Anal. Chem. 1996, 68, 473-480
Quantitative Evaluation of SERS-Active Ag Film Nanostructure by Atomic Force Microscopy Shane E. Roark, David J. Semin,† and Kathy L. Rowlen*
Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309
The reliability of an image analysis algorithm for atomic force microscopy (AFM) of thin metal films was evaluated by comparison with manual analysis of images and transmission electron micrographs of Ag films deposited on Formvar-coated Cu grids. In order to extract quantitative nanostructural information using the algorithm discussed herein, the optimal fitting parameters were found to be low-pass filtering to reject high-frequency noise, a 5 × 5 point grid for identification of particle maxima, and a linear least-squares fit to a hemispheroidal model of particle shape. Metal particle dimensions were defined from the height and radius of the hemispheroid fit. Due to the close spacing of particles in these Ag films, tip geometry causes the greatest error in the height measurements, rather than width measurements. In addition, the effect of scanning parameters such as scan rate and size, applied load, and humidity on particle count and dimensions was examined. Increasing the scan rate reduced the number of resolvable Ag particles, decreased the apparent particle height, and increased the apparent particle radius. Under conditions of low capillary force, a net repulsive force of ∼19 nN resulted in subtle tipinduced changes in the Ag surface morphology. The Ag film surface was damaged at a net repulsive force of ∼23 nN. At slow scan rates, the moisture layer did not significantly affect the quality of the AFM images obtained over a broad relative humidity range. Finally, the Ag surface structure was found to be very homogeneous over a relatively large area. Traditionally, metal film nanostructure has been characterized with transmission electron microscopy (TEM) or scanning electron microscopy (SEM). The qualitative information available from these techniques has provided invaluable insight into surface morphology over the years.1-11 In order to extract quantitative † Los Alamos National Laboratory, Los Alamos, NM 87545. (1) Cohen, R. W.; Cody, G. D.; Coutts, M. D.; Abeles, B. Phys. Rev. B 1973, 8, 3689-3701. (2) Davis, C. A.; McKenzie, D. R.; McPhedran, R. C. Opt. Commun. 1991, 85, 70-82. (3) Buchholz, S.; Fuchs, H.; Rabe, J. P. J. Vac. Soc. Am. B 1991, 9, 857-861. (4) Dawson, P.; Alexander, K. B.; Thompson, J. R.; Haas, J. W., III; Ferrell, T. L. Phys. Rev. B 1991, 44, 6372-6381. (5) Hurt, H. H.; Bennett, J. M. Appl. Opt. 1985, 24, 2712-2720. (6) Gajdardziska-Josifovska, M.; McPhedran, R. C.; Cockayne, D. J. H.; McKenzie, D. R.; Collins, R. E. Appl. Opt. 1989, 28, 2736-2743. (7) Schlegel, V. L.; Cotton, T. M. Anal. Chem. 1991, 63, 241-247. (8) Sennett, R. S.; Scott, G. D. J. Opt. Soc. Am. 1950, 40, 203-211. (9) Sharma, S. K.; Spitz, J. Thin Solid Films 1980, 65, 339-350. (10) Varnier, F.; Mayani, N.; Rasigni, G.; Rasigni, M.; Llebaria, A. J. Vac. Soc. Am. A 1987, 5, 1806-1808. (11) Yamamoto, M.; Namioka, T. Appl. Opt. 1992, 31, 1612-1621.
