Sequential Monitoring of Film Thickness Variations with Surface

Anal. Chem. 2008, 80, 891-897

Sequential Monitoring of Film Thickness Variations with Surface Plasmon Resonance Imaging and Imaging Ellipsometry Constructed with a Single Optical System Yong-Jun Li, Yi Zhang, and Feimeng Zhou*

Department of Chemistry and Biochemistry, California State University, Los Angeles, Los Angeles, California 90032

Sequential surface plasmon resonance imaging (SPRI) and imaging ellipsometry (IE) measurements are realized with a single set of optical components mounted on a goniometer. The sample situated on top of a triangular prism is positioned in the center of the goniometer. The resultant setup (SPRI/IE) can be used to examine the same surface region above (IE) and below (SPRI) the sample. Thickness values of silver stripes sputtered onto a gold substrate were determined by SPRI and IE, and the results were compared to establish the validity of the method. The SPRI/IE setup was also used to monitor the thickness of phospholipid films of different layer numbers. SPRI measurements were found to be more accurate for ultrathin films, whereas the IE results are more reliable for films whose thicknesses approach or exceed the distance encompassed by the evanescent wave of the surface plasmon. Thus, utilizing these two techniques sequentially facilitates the continuous monitoring of film thickness variation over a wide thickness range with high fidelity and provides a viable approach to image the surface regions with large topographic fluctuations. Interfaces and thin films play a central role in catalysis, coatings, semiconductor and electronic devices, biosurfaces, and sensors. Many techniques have been developed to characterize thin films and to monitor events at interfaces. These techniques include, but are not limited to, atomic force microscopy (AFM),1 profilometry (contact2,3 or optical4), X-ray reflectivity and diffraction,5 quartz crystal microbalance,6 surface plasmon resonance (SPR),7,8 and ellipsometry.9 Most of these techniques can deter* Corresponding author. Phone: 323-343-2390. Fax: 323-343-6490. E-mail: [email protected]. (1) Giessibl, F. J. Rev. Mod. Phys. 2003, 75 (3), 949-983. (2) Chappard, D.; Degasne, I.; Hure, G.; Legrand, E.; Audran, M.; Basle, M. F. Biomaterials 2003, 24 (8), 1399-1407. (3) Brown, C. A.; Savary, G. Wear 1991, 141 (2), 211-26. (4) Power, J. F. Rev. Sci. Instrum. 2002, 73 (12), 4057-4141. (5) Rafaja, D. Festkor. Adv. Solid State Phys. 2001, 41, 275-286. (6) O’Sullivan, C. K.; Guilbault, G. G. Biosens. Bioelectron. 1999, 14 (8-9), 663-670. (7) Homola, J. Surface Plasmon Resonance Based Sensors; Springer: New York, 2006. (8) Rothenhausler, B.; Knoll, W. Nature 1988, 332 (14), 615-617. (9) Azzam, R. M. A.; Bashara, N. M. Ellipsometry and Polarized Light; NorthHolland Publishing Company: New York, 1977. 10.1021/ac701873v CCC: $40.75 Published on Web 01/01/2008

