Anal. Chem. 1903, 65, 3177-3186
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Physical Structure, Optical Resonance, and Surface-Enhanced Raman Scattering of Silver-Island Films on Suspended Polymer Latex Particles Paula A. Schueler,+Jeffrey T. Ives? Fern DeLaCroix,? William B. Lacy? Patricia A. Becker,t Jianmin Li,t Karin D. Caldwel1,S Barney Drake,ll and Joel M. Harris**$ Research and Development, Boehringer Mannheim Corporation, P.O.Box 50457, 91 15 Hague Road, Indianapolis, Indiana 46250-0457, Department of Chemistry, Henry Eyring Building, University of Utah, Salt Lake City, Utah 84112, Department of Bioengineering, 2260 Merrill Building, University of Utah, Salt Lake City, Utah 84112, and Imaging Services, 520 East Montecito Street, Suite K , Santa Barbara, California 93101
A new approach to producing SERS activity on dispersed particles is described. Silver islands are deposited onto submicron latex and glass particles, and the particles a r e dispersed into aqueous suspensions. The physical structure of the metal islands is investigated by transmission electron and atomic force microscopies and compared with the optical absorption and SERS efficiencies for several particulate substrates and silver deposition methods. Thermal vapor deposition onto poly(methyl methacrylate) particles produced a latex that supported silver islands with optical properties and SERS activities similar to silver-island films deposited on flat surfaces. Like silver colloids, the material could be handled as a n aqueous suspension to generate SERS from a dispersed surface; unlike silver colloids, however, the optical and physical properties of the material were stable upon storage for periods as long as several months. INTRODUCTION Surface-enhanced Raman scattering (SERS) has been applied to the detection of surface-adsorbed or bound + Boehringer Mannheim Corp. t
Department of Chemistry, University of Utah.
I Department of Bioengineering, University of I Imaging Services.
Utah.
0003-2700/93/0365-3177$04.00/0
molecules on a variety of discontinuous metal structures. Enhancement factors of 105-106 can be observed in coinage metals (gold, silver, copper) when structural features on a nanometer scale are present and surface plasmon resonance conditions are satisfied.' Earliest examples utilized electrochemically roughened silver electrodes,Z which have the advantage of generating a renewable metal surface, but the disadvantage of requiring an electrochemical cell for measurement. Metal-island films prepared by slow evaporation of a metal onto a dielectric substrate have also been used extensively to provide strong SERS enhancement.3 The disadvantage of both of these approaches relates to the small surface area and resulting limited capacity of a flat surface for binding or adsorbing molecules. Furthermore, adsorption kinetics from a dilute solution to a flat surface are slow, limited by diffusion through a stagnant layer of solvent near the surface.4~5With flat surfaces, therefore, the dynamic range of response for high concentrations is limited while response time for low concentrations is slow. In contrast, dispersed particles have a much greater surface area and exhibit faster adsorption kinetics than flat surfaces4~6 and would be an attractive alternative for SERS studies. Considerable effort has been directed toward generating, controlling, and applying suspensions of metal colloids for (1) Moskovita, M. Rev. Mod.Phys. 1985,57,783. (2) Fleischmann, M.; Hendra,P. J.;McQuillan, A. J. Chem.Phys.Lett. 1974, 26, 163. (3) Chang, R. K.; Furtak, T.E.Surface Enhanced Raman Scattering; Plenum: New York, 1982. (4) Hechemy, K.; Michaelson, E. Lab. Manage. 1984,6,26; 1984,7,26. ( 5 ) Wolf, S. Nucleic Acids Res. 1987, 15, 2911.
0 1993 American Chemical Society
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SERS-based detection.Unfortunately, t h e stability of metal colloid suspensions is strongly influenced by temperature, ionic strength, p H , and the presence of adsorbates in solution; furthermore, aggregation of t h e metal colloid is required t o generate a long-wavelength plasmon resonance with strong S E R S enhancement. As aggregates grow in size, the enhancement shifts to longer wavelengths over time, from minutes t o hours, depending on t h e kinetics of aggregation.8~9 Despite attributes of higher surface area and better adsorption kinetics, problems with stability and reproducibility have inhibited the use of dispersed metal colloids in routine analytical applications of SERS. An ideal particulate substrate for SERS activity would produce stable suspensions of dispersed particles t o provide the advantages of increased surface area and faster kinetics. T h e surfaces of t h e particles should be able to be modified to allow for subsequent adsorption or binding studies without destroying t h e stability of t h e particle suspension or t h e plasmon resonance. T h e present work describes a new approach t o producing SERS activity on dispersed particles by depositing silver islands on submicron latex and glass particles and suspending t h e particles in solution. T h e physical structure of the metal islands, their optical absorption, and SERS efficiencies are compared for several substrates and deposition methods. With the correct evaporation conditions and t h e appropriate substrate surface, t h e silverisland-coated latexes showed optical properties and SERS activities similar to silver-island films deposited on flat surfaces.