0003-2700/96/0368-0473$12.00/0
© 1996 American Chemical Society
information about metal particle dimensions using standard TEM and SEM, particle width must be measured manually on the electron micrograph and scaled to account for the magnification factor. Using the measured mass thickness of the film, an assumed metal particle geometry, and the calculated percent metal volume on the surface, an average particle height can be estimated.12,13 For obvious reasons, electron microscopic techniques are inappropriate for investigation of metal films on dielectric surfaces and metal films in air. Our research group continues to be interested in the morphology and annealing characteristics of thin metal films on dielectric surfaces under ambient conditions. Understanding the relationship between thin metal film morphology and optical properties is particularly important for the efficient use of these films in surface-enhanced Raman scattering (SERS). Atomic force microscopy (AFM) is the ideal choice for the surfaces mentioned above.14-18 The sample need not be conductive, imaging can be conducted in air, and three-dimensional information is obtained directly. Scanning tunneling microscopy (STM) has also been employed to characterize the surface structure of metal films.3,4,19-21 However, STM also requires that the sample be electrically conductive. For this reason, STM studies of metal films have been mainly limited to films greater than 40 nm in thickness.3,4,20,21 Mo¨ller et al. used STM to study Ag films only 4 nm thick; however, in order to make the sample conductive, a glass substrate was first coated with a layer of indium tin oxide which exhibited a roughness scale on the order of the metal particle dimensions.19 In order to extract reliable quantitative measurements from AFM data, it is necessary to fully understand the effects of experimental and scanning parameters on image quality. In this work, the measured dimensions of Ag particles as a function of probe tip geometry, scan rate and size, humidity, applied load, and postimage filtering are discussed in terms of both a manual evaluation of AFM images and an automated image analysis algorithm. (12) Shawki, G. S. A.; El-Sherbiny, M. G.; Salem, F. B. Thin Solid Films 1981, 75, 29-36. (13) Kovacs, G. J.; Loutfy, R. O.; Vincett, P. S.; Jennings, C.; Aroca, R. Langmuir 1986, 2, 689-694. (14) Roark, S. E.; Rowlen, K. L. Chem. Phys. Lett. 1993, 212, 50-56. (15) Roark, S. E.; Rowlen, K. L. Anal. Chem. 1994, 66, 261-270. (16) Van Duyne, R. P.; Hulteen, J. C.; Treichel, D. A. J. Chem. Phys. 1993, 99, 2101-2115. (17) Semin, D. J.; Rowlen, K. L. Anal. Chem. 1994, 66, 4324-4331. (18) Schimmel, T.; Bingler, H.; Franzke, D.; Wokaun, A. Adv. Mater. 1994, 6, 303-307. (19) Mo ¨ller, R.; Baur, C.; Graff, U.; Ku ¨ rz, P.; Leitner, A.; Pedarnig, J. D.; Aussenegg, F. R. J. Phys. D 1990, 23, 1267-1270. (20) Baski, A. A.; Fuchs, H. Surf. Sci. 1994, 313, 275-288. (21) Gimzewski, J. K.; Humbert, A.; Bednorz, J. G.; Reihl, B. Phys. Rev. Lett. 1985, 55, 951-954.
Analytical Chemistry, Vol. 68, No. 3, February 1, 1996 473
EXPERIMENTAL SECTION Ag Particle Film Preparation. Precleaned glass microscope slides were sequentially sonicated for several minutes in hexanes, chloroform, acetone, and methanol (Mallinkrodt) followed by heating at 120 °C for 1 h. Thin Ag films were deposited with a Denton Vacuum DV-502 vapor-phase depositor. The pressure inside the deposition chamber was maintained at ∼5 × 10-6 Torr. The film thickness and deposition rate were monitored with a factory-calibrated, temperature-controlled, quartz crystal microbalance (Syncon Instruments STM-100). The microbalance crystal was placed adjacent to and in the same plane as the substrate. The substrate was positioned 22 cm above and parallel to the source. The source was positioned equally between the substrate and microbalance at an angle of 4° from each.17 The deposition rate was 0.02 ( 0.005 nm/s. AFM. AFM images were obtained in air on either a Nanoscope II, III, or E scanning force microscope (Digital Instruments). The probe tips used were of two common types: etched singlecrystal Si or Si3N4 (Digital Instruments). On average, the Si tips have a height of 10-15 µm, an internal half-cone angle of 17°, and a tip radius of curvature between 5 and 20 nm. Typical spring constants are between 0.01 and 0.3 N/m, and typical resonance frequencies are between 5 and 20 kHz. Almost all of the AFM data herein were obtained using the single-crystal Si tips. The Si3N4 tips are pyramidal in shape, 3-5 µm nominal tip height with an internal tip angle of 70° and a radius of tip curvature between 20 and 50 nm. The spring constant is between 0.01 and 0.6 N/m, and the resonance frequency is ∼20 kHz. A Model AS-8 tubetype piezoelectric scanner was used. The maximum scan range in the x- and y-directions was 12.0 µm, and the maximum scan range in the z-direction was 4.4 µm. Resolution in the z-axis was 0.01 nm. Unless otherwise stated, images were acquired in the contact mode with net repulsive forces between 5 and 15 nN. Scan rates used for imaging were below 10 Hz. For humidity measurements, the AFM was operated in a custom-built Lucite enclosure. The humidity level in the enclosure was varied by flow rate control of a mixture of dry and humidified nitrogen through an inlet port. Humidity was measured with a slow-response hygrometer (Brannan), and the measurements are considered approximate. The images were low-pass filtered using the Digital Instruments software, a weighted 3 × 3 average. Quantification of AFM Images by Manual Analysis. Manual quantification was accomplished by first selecting three small subareas within a given AFM image. For a given 1 µm × 1 µm image, three 0.25 µm × 0.25 µm areas were selected and used for quantification. Random two-dimensional line profiles were generated in each of these subareas, and all Ag particles that were bisected by the line were measured. Due to the finite size and shape of the AFM tip, it is often difficult to find the “base” of a particle. For this reason, Ag particle heights were measured relative to the average of the 10 lowest spots in the subareas. Particle radius was measured as the horizontal distance between the particle maximum on the line profile and its edge on the twodimensional image. Unless otherwise stated, a total of 30 particles were quantified for each image. TEM. Micrographs of Ag films were acquired on either a JEOL 100 CX or Phillips TEM operated at either 100 or 205 kV. Prior to TEM analysis, the Ag films were deposited onto Formvarcoated 200 mesh copper grids (Ted Pella). The magnification 474