© 2008 American Chemical Society

mine ultrathin film thickness down to the nanometer or subnanometer resolution. Among them, AFM and X-ray-based techniques are capable of measuring thickness at the atomic level. However, the measurements are either limited to a small surface area or require prior knowledge about the sample structure. Profilometry is widely used and can scan across a large surface area, but the sensitivity of a typical profilometer is not as high as those of AFM and X-ray-based techniques. SPR and ellipsometry are two optical methods which are nondestructive and possess sensitivity down to the nanometer resolution.7,8,10-15 Surface plasmon (SP) can be regarded as a charge-density wave excited by light at a metal-dielectric interface. The coupling between photons and surface plasmon is referred to as SPR. In the Kretschmann configuration,16 light polarized in the incidence plane (p direction) enters a prism of a high refractive index and irradiates onto the back of a glass slide whose top is coated with a thin layer of metal film. The SPR results in a minimum in the reflected light beam. The evanescent field associated with SP decays exponentially into the metal and the dielectric. Although this evanescent field is extremely sensitive to changes occurring in the vicinity of the interface, SPR becomes insensitive to film variation that occurs out of the evanescent field. For SPR imaging (SPRI),8,12,13,17-24 the light source is expanded (10) Arwin, H. Thin Solid Films 2000, 377-388, 48-56. (11) Lee, H. J.; Nedelkov, D.; Corn, R. M. Anal. Chem. 2006, 78 (18), 65046510. (12) Wolf, L. K.; Fullenkamp, D. E.; Georgiadis, R. M. J. Am. Chem. Soc. 2005, 127 (49), 17453-17459. (13) Wang, Z.; Wilkop, T.; Cheng, Q. Langmuir 2005, 21 (23), 10292-10296. (14) Boozer, C.; Ladd, J.; Chen, S.; Jiang, S. Anal. Chem. 2006, 78 (5), 15151519. (15) Yao, X.; Li, X.; Toledo, F.; Zurita-Lopez, C.; Gutova, M.; Momand, J.; Zhou, F. Anal. Biochem. 2006, 354 (2), 220-228. (16) Kretschmann, E. Z. Phys. 1971, 241, 313-324. (17) Flatgen, G.; Krischer, K.; Pettinger, B.; Doblhofer, K.; Junkes, H.; Ertl, G. Science 1995, 269 (5224), 668-671. (18) Homola, J.; Yee, S. S.; Gauglitz, G. Sens. Actuators, B 1999, 54 (1-2), 3-15. (19) Johansen, K.; Arwin, H.; Lundstrom, I.; Liedberg, B. Rev. Sci. Instrum. 2000, 71 (9), 3530-3538. (20) Li, Y.-J.; Oslonovitch, J.; Mazouz, N.; Plenge, F.; Krischer, K.; Ertl, G. Science 2001, 291 (5512), 2395-2398. (21) Hanken, D. G.; Jordan, C. E.; Frey, B. L.; Corn, R. M. Surface Plasmon Resonance Measurements of Ultrathin Organic Films at Electrode Surfaces. In Electroanalytical Chemistry; Bard, A. J., Rubinstein, I., Eds.; Marcel Dekker, Inc.: New York, 1998; Vol. 20, pp 141-225. (22) Lu, H. B.; Homola, J.; Campbell, C. T.; Nenninger, G. G.; Yee, S. S.; Ratner, B. D. Sens. Actuators, B 2001, 74 (1-3), 91-99.

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and collimated to have a parallel light beam that irradiates a large area of the metal film. The reflected light beam is generally recorded by a CCD camera. The principle of ellipsometric measurements is based on the change in the polarization of light reflected by the adsorbate film at a substrate.25-28 Upon reflection at the surface, the relative phase shifts and the intensity changes of the linearly polarized light in the s (direction perpendicular to the incidence) and p directions are different. By measuring phase shifts and intensity variations in the p and s directions with an analyzer, the refractive index and thickness of the adsorbate film can be deduced. A variant of ellipsometry, imaging ellipsometry (IE), also collects the light reflected off a surface with a CCD camera. SPRI and IE both offer quantitative thickness measurements and monitor events occurring across a large surface region at a relatively high spatial resolution. Although SPRI and IE are both optically based and share many common features in the experimental setup and theory (Fresnel reflection9), they have distinct and complementary features. SPRI requires the use of a thin metal film to excite the SP and monitors reflected light intensity, whereas IE is not limited to the surface type and detects polarization change (phase and relative intensity ratio of p- and s-polarized components). Since the resonance of SP enhances the evanescent field, SPRI is particularly sensitive to infinitesimal changes at or near the metal surface. Such changes in many cases are difficult to measure using ellipsometry, as the phase shift and intensity change in the reflected light are not perturbed significantly by ultrathin films (i.e., the lack of signal enhancement limits the sensitivity of ellipsometry). A large number of studies have separately utilized SPRI and IE to determine film thickness and to monitor thickness variations.27-31 Among the many different instrumental designs, positioning the optical components onto a goniometer is a popular approach. For example, Lyon et al. have constructed a SPRI instrument using a goniometer.32 Beaglehole conducted both phase-modulated ellipsometry and imaging ellipsometry with optical components mounted onto a motorized goniometer.33 A goniometer-supported instrument that can perform both SPRI and IE has been used in its IE mode to characterize a protein microarray on carboxymethylated dextran hydrogels.34 However, the range within which the optical arms are rotated on that setup is limited.34 As a result of the limited range of rotation, the possibility of or convenience in examining the same surface area (23) Fortin, E.; Defontaine, Y.; Mailley, P.; Livache, T.; Szunerits, S. Electroanalysis 2005, 17 (5-6), 495-503. (24) Shumaker-Parry, J. S.; Campbell, C. T. Anal. Chem. 2004, 76 (4), 907917. (25) Beaglehole, D. Proc. Phys. Soc., London 1966, 87 (2), 461-471. (26) Abeles, F. Surf. Sci. 1976, 56, 237-251. (27) Reiter, R.; Motschmann, H.; Orendi, H.; Nemetz, A.; Knoll, W. Langmuir 1992, 8 (7), 1784-1788. (28) Jin, G.; Jansson, R.; Arwin, H. Rev. Sci. Instrum. 1996, 67, 2930-2936. (29) Goodall, D. G.; Stevens, G. W.; Beaglehole, D.; Gee, M. L. Langmuir 1999, 15 (13), 4579-4583. (30) Striebel, C.; Brecht, A.; Gauglitz, G. Biosens. Bioelectron. 1994, 9 (2), 139146. (31) Howland, M. C.; Szmodis, A. W.; Sanii, B.; Parikh, A. N. Biophys. J. 2007, 92 (4), 1306-1317. (32) Lyon, L. A.; Holliway, W. D.; Natan, M. J. Rev. Sci. Instrum. 1999, 70 (4), 2076-2081. (33) Beaglehole, D. Rev. Sci. Instrum. 1988, 59 (12), 2557-2559. (34) Zhou, Y.; Andersson, O.; Lindberg, P.; Liedberg, B. Microchim. Acta 2004, 147 (1), 21-30.

