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A simple method for the preparation of ultrahigh sensitivity surface enhanced Raman scattering (SERS) active substrate
Applied Surface Science 240 (2005) 260–267 www.elsevier.com/locate/apsusc
A simple method for the preparation of ultrahigh sensitivity surface enhanced Raman scattering (SERS) active substrate Gang Wei, Hualan Zhou, Zhiguo Liu, Zhuang Li* State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Science, Renmin Street 5625, Changchun 130022, PR China Received in revised form 9 June 2004; accepted 21 June 2004 Available online 5 August 2004
Abstract By immersing mica modified with cetyltrimethylammonium bromide (CTAB) into the silver colloid, a high efficient surface enhanced Raman scattering (SERS) active substrate was formed within 2 h at room temperature. The limit of detection of the substrate for Rhodamine 6G is up to 110 14 M. Changing the concentration of silver colloid and the treating time, various silver aggregates such as nanocrystals, clusters and films were found, and the SERS spectra of these aggregates were also obtained. The results of SERS revealed that CTAB could accelerate aggregation of the silver colloid and cause great Raman enhancement. Bilayer of CTAB is very important for aggregation of silver colloid and the aggregation extent is the main factor for the enormous enhancement on this substrate. # 2004 Elsevier B.V. All rights reserved. PACS: 82.65.P; 61.43.H; 61.16.C; 82.70.D Keywords: Surface enhanced Raman scattering; Aggregation; Atomic force microscopy; Colloid
1. Introduction Surface enhanced Raman scattering (SERS) is a useful technique resulting in strongly increasing Raman signals from the molecules attached to nanometer sized metallic structures. Since the discovery of SERS, a key problem in analytical application of * Corresponding author. Tel.: +86 431 5262057; fax: +86 431 5262057. E-mail address: [email protected] (Z. Li).
SERS is to develop stable and reproducible SERSactive substrates that can provide a large enhancement factor. The first measurement of a surface Raman spectrum from pyridine adsorbed on an electrochemically roughened silver electrode was reported in 1974 [1]. Then Van Duyme group and Albrecht group measured Raman signal of pyridine on a roughened silver electrode [2–3]. Later other noble metals such as Au and Cu were also used as SERS-active substrates and revealed large enhancement [4–5]. Tian and coworkers generated SERS on net transition
0169-4332/$ – see front matter # 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.apsusc.2004.06.116
G. Wei et al. / Applied Surface Science 240 (2005) 260–267
metals (Pt, Ru, Rh, Pd, Fe, Co, and Ni) by developing various roughening procedures and optimizing the performance of the confocal Raman microscope [6–8]. But unfortunately, these substrates of transition metals still couldn’t be used to detect those samples of ultralow concentration, such as in single molecule level. Metal sols were used to prepare SERS-active substrates widely. It was known that silver colloids showed a higher enhancement factor in comparison with that of Au and Cu, so silver colloids revealed great advantage in studying of SERS and were used widely. Some new active silver substrates have been investigated, including silver-coated latex spheres on filter-paper [9], silver colloid on filter-paper [10–11] and silver-coated filter membrane [12]. The limit of detection for most samples was about 1 10 9 M or mg ml 1 for these substrates. A great breakthrough in studying of Raman was the detection of single molecule [13–15]. Nie and Emory confirmed the existence of Raman enhancement factors on the order of 1014 to 1015 for R6G on colloidal silver particles. By immobilizing a lager number of individual silver particles on a polylysine-coated glass, they obtained the SERS spectrum of single R6G molecule adsorbed on single silver nanoparticle [13]. Later Brus and coworkers repeated the SERS experiments of R6G adsorbed on silver particles [16–18]. They thought the larger SERS enhancement came from colloidal aggregates, such as nanocrystals and clusters. Now, more and more SERS-active substrates are prepared by forming clusters [19–21] and film [22–28] on the solid surface, such as roughened silver film prepared by silver mirror action and silver island film [25–26]. Other substrates such as aggregated gold nanorods [29] and silver nanowire monolayers [30] were used, which also showed good Raman enhancement. In this paper, we provided a simple method to prepare a SERS-active substrate. It was found that silver nanoparticles could aggregate on mica modified with CTAB and form silver nanocrystals, clusters and films by controlling the treating time. The ability of Raman enhancement of the three aggregates was different and the sequence was film > cluster > nanocrystal. SERS (53 numbers) spectra of R6G in 1 10 12 M were obtained on this substrate. The results revealed this substrate was stable and it had excellent reproducibility, and the limit of
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detection of this substrate for R6G can reach up to 1 10 14 M.
2. Experimental 2.1. Colloid preparation Silver colloids were prepared according to the standard citrate reduction of Lee and Meisal [31]. Briefly, using a heating plate and a 500 ml flask, silver nitrate (90 mg) was added to approximately 400 ml of ultrapure water (18.2 MV) and brought to boiling with rapid magnetic stirring. Ten milliliters 1% sodium citrate was added to this solution. Boiling continued for about 1 h, after which the solution was diluted with triply distilled water to obtain a final volume of 500 ml. 2.2. Preparation and characterization of SERSactive substrates CTAB-coated mica was prepared as followed: a 15 ml aliquot of CTAB (2 10 3 M) was dropped to the new cleaved mica (1 cm 1 cm), then the mica was covered with a piece of parafilm (1.5 cm 1.5 cm) to make the solution expandedness adequately. Fifteen minutes later, the mica modified with CTAB was immersed into the silver colloid for different time. Contact mode AFM imaging was performed on a Digital Instrument Multimode AFM controlled by Nanoscope IIIa (DI, Santa Barbara, CA). The cantilevers were 115 mm in length with Si3N4 tips. All of the images were presented in this article represent flattened data obtained in air. Topographical roughness profiles were extracted from digital image data using a section analysis program. Optical spectra were acquired using UV-2450 spectrophotometer (SHIMADZU, Japan). 2.3. Sample preparation Rhodamine 6G solution was diluted to various concentration from 1 10 9 M to 1 10 15 M with methanol, then 20 ml such solution was dropped to the mica modified with CTAB or polylysine and allowed the solvent to evaporate under ambient conditions.