Analytical Chemistry, Vol. 68, No. 3, February 1, 1996
factor was determined from the 8.75 nm lattice spacing on a beef liver catalase crystal (SPI Supplies). RESULTS AND DISCUSSION Quantification of AFM Images by Computer Analysis. While manual quantification of metal particle nanostructure by cross-section analysis is the most direct measure of particle dimensions, it is also tedious and time consuming, and only a fraction of the Ag particles in an image can be measured. In a previous paper, a numerical algorithm was developed so that every Ag particle in an image could be quantified automatically.22 In this paper, the use of the algorithm for quantification of particle height, width, and number is examined in detail. Each AFM image consists of a 512 × 512 lateral grid of values that correspond to vertical measurements. For a 1 µm × 1 µm scan, the image has an in-plane x-y resolution of ∼2 nm. Prior to software analysis images were trimmed (∼10% from each edge) in order to avoid edge effects. The surface function z(x,y) was modeled as either a superposition of full ellipsoids or hemispheroids.22 Each image was first scanned for all local maxima by sequentially examining both a 3 × 3 and a 5 × 5 grid of points. Once the location of all the particles was determined, the points that defined the cap of the particle were fit to either an in-plane ellipsoid or a hemispheroid using a linear least-squares method (IMSL routine DLCLSQ). Since an average particle is ∼20 nm in diameter (∼10 data points in length), a 3 × 3 grid of points at the top of the particle was considered a reasonable number to use for the least-squares fit. However, in order to obtain a better estimate of relative error in the fit, a grid of 5 × 5 data points was also examined. The norm of the residual vector indicated that the tops were well represented by an ellipsoid (estimated between 3 and 8% relative error for the 5 × 5 grid); however, the remainder of the particle can deviate from ellipsoidal shape. As with manual measurements, particle height was defined as the vertical distance between a particle maximum and the lowest points on the entire image. Interpolation between the points on the ellipsoid or hemispheroid surface was used to identify the actual particle maximum, and the average of the 10 lowest points found on the image was used as the baseline reference. The low points were averaged to counter any unevenness present on the substrate. Particle radius or width was determined from the average of the in-plane major and minor radii of the ellipsoid or hemispheroid. In order to eliminate artifactual contributions from “scan lines” (i.e., high-frequency noise) in the images, radius measurements greater than 30 nm were rejected by the algorithm. The algorithm was evaluated by comparing measured particle dimensions to those determined by manual quantification of the images shown in Figure 1. The images in (A) and (B) are of the same 5 nm Ag film on glass scanned at different resolutions. The area in image B was from the center of the area shown in (A). The results from manual quantification and the algorithm were also compared for AFM images and TEM micrographs obtained on a separate Ag film, the TEM micrograph serving as a reference independent of tip geometric effects and scan noise. A representative AFM image C and TEM micrograph D of a 5 nm thick Ag film are also shown in Figure 1. In order to make a quantitative comparison of the AFM and TEM images, the film shown in panels C and D was deposited onto a Formvar-coated TEM grid.23 (22) Roark, S. E.; Semin, D. J.; Lo, A.; Skodje, R. T.; Rowlen, K. L. Anal. Chim. Acta 1995, 307, 341-353.
Figure 1. AFM image of (A) a 1 µm × 1 µm scan of a 5 nm thick Ag film on glass and (B) a 0.5 µm × 0.5 µm scan obtained from the center of the area shown in (A). AFM image C and TEM micrograph D are of the same 5 nm thick Ag film deposited onto a Formvar-coated TEM grid. The area shown in (C) is 0.25 µm × 0.25 µm, whereas the area shown in (D) is approximately 0.25 µm × 0.25 µm. Reduced to 90% of original for publication. Table 1. Comparison of Ag Particle Count and Heighta 1 µm × 1 µm
0.5 µm × 0.5 µm
0.3 µm × 0.3 µm
0.5 µm × 0.5 µm
0.5 µm × 0.5 µm
quantificn method
particle count
height (nm)
particle count
height (nm)
particle count
height (nm)
particle count
height (nm)
particle count
height (nm)
manual raw 3 × 3 raw 5 × 5 low-pass 3 × 3 low-pass 5 × 5
1603 1939 1827 1679 1606
6(1 7(1 7(1 6(1 6(1
572 878 829 587 581
6(1 7(1 7(1 7(1 7(1
365 1797 1419 495 481
6(1 8(1 8(1 8(1 8(1
670 1568 1374 696 673
6(1 9(1 9(1 9(1 9(1
647 1474 1324 660 645
7(1 8(1 8(1 9(1 9(1
a The manual quantification method was performed on the raw, unfiltered images. The raw 3 × 3 and raw 5 × 5 methods correspond to an algorithm fit to a 3 × 3 and 5 × 5 data point grid in the unfiltered images, respectively. The low-pass 3 × 3 and low-pass 5 × 5 methods correspond to algorithm fits to the same images after low-pass filtering. The data in columns 2-7 were generated from the images shown in Figure 1A, B, and C, respectively. The data shown in columns 8-11 were generated from images obtained from the same sample shown in Figure 1C. The error in the height measurement is (1σ, and is representative of the distribution in particle height.