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with SPRI and IE without serious optical realignments and sample readjustments is difficult to achieve. In this work, we report SPRI and IE thickness measurements accomplished by rotating a single optical system above and below the sample surface. Because no optical components and samples are removed or realigned, the dual imaging affords a reliable investigation of the surface morphology of the same sample and provides an opportunity to carry out thickness measurements in the same surface area with these two complementary techniques. MATERIALS AND METHODS Materials. Concentrated H2O2 was purchased from Sigma Chemical Co. (St. Louis, MO). NH4OH (29.4%) and CH3Cl (99+%) were obtained from Thermo-Fisher Scientific (Tustin, CA). 1,2Dipalmitoyl-sn-glycero-3-[phosphor-L-serine] (sodium salt) (DPPS) (99+% purity) was acquired from Avanti Polar Lipid, Inc. (Alabaster, AL). All other chemicals were used without further purification, and all solutions were prepared with deionized water treated with a Milli-Q Simplicity system (Millipore Corp., Bedford, MA). Procedures. Fabrication of Au Substrates and Ag Stripes. Gold substrates were prepared by sequentially evaporating 2 nm of chromium at a rate of 0.05 nm/s and 50 nm of gold at a rate of 0.1 nm/s onto BK7 microscope coverslips (Thermo-Fisher Scientific) inside an CHA electron-beam evaporation system (CHA Industries, Fremont, CA) at a base pressure smaller than 8 × 10-7 Torr. Ag stripes were prepared with a sputter coater (Cressington model 208HR, Ted Pella, Inc. Redding, CA) by placing a homemade mask over the Au substrates. The thickness of Ag stripes was monitored with a quartz crystal microbalance (QCM), which accurately measured the mass of the Ag deposited and converted the mass into thickness. To avoid oxidation of Ag stripes, which takes several days to develop a noticeable oxide layer in the cleanroom environment at room temperature with low humidity, all Au substrates covered with Ag stripes were fabricated right before the SRPI/IE measurements. Formation of Lipid Monolayer and Multilayers on Au Substrates. DPPS was suspended in a CH3Cl and CH3OH mixture (9:1) with a final concentration of 1 mg/mL. The solution was sonicated and stored in a freezer (-20 °C) prior to use. DPPS layers on Au substrates were formed using a Langmuir-Blodgett trough (Nima model 311D, Nima Technology Ltd., Coventry, England). Briefly, a metal clip held the substrate vertically and the substrate surface was partially immersed in the solution. DPPS solution was first introduced onto the water surface in the trough with a glass syringe (Hamilton Company, Reno, NV). After the DPPS molecules were evenly spread out on the water surface (∼30 min), the resultant monolayer was compressed to the desired pressure at a slow and constant barrier speed in the “pressure control” mode.35,36 After maintaining this surface pressure for another 30 min, the DPPS monolayer was transferred onto the Au substrate at a constant speed and a fixed surface pressure (40 mN/m2). The substrate immersed in solution became covered with DPPS when the substrate was pulled out. Through the “dip-and-pull” motion,35 compact multilayers of DPPS films were successively produced over the surface exposed to solution. Formation of the (35) Grunfeld, F. Rev. Sci. Instrum. 1993, 64 (2), 548-555. (36) Mirley, C. L.; Lewis, M. G.; Koberstein, J. T.; Lee, D. H. T. Langmuir 1994, 10 (7), 2370-2375.

