanal. Chem. 1972, 35, 209-218.

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in solutions containing chloride ions.3-6 The dissolution of gold is of concern to the electronics industry in the design and manufacture of certain film and integrated circuits. Thus, the dissolution process of gold and the morphology of the gold surface during the dissolution process have been extensively studied by electrochemistry, surface-enhanced Raman spectroscopy (SERS), and scanning tunneling microscopy (STM). For example, Heumann and Panesar investigated the anodic behavior of a gold electrode in a sulfuric acid solution containing chloride ion of various concentrations.7 Diaz et al. investigated gold dissolution in solutions with different Au+/Au3+ molar ratios.8 Based on the kinetics and thermodynamics analyses of the current-potential relation, they thought Au+/Au3+ molar ratios in reaction product were decided by the potential region. When the potential was more positive than 0.8 V (vs SHE), AuCl4- was formed on the electrode surface. On the other hand, AuCl2- was formed when the potential was more negative than 0.8 V. Lovecek et al. studied the anodic dissolution and passivation behavior of a gold electrode in neutral and acid sulfate solutions with and without Cl- ions using a rotating disk electrode.9 They confirmed a diffusion-activation control of the dissolution process and suggested that AuCl4- was the final dissolution product. With the development of SERS and STM technology, more detailed mechanisms of gold electrodissolution were proposed. By using the SERS method, Loo found the formation of AuCl2- and AuCl4- species on a gold electrode surface in 0.1 M KCl solution in a potential region more negative and positive than 0.9 V (vs SCE), respectively.10 Li et al. studied the gold oscillatory electrodissolution processes in HCl solution by in situ Raman spectroscopy.11 They measured the spatial profile of AuCl4- concentration in the diffusion layer and the temporal (2) Kelsall, G. H.; Wleham, N. J.; Diaz, M. A. J. Electroanal. Chem. 1993, 361, 13-24. (3) Gauer, J. N.; Schmid, G. M. J. Electroanal. Chem. 1970, 24, 279-286. (4) Cadle, S. H.; Bruckenstein, S. J. Electroanal. Chem. 1973, 48, 325-331. (5) Frankenthal, R. P.; Thompson, D. E. J. Electrochem. Soc. 1976, 123, 799804. (6) Trevor, D. J.; Chidsey, C. E. D.; Loiacono, D. N. Phys. Rev. lett. 1989, 62, 929. (7) Heumann, T.; Panesar, H. S. Z. Phys. Chem. 1965, 229, 84. (8) Diaz, M. A.; Kelsall, G. H.; Welham, N. J. J. Electroanal. Chem. 1993, 361, 25-38. (9) Lovrecek, B.; Moslavac, K.; Matic, D. Electrochim. Acta 1981, 26, 10871098. (10) Loo, B. H. J. Phys. Chem. 1982, 86, 433-437. (11) Li, Z. L.; Wu, T. H.; Niu, Z. J.; Huang, W.; Nie, H. D. Electrochem. Commun. 2004, 6, 44-48. 10.1021/ac0507035 CCC: $30.25