The first test of the algorithm was to ensure correct identification of the number of Ag particles in a given image. Every particle (island) in the images shown in Figure 1, as well as two additional images of the same sample shown (A, B), was manually counted. The results for the manual vs algorithm Ag particle count, as well as particle height, are shown in Table 1. The most notable result in Table 1 is the excellent agreement in particle count between manual measurements and the 5 × 5, filtered version of the (23) Roark, S. E.; Rowlen, K. L. Appl. Spectrosc. 1992, 46, 1759-1761.
algorithm. High-frequency noise causes spikes in the images which are interpreted by the algorithm as peak maxima. The result is a large overestimation of number of Ag particles. The effect of low-pass filtering on the particle count by the algorithm is shown dramatically in Figure 2. Figure 2A is an unfiltered image of a 5 nm Ag film on glass with “tick” marks at all positions where the algorithm identified a particle. Figure 2B shows the result for the same image after low-pass filtering. It is clear that the algorithm grossly overestimated the particle count for the Analytical Chemistry, Vol. 68, No. 3, February 1, 1996
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Figure 2. The 126 nm × 126 nm areas magnified from a 1 µm × 1 µm scan. The image in (A) is unfiltered. The image in (B) has been filtered to remove high-frequency noise. Tick marks indicate points where the algorithm has identified a Ag particle. Reduced to 57% of original for publication.
unfiltered image. On average, the raw 3 × 3 fit overestimated the particle count by well over 100%. It is reasonable that the greater the degree of high-frequency noise in an image the more inaccurate the particle count. As a control comparison, particle count from AFM images and a manual count of the particles in the TEM micrograph shown in Figure 1D was conducted. The TEM results indicate 145 ( 6 particles in a 250 ( 10 nm × 250 ( 10 nm area. This value is only slightly lower than the results obtained for the corresponding AFM image (Figure 1C) processed by the 5 × 5, low-pass version of the algorithm (177 ( 11 particles in a 250 nm × 250 nm area). Within the error associated with the TEM image area and the variability on any given film, these particle counts are in good agreement. The particle count obtained by automated analysis of AFM images is therefore deemed a reliable measurement. The height measurements did not significantly depend on the fitting routine or whether the images were filtered. However, in each case the algorithm indicated larger particle heights than the manual measurements. Since the algorithm is capable of identifying the lowest points on the entire image, the height measurements from the algorithm are likely to be more accurate. In order to determine whether or not the Ag particles are best approximated by an ellipsoid or hemispheroid, radius measurements obtained manually and from the algorithm were compared. On average, fitting particle shape to a hemispheroid produced a 55% greater radius measurement than fitting to an ellipsoid. Manually determined radius values generally fell between the values determined by the ellipsoid and hemispheroid fits. For example, the average radius for the image shown in Figure 1C was 8 ( 2, 5 ( 2, and 9 ( 3 for manual, ellipsoid, and hemispheroid fits, respectively. In order to get a “true” radius measurement, one independent of tip geometric effects, the average particle radius was also determined manually from the TEM micrograph shown in Figure 1D. The average radius of 439 particles on the TEM micrograph was determined to be 8 ( 3 nm. For comparison, manual radius measurements of 90 particles from three AFM images taken on the same sample (one of those images is shown in Figure 1C) also produced an average radius of 8 ( 3 nm. The manual radius measurements from TEM and AFM also produced equivalent size distributions. Regardless of the actual shape of the Ag particles, due to tip geometric effects, particle shape in the AFM images is hemispheroidal. For this reason, in addition to the fact that the manual measurements are much closer to the values obtained with hemispheroid fit to 476 Analytical Chemistry, Vol. 68, No. 3, February 1, 1996
Figure 3. (A) A 0.5 µm × 0.5 µm scan of a 5 nm Ag film obtained with an etched single-crystal Si tip and (B) the same film with a Si3N4 tip. The plots below the images are the two-dimensional line cross sections. Reduced to 58% of original for publication.