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2.4. Raman measurements SERS spectra were measured with a Renishaw 2000 model confocal microscopy Raman spectrometer with a CCD detector and a holographic notch filter (Renishaw Ltd., Gloucestershire, UK). The microscope attachment was based on a Leica DMLM system, and a 50 objective was used to focus the laser beam onto a spot with approximately 1 mm in diameter. Radiation of 514.5 nm from an air-cooled argon ion laser was used for the SERS excitation with power of not more than a few mW at the sample position. All of the spectra
reported were the results of a single 30 s accumulation.
3. Results and discussion 3.1. Silver particles aggregates formed on the mica modified by CTAB Several kinds of aggregates formed by immersing mica modified with CTAB into silver colloid for different time. AFM was used to image the actual structure of these various aggregates. Fig. 1a–d shows
Fig. 1. Contact mode AFM images of: (a) single silver nanoparticles, (b) nanocrystals, (c) clusters, and (d) film.
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the typical AFM images of single silver nanoparticle and the three silver aggregates, nanocrystal, cluster and film, respectively. Fig. 1a shows the image of silver nanoparticles dropped onto cleaved mica directly. Section analysis was used to measure the dimension of about 50 nanoparticles. The particle size was based on height value for tip convolution could affect the accuracy of lateral measurements. The results showed that the particles have an average size of 39.8 nm with a standard deviation of 5.5 nm. With the different treating time, nanocrystals, clusters and films were formed on the mica. When the treating time is 30 min, nanocrystals were formed first. The AFM image shows that this SERS-active particle is a tightly packed aggregate of more than 10 particles (Fig. 1b), which is similar to the studies of Brus and coworkers [16–18], who reported that the nanocrystal is aggregated by 10–15 single nanoparticles. When the treating time is increased to about 1 h, silver clusters are found all over the mica scanned (Fig. 1c). It was found that the clusters are silver aggregates of several hundred silver nanoparticles. We conclude it attributed to the action of CTAB, which balanced the surface charge of silver nanoparticles and made silver nanocrystals reaggregated to form the complex structure. From the image, we find the cluster is not a single
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nanometer layer completely. Some parts of the cluster are higher than others. These higher and larger particles are the centers of the reaggregation. Moreover, aggregation is irreversible, therefore, the aggregated clusters will become larger and larger. When the treating time is increased to 2 h, we can see there is a visible yellow–gray membrane formed on the surface of mica. AFM image shows much bigger silver clusters were formed. An interesting result is that a more compact and orderly silver nanometer film is found (Fig. 1d), which is similar to the self-assembly of nanoparticles monolayer. These aggregates all show great Raman enhancement effect. In the following section, we will further discuss the issue in detail. As we have shown, aggregation of silver nanoparticles took place on the mica modified with CTAB. We also discuss the effect of different treating time to the silver aggregates in solution. After a 10-fold dilution, silver colloid was mixed with 20 ml CTAB (1 10 3 M). Fig. 2 shows the UV–vis absorbance spectra of the colloid and that of the CTAB aggregated colloid. When treating time was 0 min, there was a sharp peak at about 420 nm. When CTAB was added to the Silver colloid, it revealed a distinct change for the absorbance curve after 1 min, the high and sharp peak changed into a short and wide one. After 1 h, the
Fig. 2. UV–vis absorption changes of colloidal silver (5 ml, diluted for 10-fold) added of 20 ml CTAB (1 10
3
M) in different treating time.
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peak in 420 nm is nearly disappeared. Moreover, a new peak appears at 750 nm, which is attributed to the appearance of silver aggregates. This shows that the bulk property of the colloid has changed due to the addition of the CTAB, which is similar to the study of Graham et al. [20]. 3.2. SERS spectra of R6G on the active substrate R6G samples of ultralow concentration were used to verify the enhancement effect of this SERS-active substrate. Using the confocal microscopy Raman system, we obtained a laser beam spot with diameter of 1 mm. It is easy to measure a SERS spectrum of sample with scales of a few micrometers. With 50 objective and 10 ocular, we can observe the least silver aggregates is about 400 nm, so it is possible for us to obtain SERS spectra after we have gained the AFM images on the same area of the sample. We have applied a novel relocated technique for biology molecules by AFM [32]. Using this relocated technique, we can obtain the SERS spectra and AFM images of the Silver clusters and films on the same location. Fig. 3a– c shows the SERS spectra of R6G adsorbed onto the three aggregates film, cluster and nanocrystal, respectively. The nanocrystal is compact aggregate of more than 10 nanoparticles, so there are 4–5 nanocrystals in
an area of 2 mm 2 mm size. These disturbs that the spectra are not the real SERS spectra of single nanocrystal, but the multiple action of several nanocrystals. Different SERS signals were found for these aggregates. At the same experiment conditions (integration time: 30 s, power: 10%), films revealed greater SERS signal than other aggregates. Dropping silver nanoparticles onto the cleaved mica directly, SERS spectrum shows the lowest concentration of R6G can be detected is only about 1 10 9 M on this substrate, moreover the stability and reproducibility are poor. But for the Ag film, the greatest enhancement effect is observed and the limit of detection is up to 1 10 14 M. Even if we made the laser power reduced to 1%, an interpretable Raman signal is still obtained (dada no shown). So we deduce that the aggregation extent of silver colloid could affect SERS signals greatly. SERS (53 numbers) spectra of R6G (1 10 12 M) were also obtained. Fig. 4 displays the peak heights of the 1650 cm 1 line for the 53 SERS spectra. In the histogram, 39 of the 53 spectra show great Raman signal (>200 c/s), and only one spectrum shows low Raman signal (<10 c/s). Because the selection of the location measured is random, we conclude that this substrate is stable and can be used to detect those samples of ultralow concentration.
