Langmuir 1999, 15, 2999-3002
Anomalous Lateral Size Measurements by Atomic Force Microscopy in a Fluid Cell G. A. Neff, D. E. Gragson, D. A. Shon, and S. M. Baker* Department of Chemistry, Harvey Mudd College, Claremont, California 91711 Received September 28, 1998
Introduction Use of an atomic force microscope to image soft or deformable samples in water or other fluids has become routine.1 Such imaging is often performed in order to provide a controlled and natural environment for biological samples,2,3 but it also has applications in chemistry4,5 and geology.6,7 More generally, AFM imaging is performed in a fluid environment in order to lower or eliminate capillary forces on the tip.8,9 These forces arise from an interaction between the AFM tip and a thin water layer that often exists on the sample surface in ambient conditions. This interaction results in a strong attractive force that pulls the tip into contact with the surface. With soft or deformable samples, such as polymer films or biological samples, this additional force can change the magnitude of intended force applied to the surface and may cause the tip to damage the sample. Submersion of the tip and sample in water produces an isotropic and thus nondestructive force.8,9 Our interest in fluid cell AFM is directed toward studying monomolecular films of adsorbed diblock copolymers, specifically polystyrene-b-poly(ethylene oxide), where one block is attached to the surface while the other remains free to interact with an adjacent liquid phase. The use of a fluid cell with these systems results in a minimization of deformation of the diblock copolymer samples and allows us to compare the structure of these films in solvents of differing solvent quality. The structure of adsorbed polymers on surfaces is of great importance to many applications, including chromatography, adhesion,10 colloidal suspension and stabilization in solution,11 lubrication,12,13 and biocompatibility of artificial organs in medicine.14 (1) Drake, B.; Prater, C. B.; Weisenhorn, A. L.; Gould, S. A. C.; Albrecht, T. R.; Quate, C. F.; Cannell, D. S.; Hansma, H. G.; Hansma, P. K. Science 1989, 243, 1586. (2) Radmacher, M.; Tillmann, R. W.; Fritz, M.; Gaub, H. E. Science 1992, 257, 1900. (3) Baty, A. M.; Leavitt, P. K.; Siedlecki, C. A.; Tyler, B. J.; Suci, P. A.; Marchant, R. E.; Geesey, G. G. Langmuir 1997, 13, 5702. (4) Uchida, E.; Ikada, Y. Macromolecules 1997, 30, 5464. (5) Hong, Q.; Suarez, M. F.; Coles, B. A.; Compton, R. G. J. Phys. Chem. 1997, 101, 5557. (6) Hillner, P. E.; Gratz, A. J.; Manne, S.; Hansma, P. K. Geology 1992, 20, 359. (7) Putnis, A.; Junta-Rosso, J. L.; Hochella, M. F., Jr. Geochim. Cosmochim. Acta 1995, 59, 4623. (8) Magonov, S. N.; Whangbo, M.-H. Surface Analysis with STM and AFM; VCH Publishers: New York, 1996. (9) de Souza, E. F.; Douglas, R. A.; Teschke, O. Langmuir 1997, 13, 6012. (10) Lee, L. H. Adhesion and Adsorption of Polymers; Plenum: New York, 1980. (11) Napper, D. Polymeric Stabilization of Colloidal Dipersions; Academic: London, 1983. (12) Uyama, Y.; Tadokoro, H.; Ikada, Y. J. Appl. Polym. Sci. 1990, 39, 489. (13) Inoue, H.; Uyama, Y.; Uchida, E.; Ikada, Y. Cell. Mater. 1992, 2, 21. (14) Ruckenstein, E.; Chang, D. B. J. Colloid Interface Sci. 1988, 123, 170.