particle shape, the hemispheroid fit was used for all quantitative AFM image measurements of Ag particle films. Tip Geometric Effects. For Ag particle films it is important to realize that the individual Ag particles are oblate. Tip geometry can have a greater influence when imaging tall, steep surface features because contact between the tip and surface feature is first made well above the apex of the tip. Thus, the slope of the tip affects the observed image. However, since Ag particles have a much lower aspect ratio than the tips used in this work, and a radius of curvature greater than that of the tips, most of the contact between the tip and particles remains close to the apex of the tip. Therefore, for a single, isolated Ag particle one would expect a very accurate height measurement and a width measurement only slightly larger than the actual width. However, in a thin Ag film, the particles are close together (∼1-2 nm base to base) relative to the size of the tip, thus limiting tip access to the regions between particles. Limited tip access yields artificially low height values. For example, a cross-section analysis of Ag films of increasing thickness, 2.5-7 nm, indicates that the apparent Ag particle heights increase steadily. However, the apparent Ag particle height decreases by almost 50% from 7 to 9 nm in thickness, probably due to a decrease in particle separation. A further demonstration of tip geometry on height measurements is shown in Figure 3. Figure 3A shows a typical AFM image and cross section of a 5 nm Ag film on glass obtained with an etched Si tip. Recall that Si tips are relatively long and narrow with a tip radius of curvature of only ∼10 nm. For comparison, the image in Figure 3B is of the exact same sample imaged with a Si3N4 tip. The Si3N4 tips are shorter and have a much greater internal cone angle and a tip radius of curvature of ∼20-50 nm. Qualitatively there is a clear distinction between the two images. Quantitatively, the image in Figure 3A yielded an average particle height of 8 ( 1 nm and particle radius of 11 ( 2 nm. The image in Figure 3B yielded an average particle height of 2 ( 1 nm and particle radius of 10 ( 2 nm. Furthermore, the algorithm identified over 100 more particles in (A) than in (B). Comparison of the AFM image and the TEM micrograph in panels C and D of Figure 1 directly addresses the issue of tip geometry on particle radius measurements. Qualitatively, the particles in the TEM micrograph appear more well defined and there appears to be greater interparticle spacing. Additionally,
very small particles in the spaces between the larger particles are visible in the TEM micrograph. However, as pointed out above, manual measurements of particle radius on both the AFM images and TEM micrographs produce the same result (8 nm). It is likely that since the interparticle spacing is only ∼2 nm, particle radius measurements are less subject to error than height measurements. For example, two particles that are 2 nm apart can appear to join at the base. If radius measurements are taken from the particle maxima to the minima between them, the radius will only be overestimated by ∼1 nm. Using the image analysis algorithm, an attempt is made to partially overcome the possible inaccuracy in height measurements by identifying the lowest points on the micrograph as baseline reference points. Most images of Ag-particle films contain some regions where there is considerable space between particles. If the tip is able to make contact with the substrate in these areas, and those points are used as the base value, very accurate height information should be obtained. The uncertainty in this technique lies in the fact that not all images have such low points, and the assumption is made that the low points represent the underlying substrate. As a check of the reliability of Ag particle dimensions determined from the algorithm, the mass thickness (film thickness) of Ag was calculated and compared to the mass thickness indicated by the quartz crystal microbalance (5.0 ( 0.1 nm). Use of the average particle count and measured height and radius values to calculate a volume per particle for five films (∼5000 particles) indicated a Ag mass thickness of 5 ( 1 nm. Although this comparison is crude, it is important that the mass thickness calculated from the particle dimensions agreed with the thickness measured by the microbalance. Mathematical methods to extract true feature size and shape from an AFM image assume accurate height measurements and require detailed knowledge of the tip shape.24-29 However, for these films, the image analysis algorithm appears to provide fairly accurate measurements. Effect of Scan Rate and Scan Size. When imaging in the constant-force mode, the practical upper limit of the scan rate is limited by how fast the piezoelectric scanner can react to deflection signals in the feedback loop from the split photodiode.30,31 The resonance frequency of the cantilever can also limit the physically achievable scan rate; however, cantilever resonance frequencies are typically much higher than the resonance frequencies of the scanners. For Ag particle films, the effect of scan rate on image quality is shown in Figure 4 for a 5 nm Ag film on glass. Qualitatively, as the scan rate was increased from 1.00 to 30.5 Hz, a dramatic change in the contrast and resolution of the images was observed, along with a distortion in the surface features. Quantitatively, panels A and B of Figure 5 demonstrate that the number of Ag particles detected by the algorithm decreased rapidly as the scan rate was increased, for both 1 µm × 1 µm and (24) Keller, D. Surf. Sci. 1991, 253, 353-364. (25) Burnham, N. A.; Colton, R. J.; Pollock, H. M. J. Vac. Sci. Technol. A 1991, 9, 2548-2556. (26) Markiewicz, P.; Goh, M. C. Langmuir 1994, 10, 5-7. (27) Eppell, S. J.; Zypman, F. R.; Marchant, R. E. Langmuir 1993, 9, 22812288. (28) Vesenka, J.; Manne, S.; Giberson, R.; Marsh, T.; Henderson, E. Biophys. J. 1993, 65, 992-997. (29) Artifacts in SPM, TopoMetrix, Santa Clara, CA, 1993. (30) Jahanmir, J.; Haggar, B. G.; Hayes, J. B. Scanning Microsc. 1992, 6, 625660. (31) Butt, H. J.; Siedle, P.; Seifert, K.; Fendler, K.; Seeger, T.; Bamberg, E.; Weisenhorn, A. L.; Goldie, K.; Engel, A. J. Microsc. 1993, 169, 75-84.