Fig. 3. SERS spectra of R6G obtained on: (a) films, (b) clusters, (c) nanocrystals, and (d) nanoparticles. Experiment conditions: for (a)–(c), the concentration of R6G was 1 10 12 M, Laser power was 25 mW (1%).
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Fig. 4. Peak height of 1650 cm integration time: 30 s.
1
line for the 53 SERS spectra obtained when the concentration of R6G was 1 10
3.3. Aggregation extent is the main factor for the enormous enhancement Despite extensive studies of SERS since its discovery nearly many years ago and a literature of thousands of papers, a complete understanding of the mechanism of enhancement was lacking. SERS has historically been described in terms of electromagnetic and chemical or ‘‘first layer’’ enhancement mechanisms [33–34]. SERS intensity was found to be stronger for adsorbed molecules on surfaces covered with aggregated nanoparticles, the observed increase in the SERS intensity in the case of aggregation was attributed to the enhancement of electric field between particles in the aggregates [17,18,34,35]. Nanoparticles aggregates have been attracting growing interests for their collective electronic, optical, and magnetic properties. It was revealed that halide ions such as Cl and I were often used as aggregating agents [35–37], they appeared to reduce the surface charge on the colloid, lower ‘‘j-potential’’, then paving way for aggregation of nanopartilces [37]. In Nie’s studies about single molecule, they found there were only a few particles could show great Raman enhancement and claimed that the great
265
12
M. Power: 25 mW, 1%;
enhancement was attributed to ‘‘hot’’ nanoparticles [13–15,35]. In our experiment, we think that aggregation extent and dimension of particles are two key factors for the enhancement ability. Great SERS signals have been obtained on the aggregates of nanocrystals, clusters and films. And SERS signals increased with the increase of degree of aggregation. In theory, when concentration of R6G is 2 10 11 M, there is only one R6G molecule in 1 mm2, but the concentration of R6G is reduced to 1 10 14 M, we can still gain a clear SERS spectrum. So we can conclude that R6G was not absorbed to the SERSactive substrate averagely but absorbed to the hot sites provided by aggregates. But how silver nanoparticles were aggregated and formed various aggregates on the CTAB-coated mica? We proposed electrostatic interaction is the main force to cause aggregation. A control experiment was applied to testify the supposition. CTAB was diluted to 5 10 4 M, 1 10 4 M, 1 10 5 M and the substrates were prepared as experiment section. AFM images showed there were sparse nanoparticles adsorbed, even nearly no aggregate was observed (data not shown). Fig. 5a–c shows the SERS spectra of R6G (1 10 12 M) adsorbed on these substrates. When concentration of CTAB is 5 10 4 M, the
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Fig. 5. SERS spectra of R6G (1 10 12 M) on three different substrates. The concentration of CTAB for the preparation were: (a) 5 10 (b) 1 10 4 M, and (c) 1 10 5 M. Laser power: 25 mW, 1%; integration time: 30 s.
Scheme 1. Schematic binding mechanisms of CTAB adsorbed to cleaved mica.
4
M,
spectrum shows greatest SERS signal (Fig. 5a). But there is no SERS signal observed when concentration of CTAB is reduced to 1 10 5 M (Fig. 5c). The possible explanation is that for the surface of mica is hydrophilic and electronegative, so the hydrophilic cation group (–NH4+) of CTAB can be easily bound to the surface of mica and the hydrophobic group (CH3(CH2)n–) faces liquid phase (Scheme 1a). But when the concentration of CTAB is higher than its critical micelle concentration (about 0.94 mM), bilayer of CTAB was formed on mica. Two CTAB molecules are linked on the hydrophobic groups and a positive-charge surface is formed (Scheme 1b). On the other hand, we must consider the surface of mica cannot be coated by CTAB enough if the concentration of CTAB were too low, which made silver nanoparticles couldn’t adsorb onto the CTAB-coated mica adequately. Compared with the method of mixing aggregation reagents with silver colloid and dropping onto a solid substrate, the technique provided in this report is more applied and this substrate shows greater enhancement effect.