EXPERIMENTAL SECTION Reagents. All chemicals were used without further purification. Poly(methy1 methacrylate) (PMMA) and carboxylatemodified poly(methy1 methacrylate) (PMMA-CML) latex particles with diameters of 309 and 353 nm, respectively, were purchased from Seradyn Inc. (Indianapolis, IN). The size distributions of the latex particles were measured by scanning electron microscopy; the average particle diameters were confirmed, and the relative standard deviation of the particle size distribution was 7 % . Silver (99.9999% purity) was obtained from Aldrich (Milwaukee, WI) and from Engelhard Corp. (Edison, NJ). Sodium polytungstate (BioChemika No. 71913) was obtained from Fluka (Ronkonkoma, NY). Water was deionized, distilled, and Milli-Q filtered (Millipore, Bedford, MA) unless otherwise specified. Glass microscope slides were obtained from Baxter Diagnostics (No. M6158-1). Preparation of Latex Particles for Silver Deposition. A 0.3-mL aliquot of 10% PMMA latex solids in water was spun cast (Model No. l-EClOlD-R485; Headway Research Co., Garland, T X ) onto ethanol-washed glass microscope slides (26 mm by 78 mm). Slides were spun at 2000 rpm for 40 s. Latex-coated slides were dried in air and vacuum sealed in high-density polyethylene slide carriers. Deposition of Silver onto Latex Particles. Glass slides bearing a layer of spun-coated, dried latex particles were used as a substrate for deposition of silver-island films. Islands were deposited a t three different laboratories using two different evaporation techniques. Method 1. In a conventional vacuum chamber (Edwards Model E306A), latex-coated slides were fixed 20 cm above a molybdenum boat (Edwards H014-01-044) containing silver. The chamber was evacuated to a pressure of (1.5-3 X Torr, and the silver was vaporized by passing a current of approximately 25 A through the refractory boat. The mass of silver deposited was monitored using a quartz crystal oscillator thickness gauge (Edwards FTM5) ( 6 ) Pettinger, B.; Krischer, K. J . Electron Spectrosc Relat. Phenom. 1987, 45, 133. (7) Silman, 0.;Bumm, L. A.; Callaghan, R.; Blatchford, C. G.; Kerker, M. J . Phys. Chem. 1983,87, 1014. (8) Montes, R.; Contreras, C.; Ruperez, A,; Laserna. J. J. Anal Chem 1992, 64, 2715. (9) Henglein, A.; Mulvaney, P.;Linnert. T. Faraday Discuss 1991,92, 31.