© 2005 American Chemical Society Published on Web 08/05/2005

evolution of AuCl4- during the current oscillations in situ by confocal Raman spectroscopy and found a function relation between the vibration bands for Au-Cl-, AuCl4-, and Au-OH and potential. Honbo et al. investigated the electrodissolution of gold (111) in pure HClO4 and HClO4 in the presence of chloride ion by in situ electrochemical scanning tunneling microscopy.12 They found that strongly adsorbed chloride ions on gold (111) markedly enhanced the surface diffusion of gold, and the anodic dissolution of gold was prohibited by the formation of the oxide layer even in a solution containing chloride ions. These investigations provide a basis for the present study by using gold anodic electrodissolution. Optical surface plasmon resonance spectroscopy is a powerful tool for in situ real-time characterization of solid/liquid interfaces. This technology has been receiving increased attention because of its potential as a rapid, label-free, high-selectivity, and highsensitivity assay technique,13 and it has seen widespread use for the study of interactions of biological molecules.14 In chemistry, SPR measurements have been used to study Langmuir-Blodgett films,15-17 self-assembled organic monolayers,18,19 conductive polymer film at electrochemical interfaces,20,21 and specifically and nonspecifically adsorbed biopolymers.22,23 However, these studies are all based on a film of Au (or other noble metal) with optimum thickness. A thin film of gold with a 45-60 nm thickness deposited on a glass substrate is used for the excitation of surface plasmon modes in the common Kretschmann configuration.24 Currently, almost all SPR-active substrates are prepared by vacuum deposition or sputtering of metal. The conditions of vacuum deposition or sputtering dominantly decide the thickness of the gold film. In fact, gold film possessing different thickness was usually produced even under the same conditions of vacuum deposition or sputtering because of the weak binding forces between the metal film25 and the substrate and the uneven distribution of glasses in the chamber of vacuum deposition or sputtering. In this paper, a simple approach for producing active SPR substrates by in situ control of the thickness of the gold film is presented. This method is based on anodic electrodissolution of gold in a solution containing chloride ions. The effect of the gold film thickness on the SPR signal can be in situ monitored during the anodic electrodissolution process. This strategy can be effectively applied to thinning gold film as a SPR substrate when (12) Honbo, H.; Sugawara, S.; Itaya, K. Anal. Chem. 1990, 62, 2424-2429. (13) Deckert, F.; Legay, F. Anal. Biochem. 1999, 274, 81-89. (14) Jung, L. S.; Shumaker-Parry, J. S.; Campbell, C. T.; Yee, S. S.; Gelb, M. H. J. Am. Chem. Soc. 2000, 122, 4177-4184. (15) Lofas, S.; Malmqvist, M.; Ronnberg, I.; Stenberg, E.; Liedberg, B.; Lundstrom, I. Sens. Actuators, B. 1991, 5, 79-84. (16) Lawrence, C. R.; Martin, A. S.; Sambles, J. R. Thin Solid Films 1992, 208, 269-273. (17) Wijekoon, W. M. K. P.; Asgharian, B.; Casstevens, M.; Samoc, M.; Talapatra, G. B.; Prasad, P. N.; Geisler, T.; Rosenkilde, S. Langmuir 1992, 8, 135139. (18) Peterlinz, K. A.; Georgiadis, R. Langmuir 1996, 12, 4731-4740. (19) Lang, H.; Duschl, C.; Gratzel, M.; Vogel, H. Thin Solid Films 1992, 210, 818-821. (20) Hanken, D. G.; Naujok, R. R.; Gray, J. M.; Corn, R. M. Anal. Chem. 1997, 69, 240-248. (21) Hanken, D. G.; Corn, R. M. Anal. Chem. 1997, 69, 3665-3673. (22) Jordan, C. E.; Corn, R. M. Anal. Chem. 1997, 69, 1449-1456. (23) Peterlinz, K. A.; Georgiadis, R. M.; Herne, T. M.; Tarlov, M. J. J. Am. Chem. Soc. 1997, 119, 3401-3402. (24) Raether, H. Surface Plasmons; Springer-Verlag: Berlin, 1988. (25) Janata, J. Principles of Chemical Sensors; Plenum Press: New York 1989.