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4. Conclusions In summary, we have presented a simple method to prepare SERS-active substrate, which showed greater surface enhanced Raman scattering signals than those substrates made by the conventional method. The limit of detection for R6G can add up to 1 10 14 M and the stability and reproducibility are very good. Making use of the relocate technique, we not only obtained the AFM images of silver nanocrystal, cluster and film, but also gained the SERS spectrum of single R6G molecule on the same aggregate. The great enhancement depended on the size of silver nanoparticles and degree of aggregation. The driving force for forming these different aggregates must be electrostatic interactions in this study, which provided by bilayers of CTAB. Controlling aggregation of silver nanoparticles by changing the treating time and the concentration of CTAB, it is more significant to unveil mechanism of SERS. We hope this preparation of SERS-active substrate can be used to investigated the interaction of drug ligand with DNA at very low concentration. Further study about this is processing.
Acknowledgements This work was supported by the National Natural Science Foundation of China (no. 30070417).
References [1] M. Fleischman, P.J. Hendra, A.J. McQuillan, Chem. Phys. Lett. 26 (1997) 123. [2] D.L. Jeanmaire, R.P.V. Duyne, J. Electroanal. Chem. 84 (1977) 1. [3] M.G. Albrecht, J.A. Creighton, J. Am. Chem. Soc. 99 (1977) 5215. ¨ . Ertu¨ rk, A. Otto, Surf. Sci. 179 (1987) 163. [4] U [5] H. Baltruschat, E. Rach, J. Heitbaum, J. Electroanal. Chem. 194 (1985) 109. [6] Z.Q. Tian, B. Ren, D.Y. Wu, J. Phys. Chem. B 106 (2002) 9463. [7] Z.Q. Tian, B. Ren, B.W. Mao, J. Phys. Chem. B 101 (1997) 1338.
267
[8] D.Y. Wu, Y. Xie, B. Ren, J.W. Yan, B.W. Mao, Z.Q. Tian, Phys. Chem. Commun. 18 (2001) 1. [9] T. Vo-Dinh, M.Y.K. Hiromoto, G.M. Begun, R.L. Moody, Anal. Chem. 56 (1984) 1667. [10] J.J. Laserna, A.D. Campiglia, J.D. Winefordner, Anal. Chem. 61 (1989) 1697. [11] C.D. Tran, Anal. Chem. 56 (1984) 824. [12] J.J. Laserna, W.S. Sutherland, J.D. Winefordner, Anal. Acta Chim. 237 (1990) 439. [13] S. Nie, S.R. Emory, Science 275 (1997) 1102. [14] S.R. Emory, S. Nie, Anal. Chem. 69 (1997) 2631. [15] S.R. Emory, W.E. Haskins, S. Nie, J. Am. Chem. Soc. 120 (1998) 8009. [16] A.M. Michaels, M. Nirmal, L.E.A. Brus, J. Am. Chem. Soc. 121 (1999) 9932. [17] A.M. Michaels, J. Jiang, L. Brus, J. Phys. Chem. B 104 (2000) 11965. [18] J. Jiang, K. Bosnick, M. Maillard, L. Brus, J. Phys. Chem. B 107 (2003) 9964. [19] N. Leopold, B. Lendl, J. Phys. Chem. B 107 (2003) 5723. [20] D. Graham, W.E. Smith, A.M.T. Linacre, C.H. Munro, N.D. Watson, P.C. White, Anal. Chem. 69 (1997) 4703. [21] K. Kneipp, H. Kneipp, R. Manoharan, E.B. Hanlon, I. Itzkan, R.R. Dasari, M.S. Feld, Appl. Spectrosc. 52 (1998) 1493. [22] L. Lu, H. Wang, Y. Zhou, S. Xi, H. Zhang, J. Hu, B. Zhao, Chem. Commun. 2 (2002) 144. [23] C.H. Kwon, D.W. Boo, H.J. Hwang, M.S. Kim, J. Phys. Chem. B 103 (1999) 9610. [24] W.B. Lacy, L.G. Olson, J.M. Harris, Anal. Chem. 71 (1999) 2564. [25] Y. Saito, J.J. Wang, D.A. Smith, D.N. Batchelder, Langmuir 18 (2002) 2959. [26] C.L. Leverette, V.A. Shubert, T.L. Wade, K. Varazo, R.A. Dluhy, J. Phys. Chem. B 106 (2002) 8747. [27] J. Hu, B. Zhao, W. Xu, Y. Fan, B. Li, Y. Ozaki, J. Phys. Chem. B 106 (2002) 6500. [28] J. Hu, B. Zhao, W. Xu, Y. Fan, B. Li, Y. Ozaki, Langmuir 18 (2002) 6839. [29] B. Nikoobakht, M.A. El-Sayed, J. Phys. Chem. A 107 (2003) 3372. [30] A. Tao, F. Kim, C. Hess, J. Goldberger, R. He, Y. Sun, Y. Xia, P. Yang, Nano Lett. 3 (2003) 1229. [31] P.C. Lee, D. Meisel, J. Phys. Chem. 86 (1982) 3391. [32] A. Wu, Z. Li, L. Yu, H. Wang, E. Wang, Utramicroscopy 92 (2002) 201. [33] J.J. Laserna, Anal. Chim. Acta 283 (1993) 607. [34] K. Kneipp, H. Kneipp, I. Itzkan, R.D. Ramachandra, S.F. Michael, Chem. Rev. 99 (1999) 2957. [35] W.E. Doering, S. Nie, J. Phys. Chem. B 106 (2002) 311. [36] P. Hildebrandt, M. Stockburger, J. Phys. Chem. 88 (1984) 5935. [37] W. Cheng, S. Dong, E. Wang, Angew. Chem. Int. Ed. 42 (2003) 449.