located 17 cm above the boat (an r2distance correction was used to estimate the thickness of silver deposited on the particles). The rate of deposition was approximately 0.2 Ais as measured by the thickness gauge. Nominal silver mass thickness was 7080 A onto a flat surface a t the same distance from the source as the particles. Method 2. Silver films were deposited at Denton Vacuum Inc. (Cherry Hill, NJ) using aconventionalvacuum chamber and a resistance heated source. Latex-coated slides were located 45 cm from the molybdenum boat containing the silver. The chamber pressure was nominally 5 X 10-6 Torr. The mass of silver deposited was monitored using a quartz crystal oscillator thickness gauge (Inficon-XTC, Leybold-Heraeus, Germany) nested between the latex-coated slides. Silver mass thickness was 70 i= 5 A and was deposited over a period of 100-200 s corresponding a deposition rate of 0.4-0.8 Ais. Method 3. The silver films were deposited in a Airco-Temescal Box Coater Unit equipped with two, multipocket, shuttered, electron guns. The unit contains two Inficon quartz crystal film thickness monitors that were used to track the film thickness and control each of the guns. The latex-coated slides were mounted on a flat plate rotating fixture and rotated a t 15 rpm. A refractory crucible liner housed the high-purity silver. Typical Torr with an evaporation rate system base pressure was 5 X of 0.5-1 A/s. Film thickness calibration was accomplished by evaporating relatively thick silver films (- 2000A) and measuring the thickness with a profilometer instrument (Dektak 11). The sample-to-crystal setting was then adjusted to achieve a silver coating thickness of 30 A. Suspension of Silver-Island Latex Particles. Latex-coated slides containing deposited silver vapor were flooded with water to wet the entire slide. The silver-coated latex was loosened from the glass surface with a rubber spatula and then rinsed with water into a 50-mL polypropylene centrifuge tube (Corning No. 25331). Most of the free silver metal was removed by allowing the silver not bound to the latex to adhere to the polypropylene tube surface. The supernatant containing silver-islandlatex was subsequently transferred to 30-mL glass Corex tubes. Solutions were centrifuged a t 7500 rpm for 10 min in a Sorval HB-4 swinging bucket rotor (9OOOg) and RC5C centrifuge set a t 4 "C. The samples were washed a minimum of twice with water, and the final pellet was resuspended in water to a total volume corresponding to 1 mL times the number of slides processed. The samples were then subjected to 1G-20 s of gentle bath sonication (Bransonic Model No. 2200) to break up the macroscopic aggregates. Sodium Polytungstate Density Gradients. Aqueous sodium polytungstate gradientslo were poured stepwise to give an approximate density range of 1.0-1.6 gimL. Gradients were prepared in Ultra-Clear tubes (12.5 X 50 mm, Beckman No. 344057) with a total volume of 4 mL, and 0.25-1.0 mL of particle suspension was applied to the gradient. Centrifugation was performed in a Beckman L8-70M ultracentrifuge in an SW55Ti rotor for 1h at 30 000 rpm and 18 "C. Gradients were evaluated immediately due to visible aggregation of PMMA sample with increasing time. Sedimentation Field Flow Fractionation (sedFFF). The sedimentation field flow fractionation channel was formed by two sheets of highly polished Hastelloy C stainless steel separated by a 0.254-mm-thick Mylar sheet. The columnar space is cut out from this sheet and has dimensions of 94 X 2 X 0.254 mm. The measured channel void volume was 5.0 mL. The channel was then curved to fit inside a rotor basket allowing it to spin a t a preset centrifugal acceleration, which can be considered constant across the thin channel. Degassed, deionized water (DI water) as a carrier was delivered to the system through a PC-controlled Minipulse 3 Gilson pump with a 2.8 mL/min flow rate. A UV absorbance detector (Linear, Model No. 106)with a 254-nm light source was used for monitoring the eluent. Detailed information regarding the structure and theory of the sedFFF system has been published previously.11
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(10) Gregory, M. R.; Johnston, K. A. N. 2. J . Geol. Geophys. 1987,30,
317. (11)Caldwell, K. D. Anal. Chem. 1988,60,959A. Caldwell, K. D.; Li. J . M.; Li, J. T.; Dalgliesh, D. G. J . Chromatogr. 1992, 604, 63.