dominant wavelength of 625 nm, a bandwidth of 30 nm (full width at half-maximum), and a typical luminosity of 40 candela. Since the LED is a point light source, a collimator (a lens with 75 mm focus length and 25 mm diameter) was positioned after the LED to produce a parallel light beam. Polaroid sheets served as the rotatable polarizers, and images were collected by a CCD camera (1024 × 1024 pixels, 12 bit per pixel resolution) equipped with a long-working-distance 12× objective. The magnification of the imaging optics can be continuously adjusted up to a maximum lateral resolution of 3 µm/pixel. In the IE mode (Figure 1a), the two optical arms are rotated to a preset angle, θIE, above the sample in the typical polarizersample-retarder-analyzer configuration.9 During the measurement, the retarder rotates to different angles and the CCD camera records the corresponding images. Traditionally, the ratio of the reflected light in the p and s planes can be expressed as follows:9

r ) rp/rs ) Re(r) + iIm(r) where rp and rs are the reflectivity in the p and s direction, respectively; Re(r) and iIm(r) are the real and imaginary parts of the reflected light. The real and imaginary parts are related to ψ and ∆ values which are measured by the conventional “nullingtype” ellipsometry:9 Figure 1. Schematic representations of (a) the IE mode wherein the various optical components on the two rotation arms are positioned above the sample of interest and (b) the SPRI mode wherein the arms are below the sample. The examined area of the sample is in the center of the goniometer.

DPPS stripes of different layer numbers were accomplished by raising the substrate clip up by a given distance (e.g., to produce a 2 mm wide DPPS stripe, after a given number of DPPS layers had been made, the initial height of the substrate would be raised by 2 mm). Repeating the above process generated different DPPS stripes with various thicknesses. SPRI and IE. The SPRI/IE setup used in this work was modified from an imaging ellipsometer (Beaglehole Instruments Ltd., Wellington, New Zealand). As schematically shown in Figure 1, the entire setup was mounted onto a motorized goniometer with incidence and reflection arms in the same vertical plane. Each arm can be rotated separately with a step motor at an angular resolution of 0.01°. The goniometer allows the rotation of each arm from 0° to 180°, facilitating the sample illumination and imaging both above and below the sample. An equilaterally triangular BK7 prism (refractive index ) 1.515, Melles Griot, Carlsbad, CA), confined in a sample holder, is placed in the center of the goniometer. The choice of a prism with a relatively low refractive index, instead of that with a higher refractive index, is based on the consideration that the SPRI images collected in air will be less distorted (vide infra). The Au substrate and/or other samples can be attached to the top of the prism using a refractive index matching liquid (Cargille Laboratorie Inc., Cedar Grove, NJ). The halogen lamp on the original ellipsometer was replaced with an intense semiconductor (InGaAlP) light-emitting diode (eLED Corp., Walnut, CA). The light-emitting diode (LED) was chosen as the source for its simplicity, low cost, brightness, and freedom from laser interference patterns.13 This LED has a

Re(r) + iIm(r) ) tan ψ exp(i∆) The IE mode in our setup, which is based on phase modulation, measures two parameters x and y that are different from but closely correlated to the ψ and ∆ values. The x and y parameters are correlated to Re(r) and Im(r) in the following two equations:

x ) 2Re(r)/(1 + Re(r)2 + Im(r)2) y ) 2Im(r)/(1 + Re(r)2 + Im(r)2) Since tan ψ and tan ∆ are functions of Re(r) and Im(r), as shown in the following equations,