A simple method for the preparation of ultrahigh sensitivity surface enhanced Raman scattering (SERS) active substrate Gang Wei, Hualan Zhou, Zhiguo Liu, Zhuang Li* State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Science, Renmin Street 5625, Changchun 130022, PR China Received in revised form 9 June 2004; accepted 21 June 2004 Available online 5 August 2004
Abstract By immersing mica modified with cetyltrimethylammonium bromide (CTAB) into the silver colloid, a high efficient surface enhanced Raman scattering (SERS) active substrate was formed within 2 h at room temperature. The limit of detection of the substrate for Rhodamine 6G is up to 110 14 M. Changing the concentration of silver colloid and the treating time, various silver aggregates such as nanocrystals, clusters and films were found, and the SERS spectra of these aggregates were also obtained. The results of SERS revealed that CTAB could accelerate aggregation of the silver colloid and cause great Raman enhancement. Bilayer of CTAB is very important for aggregation of silver colloid and the aggregation extent is the main factor for the enormous enhancement on this substrate. # 2004 Elsevier B.V. All rights reserved. PACS: 82.65.P; 61.43.H; 61.16.C; 82.70.D Keywords: Surface enhanced Raman scattering; Aggregation; Atomic force microscopy; Colloid
1. Introduction Surface enhanced Raman scattering (SERS) is a useful technique resulting in strongly increasing Raman signals from the molecules attached to nanometer sized metallic structures. Since the discovery of SERS, a key problem in analytical application of * Corresponding author. Tel.: +86 431 5262057; fax: +86 431 5262057. E-mail address: [email protected] (Z. Li).
SERS is to develop stable and reproducible SERSactive substrates that can provide a large enhancement factor. The first measurement of a surface Raman spectrum from pyridine adsorbed on an electrochemically roughened silver electrode was reported in 1974 [1]. Then Van Duyme group and Albrecht group measured Raman signal of pyridine on a roughened silver electrode [2–3]. Later other noble metals such as Au and Cu were also used as SERS-active substrates and revealed large enhancement [4–5]. Tian and coworkers generated SERS on net transition
0169-4332/$ – see front matter # 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.apsusc.2004.06.116
G. Wei et al. / Applied Surface Science 240 (2005) 260–267
metals (Pt, Ru, Rh, Pd, Fe, Co, and Ni) by developing various roughening procedures and optimizing the performance of the confocal Raman microscope [6–8]. But unfortunately, these substrates of transition metals still couldn’t be used to detect those samples of ultralow concentration, such as in single molecule level. Metal sols were used to prepare SERS-active substrates widely. It was known that silver colloids showed a higher enhancement factor in comparison with that of Au and Cu, so silver colloids revealed great advantage in studying of SERS and were used widely. Some new active silver substrates have been investigated, including silver-coated latex spheres on filter-paper [9], silver colloid on filter-paper [10–11] and silver-coated filter membrane [12]. The limit of detection for most samples was about 1 10 9 M or mg ml 1 for these substrates. A great breakthrough in studying of Raman was the detection of single molecule [13–15]. Nie and Emory confirmed the existence of Raman enhancement factors on the order of 1014 to 1015 for R6G on colloidal silver particles. By immobilizing a lager number of individual silver particles on a polylysine-coated glass, they obtained the SERS spectrum of single R6G molecule adsorbed on single silver nanoparticle [13]. Later Brus and coworkers repeated the SERS experiments of R6G adsorbed on silver particles [16–18]. They thought the larger SERS enhancement came from colloidal aggregates, such as nanocrystals and clusters. Now, more and more SERS-active substrates are prepared by forming clusters [19–21] and film [22–28] on the solid surface, such as roughened silver film prepared by silver mirror action and silver island film [25–26]. Other substrates such as aggregated gold nanorods [29] and silver nanowire monolayers [30] were used, which also showed good Raman enhancement. In this paper, we provided a simple method to prepare a SERS-active substrate. It was found that silver nanoparticles could aggregate on mica modified with CTAB and form silver nanocrystals, clusters and films by controlling the treating time. The ability of Raman enhancement of the three aggregates was different and the sequence was film > cluster > nanocrystal. SERS (53 numbers) spectra of R6G in 1 10 12 M were obtained on this substrate. The results revealed this substrate was stable and it had excellent reproducibility, and the limit of
261
detection of this substrate for R6G can reach up to 1 10 14 M.
2. Experimental 2.1. Colloid preparation Silver colloids were prepared according to the standard citrate reduction of Lee and Meisal [31]. Briefly, using a heating plate and a 500 ml flask, silver nitrate (90 mg) was added to approximately 400 ml of ultrapure water (18.2 MV) and brought to boiling with rapid magnetic stirring. Ten milliliters 1% sodium citrate was added to this solution. Boiling continued for about 1 h, after which the solution was diluted with triply distilled water to obtain a final volume of 500 ml. 2.2. Preparation and characterization of SERSactive substrates CTAB-coated mica was prepared as followed: a 15 ml aliquot of CTAB (2 10 3 M) was dropped to the new cleaved mica (1 cm 1 cm), then the mica was covered with a piece of parafilm (1.5 cm 1.5 cm) to make the solution expandedness adequately. Fifteen minutes later, the mica modified with CTAB was immersed into the silver colloid for different time. Contact mode AFM imaging was performed on a Digital Instrument Multimode AFM controlled by Nanoscope IIIa (DI, Santa Barbara, CA). The cantilevers were 115 mm in length with Si3N4 tips. All of the images were presented in this article represent flattened data obtained in air. Topographical roughness profiles were extracted from digital image data using a section analysis program. Optical spectra were acquired using UV-2450 spectrophotometer (SHIMADZU, Japan). 2.3. Sample preparation Rhodamine 6G solution was diluted to various concentration from 1 10 9 M to 1 10 15 M with methanol, then 20 ml such solution was dropped to the mica modified with CTAB or polylysine and allowed the solvent to evaporate under ambient conditions.