Figure 4. Images of 0.5 µm × 0.5 µm scans of a 5 nm thick Ag film on glass at scan rates of (A) 1, (B) 12, and (C) 31 Hz. The exact same area is imaged at each scan rate. Reduced to 80% of original for publication.
0.5 µm × 0.5 µm scan sizes. The area imaged in the 0.5 µm × 0.5 µm scan corresponds to the center of the area imaged in the 1 µm × 1 µm scan. Upon increasing the scan rate from 1 to 30 Hz, the number of Ag particles decreased by ∼7 and 15% for the 1 µm × 1 µm and the 0.5 µm × 0.5 µm scan sizes, respectively. Figure 5C shows that the average Ag particle height also decreased as the scan rate was increased for both scan sizes. The lowest curve in Figure 5C is the measured height (manual analysis) for a single particle on a 0.5 µm × 0.5 µm image. Figure 5D shows that the Ag particle radius increased as a function of scan rate; however, the change in Ag particle radius was less dramatic than for height. Although the Ag particles are oblate and do not have steep edges, it is likely that the distortion of the surface features at high scan rates is the result of the z-axis response of the scanner. At high lateral scan rates, the z-response Analytical Chemistry, Vol. 68, No. 3, February 1, 1996
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Figure 5. Number of Ag particles identified by the algorithm as a function of scan rate from (A) a 1 µm × 1 µm and (B) a 0.5 µm × 0.5 µm scan area. The plots in (C) and (D) show the effect of scan rate on the apparent Ag particle height and radius. The filled circles are from 1 µm × 1 µm scans, and the unfilled circles are from 0.5 µm × 0.5 µm scans. The boxes are for a single Ag particle manually measured from the 0.5 µm × 0.5 µm scans. For a 1 µm × 1 µm scan, the image has an in-plane x-y resolution of ∼2 nm.
to the photosignal feedback is not fast enough to maintain tracking the ∼6 nm vertical range. It is reasonable that the taller the surface features are, the more severe the scan rate dependence. In fact, high scan rates are generally only useful for relatively hard, atomically flat surfaces.32,33 Additionally, at high scan rates the cantilever can experience buckling due to increased lateral forces from the surface moisture layer.33 Buckling of the cantilever can result in low height measurements and increased radius measurements due to both a change in deflection from the cantilever and a new point of contact between the tip and the sample. The effect of scan size on quantitative measurements of Agparticle films can be seen in Table 1, as well as Figure 5. In Table 1, the data in the columns 2-5 can be compared because they are from images of the same area of the same sample. The particle count for the image corresponding to the 1 µm × 1 µm scan was less than 3 times greater than the particle count for the 0.5 µm × 0.5 µm scan, although the scan area was 4 times greater. Also, the average Ag particle height was lower for the 1 µm × 1 µm scan. For the same samples, the average radius was greater in the 1 µm × 1 µm scan (e.g., 9 ( 3) than for the 0.5 µm × 0.5 µm scan (e.g., 7 ( 3). Since the 0.5 µm × 0.5 µm scan was acquired from the center of the 1 µm × 1 µm scan, one would expect the average particle dimensions to be the same. The same trend is observed in Figure 5. Although both scan sizes were performed at the same line per second rate, the tip must travel (32) Alexander, S.; Hellemans, L.; Marti, O.; Schneir, J.; Elings, V.; Hansma, P. K.; Longmire, M.; Gurley, J. J. Appl. Phys. 1989, 65, 164-167. (33) Ohnesorge, F.; Binnig, G. Science 1993, 260, 1451-1456.