tan ψ ) xRe(r)2 + Im(r)2 tan ∆ ) Im(r)/Re(r) the x and y parameters are essentially functions of ψ and ∆ commonly encountered in conventional ellipsometry. Thus, by rotating the retarder to angles corresponding to the different phase shifts (modulations) and measuring the reflected light with the analyzer set at +45° and -45° alternatively, the x and y values can be obtained and subsequently used to calculate the thickness and refractive index of the adsorbate film. The detailed working principle behind phase-modulated ellipsometry is described elsewhere.37 In the SPRI mode, the two optical arms are rotated below the sample and the light beam propagates through the triangular prism at the incidence angle θSPRI (Figure 1b). Notice that θSPRI is the difference between 180° and the arm angle. The polarizer, retarder, and analyzer are all set to 90° on their rotating mounts, (37) Acher, O.; Bigan, E.; Drevillon, B. Rev. Sci. Instrum. 1989, 60 (1), 65-77.

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Figure 2. (a) SPRI image of three Ag stripes (5, 10, and 15 nm thick, from left to right) sputtered onto a Au substrate and two IE images of the same surface area expressed in the x (b) and y (c) parameters. The cross-sectional contours along the dashed black lines in the three images are shown in (d). The SPRI image was obtained at θSPRI of 35.00°, whereas the IE images were recorded at θIE of 60.00°.

respectively, allowing only the p-polarized light to pass through. Fresnel calculations of the SPR resonance angles and film thickness were conducted using WINSPALL 3.0138 with different numbers of phases (layers).21 Simulations of the IE data were carried out with a program written in the IgorPro software. Rotations of the polarizer, retarder, and analyzer and movement of the optical arms are all controlled by the program developed with the IgorPro software. Both SPRI and IE were conducted in air (i.e., the sample on the Au substrate was exposed to ambient atmosphere). RESULTS AND DISCUSSION SPRI and IE Studies of Ag Stripes Deposited onto a Au Substrate. To establish the SPRI/IE for thickness measurements, we coated Ag stripes onto a gold substrate and sequentially imaged the resultant pattern with SPRI and IE. This pattern serves as a model system because the thickness values of the Ag stripes are all known (measured with QCM equipped on the sputter coater) and the refractive indices of Cr, Ag, and Au can be obtained from literature. Moreover, Ag/Au bilayers have been shown to enhance the SPR sensitivity due to the sharper SPR profile.39 Figure 2 displays the SPRI and IE images of a gold (38) WINSPALL (Analysis Package for Surface Plasmon Data). http://www.mpipmainz.mpg.de/∼johanns/ak_knoll_Software.htm. (Accessed June 8, 2007). (39) Zynio, S. A.; Samoylov, A. V.; Surovtseva, E. R.; Mirsky, V. M.; Shirshov, Y. M. Sensors 2002, 2, 62-70.

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substrate covered with silver stripes of three different thicknesses. A representative cross-sectional contour extracted from the SPRI image is shown as the blue curve in Figure 2d. In recording the SPRI image, θSPRI was set to 35.00° (i.e., the two arms of the goniometer were both positioned at 145.00°; see also Figure 1b). At such an angle, the incident light, upon penetrating through the equilaterally triangular prism, irradiates onto the back of the gold chip at an angle of 43.8° (SPR resonance angle) inside the prism. The rotating polarizer, retarder, and analyzer were all set to 90° on their rotating mounts; thus, light is polarized linearly and parallel to the incidence plane (p-polarized light). The blue line curve in Figure 2d, which was represented in the intensity ratio of reflected p- and s-polarized light (Rp/Rs) without considering phase change as in the rp/rs ratio of IE, depicts well-defined plateaus and edges of the Ag stripes. By subtracting the reflectivity at the bare gold region from those at the Ag stripes and comparing the results to the values predicted by the Fresnel calculation,21,38 the stripe thicknesses were deduced to be 5.3, 9.6, and 13.0 nm. These values are in good agreement with those measured by the QCM on the sputter coater (5.0, 10.0, and 15.0 nm). The IE images expressed in the x (Figure 2b) and y (Figure 2c) parameters were obtained by simply rotating the two arms to 60° (θIE) on the goniometer (cf., Figure 1a) and setting the polarizer and analyzer both at 45° with respect to the incidence plane and rotating the retarder. Contours of the same cross section in the two IE images (red curve for the x parameter and green