the thickness of the film is larger than 50 nm and smaller than 100 nm. Most importantly, this method provides a chance to remedy the metal films with greater thickness to active SPR substrates for SPR sensing and converts wastes into useful materials. EXPERIMENTAL SECTION Reagents. All solutions were made with deionized water, which was further purified with a Milli-Q system (Millipore). 11Mercaptoundecanic acid (MUA) and cytochrome c (Cyt c) were obtained from Sigma. KCl, H2SO4 (double distilled, 98%), and glass microscope slides (1.8 × 1.8 cm2) were obtained from China. The phosphate-buffered saline solution (pH 7.2, 0.01 M) was used for preparing Cyt c solution (1 mg/ml). All of the chemicals, unless mentioned otherwise, were of analytical grade and were used as received. In Situ SPR Measurements. SPR measurements were performed with a home-built electrochemical SPR system.26 It was based on the Kretschmann configuration to achieve the resonant condition by attenuated total internal reflection spectroscopy. The SPR-active substrates were prepared by sputtering of gold. In detail, the gold films with different thicknesses were sputtered on one side of a glass slide for the excitation of surface plasmons. An underlayer of chromium (1.5 nm) was used to ensure mechanical stability of the gold film. For further experiments, SPRactive substrate cleaned with piranha solution was pressed onto the base of a half-cylindrical lens (BaK4, n ) 1.61) via an index matching oil (n ) 1.61). Linearly p-polarized light having a wavelength of 670 nm from a diode laser was directed through the prism onto the gold film. The intensity of the reflected light was measured as a function of the angle of incidence, θ, using a photodiode with a chopper/lock-in amplifier technique. The gold surface of the glass slide was mounted against the Teflon cell with use of a Kalrez O-ring (the apparent electrode area was 0.38 cm2). The small, single-compartment, three-electrode Teflon cell provided a liquid-tight seal and an electrolyte contact. The Teflon cell allowed for the simultaneous recording of SPR and electrochemical data and the application of a voltage to the sample. Electrochemical Experiments. Electrochemical experiments were carried out with a CHI600 electrochemical system in a Teflon cell. The Teflon cell was provided with a saturated Ag/AgCl reference electrode and a platinum counter electrode. Electrochemical measurements were all recorded and reported versus the KCl-saturated Ag/AgCl reference electrode. All experiments were done at room temperature. Atomic Force Microscopy (AFM) Characterization. Surface images of gold film substrates were acquired in the tapping mode under ambient conditions (SPA400; Seiko Instruments Industry Co., Tokyo, Japan) Si3N4 cantilevers having integral tips (spring contant, 0.02 N/m) were used. Images were obtained by oscillating the cantilever slightly below its resonance frequency (typically, 200-300 kHz) and raster scanning across the surface. RESULTS AND DISCUSSION Electrochemical Behavior of Gold Film in KCl and H2SO4 Solutions. The electrochemistry of gold in aqueous solution has (26) Kang, X. F.; Jin, Y. D.; Cheng, G. J.; Dong, S. J. Langmuir 2002, 18, 17131718.

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Figure 1. CVs of a gold film in 0.1 M/L H2SO4 (solid line) and 0.05 M/L KCl (dotted line) solutions. Scan rate is 0.2 V/s.

Figure 3. SPR reflectivity-incident angle (R-θ) curves at the gold film with an initial thickness of 95 nm recorded in H2O after each potential scan in 0.05 M/L KCl solution. Potential range is from 0 to 1.5 V, and scan rate is 0.04 V/s.

Figure 2. SPR reflectivity-incident angle (R-θ) curves at the gold film in 0.03 M/L KCl solution after each potential scan. Potential range is 0-1.5 V, and scan rate is 0.04 V/s.

been extensively studied and applied to clean the surface of gold electrode. Figure 1 shows cyclic voltammograms (CVs) of gold film obtained in 0.05 M KCl solution and 0.1 M H2SO4 solution in the potential range from 0 to 1.5 V with a scan rate at 0.2 V/s, respectively. The solid line represents a CV curve of gold film in 0.1 M H2SO4. The peaks at 1.4 and 0.9 V correspond to the redox processes of polycrystalline gold. In contrast, the CV curve of gold film in 0.05 M KCl solution in Figure 1 (dotted line) shows a broad anodic peak near 1.3 V in the positive potential sweep, which corresponds to oxidation of gold. The faradaic current for gold oxidation in 0.05 M KCl solution is much larger than that in 0.1 M H2SO4. It indicates that Cl- ion can greatly accelerate the electrodissolution of gold and enhance the diffusion of gold from surface to solution.12 Two cathodic peaks appear in the negative potential sweep. A smaller peak appears near 0.93 V that corresponds to the reduction of gold oxide, and a larger peak occurs at 0.6 V that corresponds to the dissolution of gold. As is shown by Gaur and Schmid, gold dissolution and oxide formation are competitive reactions.3 We can see that the peak near 0.6 V is much larger than the peak near 0.9 V; it proves that the electrodissolution of gold is the major reaction process during the negative potential scan when the electrolyte contains Cl- ions. As one of the reacting substances, the amount of Cl- ions can greatly affect the diffusion of the gold atom from surface to 5762 Analytical Chemistry, Vol. 77, No. 17, September 1, 2005