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2.4. Raman measurements SERS spectra were measured with a Renishaw 2000 model confocal microscopy Raman spectrometer with a CCD detector and a holographic notch filter (Renishaw Ltd., Gloucestershire, UK). The microscope attachment was based on a Leica DMLM system, and a 50 objective was used to focus the laser beam onto a spot with approximately 1 mm in diameter. Radiation of 514.5 nm from an air-cooled argon ion laser was used for the SERS excitation with power of not more than a few mW at the sample position. All of the spectra
reported were the results of a single 30 s accumulation.
3. Results and discussion 3.1. Silver particles aggregates formed on the mica modified by CTAB Several kinds of aggregates formed by immersing mica modified with CTAB into silver colloid for different time. AFM was used to image the actual structure of these various aggregates. Fig. 1a–d shows
Fig. 1. Contact mode AFM images of: (a) single silver nanoparticles, (b) nanocrystals, (c) clusters, and (d) film.
G. Wei et al. / Applied Surface Science 240 (2005) 260–267
the typical AFM images of single silver nanoparticle and the three silver aggregates, nanocrystal, cluster and film, respectively. Fig. 1a shows the image of silver nanoparticles dropped onto cleaved mica directly. Section analysis was used to measure the dimension of about 50 nanoparticles. The particle size was based on height value for tip convolution could affect the accuracy of lateral measurements. The results showed that the particles have an average size of 39.8 nm with a standard deviation of 5.5 nm. With the different treating time, nanocrystals, clusters and films were formed on the mica. When the treating time is 30 min, nanocrystals were formed first. The AFM image shows that this SERS-active particle is a tightly packed aggregate of more than 10 particles (Fig. 1b), which is similar to the studies of Brus and coworkers [16–18], who reported that the nanocrystal is aggregated by 10–15 single nanoparticles. When the treating time is increased to about 1 h, silver clusters are found all over the mica scanned (Fig. 1c). It was found that the clusters are silver aggregates of several hundred silver nanoparticles. We conclude it attributed to the action of CTAB, which balanced the surface charge of silver nanoparticles and made silver nanocrystals reaggregated to form the complex structure. From the image, we find the cluster is not a single
263
nanometer layer completely. Some parts of the cluster are higher than others. These higher and larger particles are the centers of the reaggregation. Moreover, aggregation is irreversible, therefore, the aggregated clusters will become larger and larger. When the treating time is increased to 2 h, we can see there is a visible yellow–gray membrane formed on the surface of mica. AFM image shows much bigger silver clusters were formed. An interesting result is that a more compact and orderly silver nanometer film is found (Fig. 1d), which is similar to the self-assembly of nanoparticles monolayer. These aggregates all show great Raman enhancement effect. In the following section, we will further discuss the issue in detail. As we have shown, aggregation of silver nanoparticles took place on the mica modified with CTAB. We also discuss the effect of different treating time to the silver aggregates in solution. After a 10-fold dilution, silver colloid was mixed with 20 ml CTAB (1 10 3 M). Fig. 2 shows the UV–vis absorbance spectra of the colloid and that of the CTAB aggregated colloid. When treating time was 0 min, there was a sharp peak at about 420 nm. When CTAB was added to the Silver colloid, it revealed a distinct change for the absorbance curve after 1 min, the high and sharp peak changed into a short and wide one. After 1 h, the
Fig. 2. UV–vis absorption changes of colloidal silver (5 ml, diluted for 10-fold) added of 20 ml CTAB (1 10
3
M) in different treating time.
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peak in 420 nm is nearly disappeared. Moreover, a new peak appears at 750 nm, which is attributed to the appearance of silver aggregates. This shows that the bulk property of the colloid has changed due to the addition of the CTAB, which is similar to the study of Graham et al. [20]. 3.2. SERS spectra of R6G on the active substrate R6G samples of ultralow concentration were used to verify the enhancement effect of this SERS-active substrate. Using the confocal microscopy Raman system, we obtained a laser beam spot with diameter of 1 mm. It is easy to measure a SERS spectrum of sample with scales of a few micrometers. With 50 objective and 10 ocular, we can observe the least silver aggregates is about 400 nm, so it is possible for us to obtain SERS spectra after we have gained the AFM images on the same area of the sample. We have applied a novel relocated technique for biology molecules by AFM [32]. Using this relocated technique, we can obtain the SERS spectra and AFM images of the Silver clusters and films on the same location. Fig. 3a– c shows the SERS spectra of R6G adsorbed onto the three aggregates film, cluster and nanocrystal, respectively. The nanocrystal is compact aggregate of more than 10 nanoparticles, so there are 4–5 nanocrystals in
an area of 2 mm 2 mm size. These disturbs that the spectra are not the real SERS spectra of single nanocrystal, but the multiple action of several nanocrystals. Different SERS signals were found for these aggregates. At the same experiment conditions (integration time: 30 s, power: 10%), films revealed greater SERS signal than other aggregates. Dropping silver nanoparticles onto the cleaved mica directly, SERS spectrum shows the lowest concentration of R6G can be detected is only about 1 10 9 M on this substrate, moreover the stability and reproducibility are poor. But for the Ag film, the greatest enhancement effect is observed and the limit of detection is up to 1 10 14 M. Even if we made the laser power reduced to 1%, an interpretable Raman signal is still obtained (dada no shown). So we deduce that the aggregation extent of silver colloid could affect SERS signals greatly. SERS (53 numbers) spectra of R6G (1 10 12 M) were also obtained. Fig. 4 displays the peak heights of the 1650 cm 1 line for the 53 SERS spectra. In the histogram, 39 of the 53 spectra show great Raman signal (>200 c/s), and only one spectrum shows low Raman signal (<10 c/s). Because the selection of the location measured is random, we conclude that this substrate is stable and can be used to detect those samples of ultralow concentration.