Figure 3. SPR image (a) of the boundary between a DPPS monolayer and the bare Au region, the cross-sectional contour (b) corresponding to the dashed line in the image, and the SPR profiles (c) recorded at the bare (thick line curve) and DPPS monolayer covered Au regions (thin line curve). The ellipsometric images of the same surface area, expressed in x and y parameters, are shown in panels (d) and (e), respectively. The corresponding cross-sectional contours extracted from the IE images are overlaid in panel (f). Note that the DPPS monolayer is located at the bottom part of the SPR image in (a) but appeared in the top of the images in (d) and (e).

curve for the y parameter) are overlaid in Figure 2d with that extracted from SPRI. By simulating the x and y parameters, the Ag stripe thicknesses were determined to be 5.0, 10.2, and 12.4 nm. These results are also consistent with both the values measured by SPRI and the QCM. Thus, both the SPRI and IE modes yielded accurate thickness values and can be used to examine the same area of a surface using a single optical system. Three additional points are worth noting upon making close comparisons between the SPRI and IE results in Figure 2. First, for ultrathin films, the SPR image produced a higher contrast (e.g., the boundaries of the Ag stripes in the SPR image are more discernible) and the contour (blue curve in Figure 2d) has a greater signal-to-noise ratio. The contrast is more obvious at the boundary between the 10 and 15 nm thick Ag stripes. The greater contrast can be attributed to the higher sensitivity inherent in SPRI, since the ultrathin Ag stripes are well within the field of the evanescent wave. In contrast, a small thickness variation generally does not alter the polarization of the light significantly, and consequently IE is less sensitive for ultrathin films. We should also note that the boundary between two stripes is typically greater than 100 µm, which far exceeds the lateral dimension measurable by most AFM scanners. Second, ellipsometric images of the greatest signal-to-noise ratio are generally obtained at an angle where the x and y parameters have the highest dependence on thickness9 (60° in the present case). Since this angle is much different than 45° at which collimated light travels for the same distance upon reflection, the resultant ellipsometric image is distorted. In the SPRI mode, the highest sensitivity is obtained at an angle close to the resonance angle (43.8°). Such a value is much closer to 45°. As a consequence, little distortion was observed. Third, notice that the orientation of the Ag stripes with respect to the dotted black line in Figure 2a is different than those in Figure 2, parts b and c, even though the same surface area

was examined. This difference is resulted from the fact that the same area was viewed with the optics below (in the SPRI mode) and above (in the IE mode) the sample. SPRI and IE Studies of Multilayered DPPS Films. We further extended SPRI/IE to the imaging and thickness measurements of Langmuir-Blodgett DPPS films. With the LangmuirBlodgett trough, the number of DPPS layers can be controllably varied and the structure of the multilayered films is compact and well organized. Figure 3a shows an SPRI image of a surface containing both bare and DPPS monolayer covered Au regions. The cross-sectional contour corresponding to the dashed white line (2 mm across) is presented in Figure 3b, whereas the SPR reflectivity curves at the bare and the DPPS-covered Au regions are shown in Figure 3c. In Figure 3c, a shift of SPR resonance angle by 0.28° was observed. By fitting the experimental results to the value predicted by the Fresnel calculation,21,38 the thickness of the DPPS monolayer was determined to be 2.6 ( 0.2 nm. The resonance angles of the SPR profiles shown in Figure 3c are in good agreement with the simulated SPR profile at the incidence angle and prism used in this study. However, the reflectivity minimum is greater than the theoretical value. This deviation is due to the broad bandwidth of the LED light source we used. Since a DPPS monolayer has a thickness of 2.5 nm and the goniometer has a resolution down to 0.01°, it is clear that our instrument is capable of measuring thicknesses of ultrathin films down to the subnanometer resolution. Juxtaposed in the second row of Figure 3 are the IE images of the same surface area as in Figure 3a that are expressed in the x (Figure 3d) and y (Figure 3e) parameters and the contours (Figure 3f) of the same cross section as that in Figure 3a. Notice that the orientation of the boundary in Figure 3, parts d and e, is again different than that in Figure 3a, due to the aforementioned fact that the SPRI and IE images were collected below and above the sample. Because the Analytical Chemistry, Vol. 80, No. 3, February 1, 2008

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Figure 4. (a) Angle-resolved SPRI measurements of three DPPS multilayers formed at a Au substrate, (b) simulated SPR curves, (c) profiles of IE images of the same three DPPS multilayers, and (d) simulated ellipsometric parameters for determining DPPS multilayers between 0 and 120 nm.