Figure 4. SPR reflectivity-incident angle (R-θ) curves at the gold film with an initial thickness of 77 nm before (solid line) and after (dotted line) anodic electrodissolution.

solution when the potential range is confined from 0 to 1.5 V and scan rate is fixed at 0.04 V/s. In the following experiments, we optimized the experimental conditions. The different concentrations of Cl- ions were used for thinning the gold film with different thickness. Characterization of the Variation of Gold Film Thickness by Electrochemistry and in Situ SPR. In principle, the SPR spectrum of a bare gold film demonstrates that the position of the resonance is mostly a function of the extinction coefficient, K; the width of the resonance is mainly determined by the value of the refractive index, n; and the depth of the SPR spectrum is predominantly controlled by the thickness of the metal film, d.27 Of these factors, the refractive index of gold and the extinction coefficient are usually variable in a limited range. Thus, the thickness of bare gold film becomes a crucial factor for the SPR signal. As a rule, the gold film with ∼50 nm thickness can give (27) Salamon, Z.; Macleod, H. A.; Tollin, G. Biochim. Biophys. Acta 1997, 1331, 117-129.

Figure 5. AFM images of the gold film with an initial thickness of 77 nm after anodic electrodissolution. (a) Taping mode AFM image across the edge of gold/glass. (b) Analysis of the profilometer across the edge of gold/glass.

the best SPR signal. When the thickness of the gold film is larger than 50 nm, the depth of the SPR spectrum gradually reduces with increasing gold film thickness; at the same time, the minimum resonance angle slightly increases. The width of the resonance increases and the depth of the SPR spectrum decreases when the gold film thickness is smaller than 50 nm. In the present experiment, we investigated the changes of SPR signal when the thickness of the gold film changed from ∼95 to ∼55 nm by in situ electrochemical surface plasmon resonance. In our initial experiment, a gold film with a thickness of ∼85 nm prepared by sputtering is used as a SPR substrate. Figure 2 shows the changes of SPR signal in 0.03 M KCl solution when the potential scan is continuously applied. The R-θ curve of SPR is recorded after each potential scan in 0.03 M KCl solution containing the halide of gold produced in the process of the potential scan. From Figure 2, we can see that both the increase of the depth of the SPR spectrum and the shift of the position of the resonance appear after each potential scan. The increase of the depth of the SPR spectrum can be attributed to the decrease of gold film thickness resulting from the electrodissolution of the surface gold atom. As for the shift of SPR angle to a larger angle, two reasons might to be used to explain for this phenomenon. One is the increase of the refractive index of solution resulting in the formation of gold halide during the potential scan. The major reason originates from the adsorption of gold halide and Cl- ions on the gold surface.10,28 This nonspecific adsorption not only reduces the effective area of gold film for SPR sensing but also results in a larger error for the extraction of an optical constant by nonlinear least-squares fitting to the full Fresnel equations.29 So it is necessary to reduce the angle shift resulting from the adsorption of gold halide and Clions produced in the process of obtaining the potential scan. In our further experiments, we effectively reduce this angle shift. First, the ∼95 nm gold film was used as the work electrode. The electrolyte solution of 0.05 M KCl was replaced by 0.1 M H2SO4 after each potential scan. And then, three cyclic potential scans were carried out in 0.1 M H2SO4. Last, H2O was add to the cell (28) Lei, H. W.; Uchida, H.; Watanabe, M. Langmuir 1997, 13, 3523-3528. (29) Damos, F. S.; Luz, R. C. S. L.; Kubota, T. Langmuir 2005, 21, 602-609.

Figure 6. AFM topographic images of the same polycrystalline gold film with an initial thickness of 95 nm before (a) and after (b) anodic electrodissolution of gold with 6 times in 0.05 M/L KCl solution.