Fig. 3. SERS spectra of R6G obtained on: (a) films, (b) clusters, (c) nanocrystals, and (d) nanoparticles. Experiment conditions: for (a)–(c), the concentration of R6G was 1 10 12 M, Laser power was 25 mW (1%).
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Fig. 4. Peak height of 1650 cm integration time: 30 s.
1
line for the 53 SERS spectra obtained when the concentration of R6G was 1 10
3.3. Aggregation extent is the main factor for the enormous enhancement Despite extensive studies of SERS since its discovery nearly many years ago and a literature of thousands of papers, a complete understanding of the mechanism of enhancement was lacking. SERS has historically been described in terms of electromagnetic and chemical or ‘‘first layer’’ enhancement mechanisms [33–34]. SERS intensity was found to be stronger for adsorbed molecules on surfaces covered with aggregated nanoparticles, the observed increase in the SERS intensity in the case of aggregation was attributed to the enhancement of electric field between particles in the aggregates [17,18,34,35]. Nanoparticles aggregates have been attracting growing interests for their collective electronic, optical, and magnetic properties. It was revealed that halide ions such as Cl and I were often used as aggregating agents [35–37], they appeared to reduce the surface charge on the colloid, lower ‘‘j-potential’’, then paving way for aggregation of nanopartilces [37]. In Nie’s studies about single molecule, they found there were only a few particles could show great Raman enhancement and claimed that the great
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12
M. Power: 25 mW, 1%;
enhancement was attributed to ‘‘hot’’ nanoparticles [13–15,35]. In our experiment, we think that aggregation extent and dimension of particles are two key factors for the enhancement ability. Great SERS signals have been obtained on the aggregates of nanocrystals, clusters and films. And SERS signals increased with the increase of degree of aggregation. In theory, when concentration of R6G is 2 10 11 M, there is only one R6G molecule in 1 mm2, but the concentration of R6G is reduced to 1 10 14 M, we can still gain a clear SERS spectrum. So we can conclude that R6G was not absorbed to the SERSactive substrate averagely but absorbed to the hot sites provided by aggregates. But how silver nanoparticles were aggregated and formed various aggregates on the CTAB-coated mica? We proposed electrostatic interaction is the main force to cause aggregation. A control experiment was applied to testify the supposition. CTAB was diluted to 5 10 4 M, 1 10 4 M, 1 10 5 M and the substrates were prepared as experiment section. AFM images showed there were sparse nanoparticles adsorbed, even nearly no aggregate was observed (data not shown). Fig. 5a–c shows the SERS spectra of R6G (1 10 12 M) adsorbed on these substrates. When concentration of CTAB is 5 10 4 M, the
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Fig. 5. SERS spectra of R6G (1 10 12 M) on three different substrates. The concentration of CTAB for the preparation were: (a) 5 10 (b) 1 10 4 M, and (c) 1 10 5 M. Laser power: 25 mW, 1%; integration time: 30 s.
Scheme 1. Schematic binding mechanisms of CTAB adsorbed to cleaved mica.
4
M,
spectrum shows greatest SERS signal (Fig. 5a). But there is no SERS signal observed when concentration of CTAB is reduced to 1 10 5 M (Fig. 5c). The possible explanation is that for the surface of mica is hydrophilic and electronegative, so the hydrophilic cation group (–NH4+) of CTAB can be easily bound to the surface of mica and the hydrophobic group (CH3(CH2)n–) faces liquid phase (Scheme 1a). But when the concentration of CTAB is higher than its critical micelle concentration (about 0.94 mM), bilayer of CTAB was formed on mica. Two CTAB molecules are linked on the hydrophobic groups and a positive-charge surface is formed (Scheme 1b). On the other hand, we must consider the surface of mica cannot be coated by CTAB enough if the concentration of CTAB were too low, which made silver nanoparticles couldn’t adsorb onto the CTAB-coated mica adequately. Compared with the method of mixing aggregation reagents with silver colloid and dropping onto a solid substrate, the technique provided in this report is more applied and this substrate shows greater enhancement effect.