ANALYTICAL CHEMISTRY, VOL. 65, NO. 22, NOVEMBER 15, 1993
For sedFFF system calibration, polystyrene latex (d = 272 nm, p = 1.05 g/mL, Seradyn Inc.) was diluted to 1% (w/v) with DI water and analyzed in 0.1 % FL-70 detergent (FisherScientific, Fair Lawn, NJ) solution. A 25-pL aliquotof suspendedAg-coated PMMA particles (0.05 % w/v) was injected with a syringedirectly into the separation chamber while the rotor was held stationary. After a 30-8 delay, the rotor was accelerated to 450 rpm (35g). Transmission Electron Microscopy (TEM). Silver-coated latex samples were concentrated using an Eppendorf 5414 centrifuge, operating at 14 OOO rpm for 10 min; the supernatant liquid was subsequentlyremoved. The concentratedsuspensions were then dropped on Formvar film-coatedstandard copper grids (200 mesh) at room temperature. After 1min, excess fluid was removed using fiiter paper and the grid was allowed to air-dry. There was no staining of the sample before TEM observation. The microscope used was a JEOL-100CX I1 operated under 80kV acceleration voltage. Photographs were taken with at least 2oooOX magnification. Tapping-Mode Atomic Force Microscopy (AFM). Sample preparation of coated and uncoated latex spheres consisted of air-drying 20-40 pL of a concentrated (- 10% ) solution onto 12 X 12 mm2 pieces of bare silicon wafer. Air-drying typically required 1-3 h. Sampleswere imaged using a Digital Instruments NanoScope 111MultiMode atomic force microscope (AFM) used in “tapping mode”. Single-crystal, 120-pm-long silicon cantilevers (Digital Instruments) with spring constants of 20-50 N/m were used on all samples. Typical cantileverresonant frequenciesranged from 300 to 360 kHz. The operation of contact and noncontactAFMs has been described in detail in previous publications,12and the use of tapping mode AFM to image fragments of a low-modulus polymer has recently appeared.** In tapping mode, a relatively stiff cantilever is driven into Oscillation near its natural resonant frequency. This resonating cantilever is then brought down to within 5-10 nm from the surface so the tip will touch the sample at the bottom of its oscillation. As the cantilever comes in contact with the sample surface,the amplitude of the oscillation is reduced to a fiied ‘set point”value which is held constant as the sampleis raster scanned beneath the cantilever. Similar to standard contact-modeAFM, the tapping mode AFM feedback circuit holds the amplitude of the cantilever oscillation constant as the sample’s surface topography is mapped out. This hybrid mode of both partial contact and noncontact AFM allows for high-resolutionimaging vertically and laterally,while no shear or lateral forces are applied to the sample. Typical forces applied to the sample are 0.1-1 nN. Determination of Optical Resonance of Silver-Island Latex Particles. Absorbance scans were performed on SUBpended silver-island particles using a Beckman DU-7500 photodiode array spectrophotometer (at Boehriiger Mannheim) or a Hewlett-Packard8542 (at the University of Utah, Department of Chemistry). A suspension of uncoated latex particles scattermatched at 300 nm was scanned with each preparation for comparison. Surface-Enhanced Raman Scattering Using Silver-Island Latex Particles. The Raman instrument used for the SEW measurements was configured as follows: Raman scatter was generated by laser excitation of the sample at 514.5 nm, collected at 90°, focused into a monochromator, and detected with a charge-coupleddevice (CCD). A detailed description of the instrument has been presented previ0usly.1~The following modifications were made for the pyridine adsorption experiments: a 50-mW beam was focused with a plano-convex lens cf = 75 mm) onto the sample, which was contained in a quartz microfluorometer cell (250 pL, Type 607, NSG Precision Cells, Inc.); the beam was directed off the front face of the sample cell at 57O. For the PMMA substrate experiments,a 250-mW beam was focused with a cylindrical lens cf = 75 mm) onto the sample, which was contained in a quartz fluorometer cell (490 pL, Type 21, NSG Precision Cells, Inc.). The long axis of the cylindrically focused light was parallel to the spectrometer slit, and the cell (12)Rugar, D.; Hansma, P. Phys. Today 1990, Oct,23. (13)Zhong, Q.;Inniss, D.; Kjoller, K.; Elings, V. B. Surf. Sci. Lett. 1993,290, L688. (14)Lacy,W. B.; Rowlen, K. L.; Harris,J. M.Appl. Spectrosc. 1991, 45,1598.