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twice as fast to image a 1 µm × 1 µm area than a 0.5 µm × 0.5 µm area. Since for each “line” the sample is moved left to right and then right to left, for a scan rate of 5 Hz (lines/s) in a 1 µm × 1 µm area, the sample is being moved at a velocity of 10 µm/s. At 5 Hz in a 0.5 µm × 0.5 µm area, the sample is being moved at a velocity of 5 µm/s. Since the trend in the scan rate data shows that at higher scan rates there appear to be fewer particles and the particles are shorter and wider, it is logical that at a given scan rate an image of a larger scan area will appear to have fewer particles and the particles will be shorter and wider than an image of a smaller area. However, this explanation does not completely account for the discrepancy in quantitative data between the 1 µm × 1 µm and 0.5 µm × 0.5 µm scan sizes. For example, the data from the 1 µm × 1 µm area scanned at 5.1 Hz can be compared to the data for the 0.5 µm × 0.5 µm area scanned at 10.2 Hz since, for both scans, the sample is traveling at 10.2 µm/ s. In this comparison, both data sets yielded the same height (6.3 ( 1 nm). However, the radius measurement from the 1 µm × 1 µm scan was ∼2 nm greater than for the 0.5 µm × 0.5 µm scan. Furthermore, there were only 3 times as many particles in the 1 µm × 1 µm scan than the 0.5 µm × 0.5 µm scan. It is likely that, for small scan areas, irregularities in the particle surfaces that are not resolved in images of larger areas are being identified by the algorithm. This is reasonable since the 5 × 5 grid of data points that are fit to the particles corresponds to only ∼5 nm × 5 nm in the 0.5 µm × 0.5 µm scans, whereas for the 1 µm × 1 µm scan the 5 × 5 grid corresponds to ∼10 nm × 10 nm. Therefore, for smaller scan areas, the 5 × 5 grid covers a smaller fraction of the particle. This effect would be even more severe for low scan rates where particle fine structure is more apparent and might explain why the particle count was exceptionally high (695) for the 1 Hz scan rate of the 0.5 µm × 0.5 µm area in Figure 5B. Increasing the scan rate to 3.1 Hz reduced the resolution of irregularities in the particle surface considerably and dropped the particle count by ∼15%. Effect of Force. AFM images of a Ag film were acquired under a range of applied loads to determine whether or not the force between the tip and sample affects image quality. Also, the force required to promote tip-induced perturbations of the Ag surface was determined. The ambient relative humidity was typically ∼25%, which generated a capillary force between the tip and Ag film of ∼5 nN (using a spring constant of 0.2 N/m). As long as the tip and sample are in contact, the capillary force contributes to the net force acting between them. From this base capillary load, additional loads were applied in ∼3 nN increments. Figure 6A shows a 1 µm × 1 µm area image of a Ag film on glass obtained at 4.8 nNsunder capillary forces only (no cantilever deflection). Figure 6B shows an image of the same area under a net load of ∼23 nN. At 23 nN of net repulsive force, the tip began to seriously perturb the Ag surface, as is evident by the hole formed in the lower right-hand corner. The size of the hole was between 50 and 100 nm in diameter and was formed within the ∼2 min required to obtain the image. Other less obvious indications of tip-sample interaction can be seen in various areas in the images. Figure 6C is the image of a large scan area that encompassed the 1 µm × 1 µm area used for the study. A square pattern from the 1 µm × 1 µm scanning area was imprinted into the surface. The pattern of the scan area became apparent at a net load of ∼19 nN. Therefore, a subtle influence of the tip on the sample occurs before the point of sample destruction. It
Figure 7. Effect of the net repulsive force applied to the tip on the (A) apparent Ag particle count, (B) height, and (C) radius.
Figure 6. (A) A 1 µm × 1 µm area scanned at a net repulsive force of ∼5 nN (capillary forces only) and (B) the same area as in (A) scanned at a net applied load of ∼23 nN. The image in (C) is a larger area scan, encompassing the 1 µm × 1 µm area in (A) and (B). Reduced to 80% of original for publication.