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Figure 7. CVs of an evaporation-prepared Au film with an initial thickness of 95 nm in 0.1 M/L H2SO4 before (solid line) and after (dotted line) anodic electrodissolution of gold with 6 times in 0.05 M/L KCl solution. Scan rate is 0.04V/s.

and the R-θ curve of SPR was recorded. As shown in Figure 3, the depth of the SPR spectrum gradually increases with increasing number of potential scans and the minimum resonance angle almost keeps constant. This phenomenon accords with the above rule reported by Salamon et al.,27 and it confirms the feasibility that regulates the thickness of SPR gold film through controlled electrodissolution of gold in KCl solution. Compared with Figure 2, the depth of the SPR spectrum is visibly increased after the same number of potential scans. These differences of depth resulted from the different Cl- ion concentrations. To further compare the influence of the change of gold film thickness on the SPR signal, the thickness of the gold substrate is in situ simulated by nonlinear least-squares fitting to the full Fresnel equations after each potential scan. A four-layer (prism/bulk gold layer/surface gold layer/water) complex Fresnel calculation is used to extract the film thickness and the optical constants of gold film.30 At a wavelength of 670 nm, the refractive index values used for the SPR modeling calculations are 1.61 and 1.3301 for BaK4 lens and water, respectively. The real part of the dielectric constant used for bulk gold film is -0.1672, and the imaginary part of the dielectric constant is 3.4826. The thickness of bulk gold film is 40 nm. The optical constant and thickness of surface gold film are variable because the character of the surface gold layer is changed with potential scan. Based on these parameters, the initial thickness of gold film is estimated as 94.6 nm. The thickness of gold film is changed to 86.4, 78.8, 72.0, 66.8, 60.8, and 56.0 nm after each potential scan. The imaginary part of the dielectric constant for surface gold film is changed from 3.4826 to 3.5617, and the real part of the dielectric constant is changed from -0.1672 to -0.4137. To confirm this method can be used as a general approach for thinning gold film, a gold film with a thickness of ∼77 nm is thinned. The results are shown in Figure 4. The depth of the SPR spectrum obviously increases after anodic electrodissolution in KCl solution, and the thickness of gold substrate is changed from 77 to 59.4 nm. At the same time, the validity of gold film thickness simulated by the full Fresnel equations is confirmed at this gold film. The section plane of the gold film after anodic electrodissolution is obtained by conven(30) Jin, Y. D.; Dong, S. J. J. Phys. Chem. B 2003, 107, 13969-13975.

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Figure 8. SPR reflectivity-incident angle (R-θ) curves at a treated gold film in situ recorded after various surface modification steps. (1) The bare gold film treated with anodic electrodissolution. (2) Adsorbed MUA on the bare gold film. (3) Adsorbed cytochrome c on the gold film modified with MUA. All curves were recorded in pure water.

tional photolithography that was performed by spin-coating the photoresist (RZJ-390) on the gold film, exposing it to UV light under the mask containing the desired pattern, which was designed in a computer and printed onto a transparent film by a commercial print service with a high-resolution laser printer, developing, and etching by a wet chemical procedure carried out with aqua regia. After removing the remaining photoresist with acetone, AFM of tapping mode was used to show the profilometer across the edge of the gold/glass, and the experimental results are shown in Figure 5. A clear interface can be seen between the glass and the gold film in Figure 5a. Furthermore, we measure the thickness of gold film at this interface. As is shown in Figure 5b, the real thickness (∼59.94 nm) of the gold film after anodic electrodissolution is close to our simutated thickness (59.4 nm). From Figure 5b, we also find the real thickness of the gold film show little variation around 59.94 nm; this change mainly results from the surface roughness of the gold film. Although the surface roughness of the gold film is slightly increased, it hardly affects the angle shift of SPR curves (shown in Figure 4). On the basis of these experimental results, we confirm that this method is suitable for thinning gold film to an active SPR substrate. However, it must be pointed out that this method cannot be used when the thickness of the gold film is larger than 100 nm. That means that, if the substrate is too thick, the SPR resonance phenomenon cannot appear even after repetitively scanning in KCl solution. AFM Images and CVs of Gold Film in H2SO4 by Anodic Electrodissolution. The AFM topographic images (Figure 6) show the evidence of surface roughing of the gold film after electrochemical treatment. The change of gold surface morphology is distinctly observed by comparing panels a and b in Figure 6. The surface roughness of the same gold film before and after anodic electrodissolution is 1.179 and 2.767 nm, respectively. The increase of surface roughness comes from the diffusion of gold atoms on the gold film surface to solution in the presence of Clions. The degree of diffusion is mainly decided by the concentrations of Cl- ions. When the concentration of Cl- ion is smaller