317Q
was tilted so the incident light struck the front face at an angle of 60” relative to the surface normal. In both experiments, scattered light from the sample cell was collectedand collimated by anfll.4 camera lens (25mm, JML Optics) and focused by an achromatic doublet cf = 100.00 mm, Newport Corp.) into the entrance slit of the monochrometer. The slit width of the monochromator was 50 pm for the pyridine adsorption data and 100 pm for the PMMA substrate study. A holographic notch filter (HNF-514-1.0, Kaiser Optical Systems, Inc.) for the 514.5nm line was placed between the collection and focusing leto eliminate Rayleigh scatter interference. A 1200-groove/mmgrating dispersed the scattered radiation across a Thomson TH 7882 CCD chip (Series 200, Photometrics, Ltd., Tuscon, AZ). The temperature of the CCD chip was -120 f 0.1 “C. The format of the silicon/metaloxide chip is 576 row8 by 384 columns. The short axis of the chip was oriented parallel to the direction of wavelength dispersion, and the long axis along the slit of the monochrometer. For all spectra, 576 rows were ‘binned” (on-chip signal averaging) along the vertical axis to improve the signal-to-noise ratio. Spectra for the pyridine experiments were taken in groups of five and subsequently coadded to improve the signal-to-noise ratio and aid in data manipulation. Data were processed off-line on a -based PC running Lotus 1-2-3and Matlab. Peak modeling was accomplished by a xa minimization routine constructed in Lotus 1-2-3 and subsequent matrix linear least-squares fitting in Matlab (using fixed, nonlinear peak parameters obtained from Lotus 1-2-3). Prior to Raman measurements, the absorbance spectra of suspensionsof dispersed latex particleewere measured on adiode array spectrometer (Mode18542,Hewlett Packard). Suspensions were diluted in l&MQ water (quartz-distilled and filtered; Nanopure 11, Barnstead) until the scattering at 300 nm was matched for all samplesto assure equal concentrationsof PMMA. The pyridineisothermexperimentswere all performed on samples from the same batch of PMMA substrate prepared by method 2 and immersed for at least 5 min in pyridine (Baker)/l&MQ water solutions.
RESULTS AND DISCUSSION The high surface area and efficient transport for rapid binding kinetics make dispersed particulate surfaces attractive for many analytical chemistry applications. To bring these attributes to SERS measurements, small (300 nm) polymer latex particles were coated with silver, and the SERS activity of the material was investigated. Due to the unique challenge of manipulating these particles and depositing silver onto their surfaces, the physical structure and optical properties of the material were also characterized to establish the physical basis of their performance and understand differences between the three different silver deposition methods. Quantifying Silver on Latex Particles. SERS signals are related to the local electromagnetic fields near a discontinuous metal surface. For metal-island fiis formed by vapor deposition, the surface electromagnetic fields depend on structure of the islands and interactions between them. The extent of silver coverage on each particle and the distribution of coverages among the population of particles, therefore, influence the SERS activity. Due to the deposition method used in this work, whereby particles were immobilized on a flat surface and then coated with silver by vapor deposition, an approximate hemispherical shell or half-shell of silver islands is expected on each particle. Depending on the packing density of immobilized particles, some particles may receive little or no silver due to shielding by neighboring or overlying particles. Sodium polytungstate density gradients were used to resolve silver-coated latex from uncoated latex and free silver colloid, to determine the average density of the coated material. Separation by sodium polytungstate gradients relies on the large density difference between silver (10.49 g/mL) and bulk PMMA (1.19 g/mL). Figure 1shows an example of the gradient separations. The density gradients were linear (Ra
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Elution volume (mL) Figure 2. SedFFF elution profile of method 1- and Pdeposited PMMA latexesand bare PMMA latex in water. The higherdensity PMMA particles after method 1 and 2 deposition (indicated by the numbers 1 and 2) elute later than the bare PMMA.
Figure 1. Photographs of sodium polytungstate density gradients. Samples are designated by the number labeled under each tube, and the associated sample is described in the following text: (A, top) (1) Method ldeposited PMMA, (2) and (3) two different preparations of method 2deposited PMMA, (4) method ldeposited PMMA-CML, (5) bare PMMA latex, and (6) bare PMMA-CML latex. The bare latex position is designated by the upper arrow and the band of highest density by the lower arrow. (6, bottom) (7) Method 3deposited PMMA latex, (8) bare PMMA latex, and (9) and (10) a different run of the same two samples of method 2deposited PMMA latex as in (2) and (3).
= 0.96, N = 4) from 1.0 to 1.5 g/mL. All the deposition methods on PMMA and PMMA-CML (only method 1 was tested on PMMA-CML)yield approximatelythe same pattern of bands in these gradients. The primary band for each of the silver-coated latex preparations was found at one distinct density level: 1.21 g/mL for method 3 (electron-beam sputtering) on PMMA and 1.27 g/mL for methods 1 and 2 (thermal vapor deposition). In each case, a band corresponding to uncoated latex was also present a t a density of 1.14g/mL. For method 3, the uncoated latex band comprised 50 % of the total sample loaded on the gradient; in all other preparations, this band corresponded to