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4. Conclusions In summary, we have presented a simple method to prepare SERS-active substrate, which showed greater surface enhanced Raman scattering signals than those substrates made by the conventional method. The limit of detection for R6G can add up to 1 10 14 M and the stability and reproducibility are very good. Making use of the relocate technique, we not only obtained the AFM images of silver nanocrystal, cluster and film, but also gained the SERS spectrum of single R6G molecule on the same aggregate. The great enhancement depended on the size of silver nanoparticles and degree of aggregation. The driving force for forming these different aggregates must be electrostatic interactions in this study, which provided by bilayers of CTAB. Controlling aggregation of silver nanoparticles by changing the treating time and the concentration of CTAB, it is more significant to unveil mechanism of SERS. We hope this preparation of SERS-active substrate can be used to investigated the interaction of drug ligand with DNA at very low concentration. Further study about this is processing.
Acknowledgements This work was supported by the National Natural Science Foundation of China (no. 30070417).
References [1] M. Fleischman, P.J. Hendra, A.J. McQuillan, Chem. Phys. Lett. 26 (1997) 123. [2] D.L. Jeanmaire, R.P.V. Duyne, J. Electroanal. Chem. 84 (1977) 1. [3] M.G. Albrecht, J.A. Creighton, J. Am. Chem. Soc. 99 (1977) 5215. ¨ . Ertu¨ rk, A. Otto, Surf. Sci. 179 (1987) 163. [4] U [5] H. Baltruschat, E. Rach, J. Heitbaum, J. Electroanal. Chem. 194 (1985) 109. [6] Z.Q. Tian, B. Ren, D.Y. Wu, J. Phys. Chem. B 106 (2002) 9463. [7] Z.Q. Tian, B. Ren, B.W. Mao, J. Phys. Chem. B 101 (1997) 1338.
267
[8] D.Y. Wu, Y. Xie, B. Ren, J.W. Yan, B.W. Mao, Z.Q. Tian, Phys. Chem. Commun. 18 (2001) 1. [9] T. Vo-Dinh, M.Y.K. Hiromoto, G.M. Begun, R.L. Moody, Anal. Chem. 56 (1984) 1667. [10] J.J. Laserna, A.D. Campiglia, J.D. Winefordner, Anal. Chem. 61 (1989) 1697. [11] C.D. Tran, Anal. Chem. 56 (1984) 824. [12] J.J. Laserna, W.S. Sutherland, J.D. Winefordner, Anal. Acta Chim. 237 (1990) 439. [13] S. Nie, S.R. Emory, Science 275 (1997) 1102. [14] S.R. Emory, S. Nie, Anal. Chem. 69 (1997) 2631. [15] S.R. Emory, W.E. Haskins, S. Nie, J. Am. Chem. Soc. 120 (1998) 8009. [16] A.M. Michaels, M. Nirmal, L.E.A. Brus, J. Am. Chem. Soc. 121 (1999) 9932. [17] A.M. Michaels, J. Jiang, L. Brus, J. Phys. Chem. B 104 (2000) 11965. [18] J. Jiang, K. Bosnick, M. Maillard, L. Brus, J. Phys. Chem. B 107 (2003) 9964. [19] N. Leopold, B. Lendl, J. Phys. Chem. B 107 (2003) 5723. [20] D. Graham, W.E. Smith, A.M.T. Linacre, C.H. Munro, N.D. Watson, P.C. White, Anal. Chem. 69 (1997) 4703. [21] K. Kneipp, H. Kneipp, R. Manoharan, E.B. Hanlon, I. Itzkan, R.R. Dasari, M.S. Feld, Appl. Spectrosc. 52 (1998) 1493. [22] L. Lu, H. Wang, Y. Zhou, S. Xi, H. Zhang, J. Hu, B. Zhao, Chem. Commun. 2 (2002) 144. [23] C.H. Kwon, D.W. Boo, H.J. Hwang, M.S. Kim, J. Phys. Chem. B 103 (1999) 9610. [24] W.B. Lacy, L.G. Olson, J.M. Harris, Anal. Chem. 71 (1999) 2564. [25] Y. Saito, J.J. Wang, D.A. Smith, D.N. Batchelder, Langmuir 18 (2002) 2959. [26] C.L. Leverette, V.A. Shubert, T.L. Wade, K. Varazo, R.A. Dluhy, J. Phys. Chem. B 106 (2002) 8747. [27] J. Hu, B. Zhao, W. Xu, Y. Fan, B. Li, Y. Ozaki, J. Phys. Chem. B 106 (2002) 6500. [28] J. Hu, B. Zhao, W. Xu, Y. Fan, B. Li, Y. Ozaki, Langmuir 18 (2002) 6839. [29] B. Nikoobakht, M.A. El-Sayed, J. Phys. Chem. A 107 (2003) 3372. [30] A. Tao, F. Kim, C. Hess, J. Goldberger, R. He, Y. Sun, Y. Xia, P. Yang, Nano Lett. 3 (2003) 1229. [31] P.C. Lee, D. Meisel, J. Phys. Chem. 86 (1982) 3391. [32] A. Wu, Z. Li, L. Yu, H. Wang, E. Wang, Utramicroscopy 92 (2002) 201. [33] J.J. Laserna, Anal. Chim. Acta 283 (1993) 607. [34] K. Kneipp, H. Kneipp, I. Itzkan, R.D. Ramachandra, S.F. Michael, Chem. Rev. 99 (1999) 2957. [35] W.E. Doering, S. Nie, J. Phys. Chem. B 106 (2002) 311. [36] P. Hildebrandt, M. Stockburger, J. Phys. Chem. 88 (1984) 5935. [37] W. Cheng, S. Dong, E. Wang, Angew. Chem. Int. Ed. 42 (2003) 449.