Figure 9. CVs of horse heart Cyt c adsorbed on a monolayer of MUA at the gold film treated with anodic electrodissolution. Scan rate is 0.1 V/s. The electrolyte is 0.01 M/L PBS.

than 10-4 mol/L, small defects were formed on the gold surface,12 and the large numbers of gold atoms would dissolve into solution when the concentrations of Cl- ions were high enough. According to our simulated results, the dissolution region of gold was 5-8 nm after each potential scan in our experiment. Electrochemical characterization was also applied to study the change of electrode surface before and after the anodic electrodissolution of gold. Figure 7 shows two CV curves of gold film in 0.1 M H2SO4. The solid line and dotted line represent the CVs of gold film before and after anodic electrodissolution, respectively. From Figure 7, we can see that the current of the oxidation peak after anodic electrodissolution is larger than before anodic dissolution. The increase of oxidation peak current potentially indicates the presence of trace Cl- ions. Because Cl- ions can strongly adsorb on the gold surface, the H2SO4 rinse could not get rid of Cl- ion entirely.31 The residual Cl- ion resulted in an increase of the oxidation peak current. The reduction peak slightly shifted to higher potential. This shift is also from the effect of Cl- ion, and the result is fairly consistent with the CVs that reported for a gold electrode in the presence of Cl- ion by Cadle and Bruckenstein and Shi and Lipkowski.31,32 It further confirms the adsorption of trace Cl- ion on the gold surface. The adsorption of trace Cl- ion is unable to affect the functionality of the gold surface, on which (31) Cadle, S. H.; Bruckenstein, S. Anal. Chem. 1974, 46, 16-20. (32) Shi, Z. C.; Lipkowski, J. J. Electroanal. Chem. 1996, 403, 225-239. (33) Stenberg, E.; Persson, B.; Roos, H.; Urbaniczky, C. J. Colloid Interface Sci. 1991, 143, 513-526. (34) Chen, X. X.; Ferrigno, R.; Yang, J.; Whitesides, G. M. Langmuir 2002, 18, 7009-7015.

the monolayer of MUA was fabricated. After then, an adsorption experiment with Cyt c was carried out. Figure 8 shows experimental results of R-θ curves of SPR at a treated gold substrate in situ recorded after various surface modification steps. The minimum of the R-θ curves was shifts to higher angle after adsorption of MUA and Cyt c. The value of angle shift was decided by molecular weight and surface coverage of the molecule adsorbed on the gold film. MUA shows an angle shift of 0.03°, and Cyt c, possessing a larger molecular weight, results in an angle shift of 0.2°, which corresponds to a surface density of 2.0 ng/mm2. This estimation is based on the generally accepted and used assumption that a 0.1° angle shift in the SPR sensorgram corresponds to 1 ng/mm2 mass change at the sensor surface.33 This result is similar to the value reported by Chen et al.34 The electrochemistry of Cyt c at this treated gold surface was carried out as complementary information. As shown in Figure 9, the redox potential of Cyt c is ∼0.03 V, which also accords with the results of Chen et al.34 These results strongly confirm that the anodic electrodissolution does not change the characterization of the gold film, and this approach can be adopted as a general method for improving the SPR signal of gold film. CONCLUSIONS We have described in this paper a method for controlling the thickness of gold film as an active SPR substrate. This method is based on the anodic electrodissolution of gold film in electrolyte containing chloride ions. The work reported here is important as both basic research and application. First, we have successfully used the anodic electrodissolution of gold in electrolyte containing Cl- ion for SPR sensing; the change of the gold film thickness after each potential scan was computed by nonlinear least-squares fitting to the full Fresnel equations of SPR. This computation was beneficial for regulating the thickness of the gold film in a few nanometers accurately. Second, we have investigated the influence of gold film thickness on SPR signal monitored by in situ electrochemical surface plasmon resonance. It was of benefit for understanding the relation between the thickness of SPR gold film and SPR signal. Finally, this method provides a way to convert the wastes of gold film with larger thickness into useful materials as an active SPR substrate. ACKNOWLEDGMENT This work was supported by special funds for major state basic research of China (2002CB713803).

Received for review April 23, 2005. Accepted July 6, 2005. AC0507035

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