should be noted that the force values reported are approximate due to the uncertainty in the spring constants of the cantilevers (range of 0.01-0.3 N/m). Figure 7 contains plots of the quantitative results for the applied load study. From a net force of 5-19 nN, the Ag particle height and radius both increased only by ∼7%. At 23 nN, there was a large increase in the particle height and radius. The increase in height is likely to be partially the result of the formation of the hole since, along the perimeter of the hole, particles appear piled upon one another. It is also likely that a contribution to the increase in particle height results from particles being forced onto other particles throughout the entire scan area. The number of particles decreased steadily over the force range, which supports the suggestion that the surface morphology was influenced by the tip well before surface perturbation was obvious. At 23 nN,
there was a large decrease in the number of particles, coincident with the formation of the hole. Both the strength of the chemical interaction between the Ag and the substrate and the substrate surface roughness likely affect the force required to move Ag particles. The effect of the adhesion (attractive) force between the tip and sample on the quality of the AFM images was also investigated. The greatest contribution to the adhesion force when imaging in air is from the capillary forces between the tip and sample.25,34 The magnitude of the capillary force is proportional to the thickness of the moisture layer on the surface. The capillary force can be determined by measuring the force required to pull the tip out of contact with the surface. In addition to the capillary force, the thickness of the moisture layer also changes the viscous and frictional forces acting on the tip, which can produce distortions in the image.25,34 For this experiment, the applied load was held constant at less than 1 nN and the adhesive force was varied by changing the relative humidity in the AFM enclosure. A plot of the adhesion force vs relative humidity is shown in Figure 8. From ∼20 to 35% RH the adhesion force remained constant at ∼4 nN. Beyond 35%, the capillary force increased steadily to ∼13 nN at 62% RH. These results were reasonably close to those obtained by Thundat et al. for adhesion force measurements on a mica surface.34 In addition, the rather dramatic change in adhesive force near 30% RH is consistent with the results of Hu et al., in which a phase change in the water structure on mica is observed.35 In order to investigate the effect of humidity on image quality, images of a 5 nm Ag film were obtained at relative humidity levels from 6 to 60%. In contrast to the observations of Thundat et al., within the (34) Thundat, T.; Zheng, X.-Y.; Chen, G. Y.; Warmack, R. J. Surf. Sci. Lett. 1993, 294, L939-L943. (35) Hu, J.; Xiao, X. D.; Ogletree, D. F.; Salmeron, M. Science 1995, 268, 267269.
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Figure 8. Plot of adhesion forces versus relative humidity. Force curves were obtained on a 5 nm thick Ag film deposited onto glass. The adhesive forces were calculated from the corresponding force curves assuming a cantilever spring constant of 0.2 N/m, the manufacturer-reported average for a group of tips. Error bars represent (1σ and were obtained from three force measurements at each point.
error in the measurements (from three images at each humidity), no apparent relationship between image quality and humidity was observed. The most likely explanation for the observed differences is that since the Ag particles are very large compared to atomic dimensions, any buckling of the cantilever at the humidity levels investigated is negligible.25 It can be concluded that, over a broad humidity range representative of typical laboratory air, lateral forces resulting from the moisture layer do not significantly affect the measurements of Ag-particle films. Quantification of Ag Film Surface Variability. The homogeneity of the surface morphology of a 5 nm thick Ag film on mica was quantified by obtaining several 1 µm × 1 µm scans over an area of ∼20 mm2. Each sample area was a 12 µm × 12 µm region within a 20 mm2 area. The values for each sample area were composed of the averages for five separate 1 µm × 1 µm scans taken within the corresponding 12 µm × 12 µm region. Thus, a total of 25 images were quantified over the ∼20 mm2 area. The standard deviation in particle height, radius, and count was only slightly higher over the 20 mm2 area than for the 12 µm × 12 µm regions. The average relative standard deviation for particle
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height, radius, and count over the 12 µm × 12 µm regions was 8, 4, and 4%, respectively. For the 20 mm2 area, the relative standard deviation in particle height, radius, and count from 25 measurements was 12, 6, and 7%, respectively. Thus, the deviation in the Ag particle surface structure over a ∼20 mm2 area is less than 10% and the surface is very homogeneous on a size scale relative to AFM and optical measurements.22 For comparison, the variability in the Ag surface structure for a single 1 µm × 1 µm area as a function of time was determined. A 5 nm thick Ag film on glass was scanned continuously for 1 h at room temperature and images were quantified every 10 min. From seven images, the relative standard deviation in Ag particle height, radius, and count was 10, 7, and 3%, respectively. Again, the greatest error was in the height measurement. Because there was no clear trend in the values for height, radius, or count as a function of time, the deviations are not likely due to tip-induced modifications of the morphology or surface annealing from the laser. However, variations in image quality between simultaneous scans is quite common. A possible explanation for this observation is that debris from the surface occasionally adheres to the tip causing a reduction in resolution. Eventually, the debris comes loose and image quality is restored. In conclusion, automated analysis of AFM images obtained under appropriate scanning conditions provides quantitatively meaningful measurements of Ag particle nanostructure. ACKNOWLEDGMENT This work was supported by a grant from the National Science Foundation (CHE-9311638). The authors gratefully acknowledge Alan Lo and Rex Skodje for the image analysis algorithm and helpful discussions. Received for review September 8, 1995. November 17, 1995.X
Accepted
AC950909D X
Abstract published in Advance ACS Abstracts, January 1, 1996.