DPPS monolayer is rather thin, the SPRI image is of a greater contrast than the IE images. As a result, the DPPS layer thickness deduced from the SPRI image (2.6 ( 0.2 nm) is in a much better agreement with the theoretical DPPS thickness (2.7 nm)40 than that from the IE measurement (2.0 ( 0.2 nm). The measurement of a relatively thick film by SPRI, on the other hand, can be problematic when the thickness has exceeded certain values. Figure 4a exemplifies such a scenario. When the DPPS layer number was increased from 3 to 15 (or thickness from 7.4 to 46 nm), the SPR resonance angle shifted from 36.50° to 52.60°, and the SPR reflectivity curve became broader. These characteristics are consistent with those predicted from the Fresnel calculation (cf., the red and green curves in Figure 4b simulated using a refractive index of 1.45 and thicknesses of 7.4 and 46 nm, respectively). As the DPPS layer number was further increased to 31 (corresponding to a thickness of 94 nm), the SPR resonance angle shifted to 82.42° (cf., the blue curve in Figure 4b). Such a high resonance angle prohibits a complete SPR reflectivity curve from being recorded on our instrument (see the blue curve in Figure 4a). As abovementioned, the field strength of the evanescent wave decays exponentially with the distance from the Au substrate. With an incident light of 625 nm, the thickness range within which SPR remains sensitive is about 1/e of the wavelength or approximately 160 nm.41 At 94 nm the SPRI sensitivity has already much attenuated, and an infinitesimal thickness variation has become more difficult to measure. The above SPRI limitation can be readily overcome by IE, whose dynamic range is much wider. Figure 4c depicts the crosssectional contours of three stripes of DPPS films composed of 3, 15, and 31 layers. The distinct x and y values at the terraces of (40) Fanucci, G. E.; Backov, R.; Fu, R.; Talham, D. R. Langmuir 2001, 17 (5), 1660-1665. (41) Ordal, M. A.; Bell, R. J.; Alexander, R. W., Jr.; Long, L. L.; Querry, M. R. Appl. Opt. 1985, 24, 4493-4499.

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the three DPPS stripes demonstrate that IE can easily distinguish the layers of large thickness differences. Notice that both the experimental and theoretical x and y values do not linearly increase with the layer number (thickness). This can be explained on the basis that IE detects the phase shift associated with thickness change and the ellipticity exhibits periodic variation over a wide range of thickness values.9 To illustrate this point, we plotted the theoretical x and y parameters in Figure 4d as a function of film thickness between 0 and 120 nm. Notice that the x and y parameters vary differently with the film thickness. In the region where one parameter (e.g., x) is relatively invariant, the other parameter (e.g., y) changes substantially. Thus, comparisons between the x and y parameters enable one to continuously measure the film thickness variation over a wide range. This is in contrast to the SPRI measurements, which are subject to the coverage of the evanescent wave associated with the surface plasmon. CONCLUSIONS SPRI and IE constructed with a single optical system mounted on a goniometer have been developed. The SPRI and IE imaging and thickness measurements can be sequentially performed on the same area of a sample. Switching between the two techniques or modes can be conveniently accomplished by simply rotating the two arms that hold the light source, polarizer, retarder, analyzer, and detector above (in the IE mode) and below (in the SPRI mode) the sample. In the SPRI mode, the retarder is fixed at 90° normal to the incidence plane, whereas the retarder is rotated at a predetermined rate in the IE mode. Through the comparative studies of ultrathin Ag stripes sputtered onto a Au substrate and phospholipid films comprising various layer numbers, the two techniques were shown to be highly complementary to each other. Whereas SPRI is extremely sensitive to thickness variation of films of a few nanometers, IE is more suitable to the

measurement of thicker films. Thus, the capability of this setup in determining both thin and thick films provides an approach to investigate cases where continuous monitoring of film growth and imaging of samples with large surface undulations are required. ACKNOWLEDGMENT Partial support by a NSF-RUI Grant (0555244), the NIH-RIMI Program at California State UniversitysLos Angeles (P20

MD001824-01), and a Dreyfus Teacher-Scholar Award (TH-01025) is gratefully acknowledged.

Received for review September 5, 2007. Accepted October 26, 2007. AC701873V

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