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While investigating PS/PEO films on Si surfaces in various solvents with our commercial AFM, we observed that images taken in air using the fluid cell attachment were significantly different compared to those taken without the fluid cell attachment. Specifically, the images obtained with the fluid cell appeared laterally magnified compared to those taken without the fluid cell. In an effort to quantify these observed differences, a holographic grating was imaged both with and without the fluid cell in air. A comparison between the images obtained from the holographic grating with and without the fluid cell shows marked differences in the observed groove density. We attribute these differences to effects resulting from the O-ring that is used to seal fluids in the fluid cell attachment. Our experiments exploring the effects of solvent quality require that we use a more chemically inert O-ring material such as viton, rather than the standard silicone O-ring. Thus we have characterized this O-ring-induced magnification for both silicone and viton O-rings. In the fluid cell attachment the O-ring must be compressed against the sample (which is held magnetically against the piezoelectric scanner) in order to obtain a good seal. The piezoelectric scanner controls the fine positioning of the sample during AFM imaging by expanding or contracting a known fractional displacement as a function of applied voltage.15 These scanners are thus calibrated by the manufacturer to account for the characteristic response of the material for a particular geometry. The additional force of the O-ring against the piezo results in smaller displacements as a function of the applied voltage than that of the calibration of the piezoceramic relative to free expansion. The distortion due to the additional force from the O-ring results in lateral magnification of images observed when using the fluid cell attachment. While distortion in the z direction likely results from use of the fluid cell as well, such effects will not be addressed here. As we discuss in detail below, the observed lateral distortions are not quantitatively reproducible in their magnitude. Thus, we suggest that images should be obtained without the fluid cell prior to fluid cell experiments for the purposes of calibration. Experimental Section The holographic film grating used as a lateral calibration standard for AFM was cut from a card-mounted linear diffraction grating with nominally 1000 lines/mm from Edmunds Scientific. The silicon substrates used were 0.5 in. diameter p-type Si(100) disks from Silicon Quest International. The diblock copolymer used was 182.7K MW PS(95.8%)/PEO, Mw/Mn ) 1.07, from Polymer Laboratories, Inc., Amherst, MA. After routine cleaning of the Si substrates,16 polymer films were adsorbed at room temperature from approximately 0.1 mg/mL solutions of the PS/ PEO in toluene for 1 week. Following adsorption, the samples were rinsed in toluene and allowed to dry. Toluene, 99.8% HPLC grade, was used as received from Sigma-Aldrich. All images shown were obtained using a Digital Instruments (Santa Barbara, CA) Nanoscope E with a scanning head of nominally 15.0 × 15.0 µm2 scan range. Samples were imaged in air in contact mode, keeping the applied force constant and using commercially available Si3N4 V-shaped cantilevers (nominally 0.06 N/m). Samples were attached to the substrate using both a soft adhesive (double-sided tape) and a hard adhesive (Su(15) Howland, R.; Benatar, L. A Practical Guide to Scanning Probe Microscopy; Park Scientific Instruments: 1996. (16) Frantz, P.; Granick, S. Langmuir 1992, 8, 1176.
10.1021/la981334t CCC: $18.00 © 1999 American Chemical Society Published on Web 03/19/1999
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perGlue, cyanoacrylate). All images are shown without any image processing other than flattening and plane fitting.
Results and Discussion A holographic grating was chosen as a standard because it is relatively nondeformable and has features of known lateral dimensions. With nominally 1000 grooves/mm, we should observe approximately 1 groove/µm. Figure 1a shows an AFM image of the grating in air obtained without use of the fluid cell. In this image 13 grooves are observed in the 15.0 × 15.0 µm2 area. Figure 1b shows a 15.0 × 15.0 µm2 AFM image of the same grating taken in air using the fluid cell attachment with a silicone O-ring. Here, only 12 grooves are observed. Thus, in this case the lateral feature size observed using the fluid cell with the silicone O-ring appears to be 1.08 times larger than the image obtained without the fluid cell. Figure 1c shows a 15.0 × 15.0 µm2 AFM image obtained in air in the fluid cell with a viton O-ring, producing features 1.23 times larger (only 9-10 grooves are observed) than those observed without the fluid cell. The observed distortions are not reproducible in their magnitude from experiment to experiment. Upon changing the seating of the O-ring on the piezo, different magnitudes of distortion are often observed, presumably resulting from different loads applied to the piezo with different seatings. We have observed (in a limited sampling of gratings) amplifications of the grating features of 1.04-1.11 with the silicone O-ring and amplifications of 1.23-1.45 with the viton O-ring. Thus, the amplifications observed in using both the silicone and viton O-rings (Figure 1b and c, cannot be treated as constants or used as calibration corrections. While piezoelectric scanners can exhibit intrinsic nonlinearity and hysteresis from ideal behavior that can result in distortion of images,15 the observed magnification is not a consequence of either phenomenon. Nonlinearity would result in nonuniform spacings and/or curvature of linear structures, while hysteresis could cause shifting of data collected in one scan direction relative to that collected in the opposite direction. Creep is another phenomenon known to occur in piezoelectric materials and results when an abrupt change in voltage is applied to the material. The material changes dimensions in two steps, the first very fast and the second very slow. Creep, the slow step, can cause the observation of different length scales (magnification) in images taken at different scan speeds.15 All the images which we compare to one another were taken at the same scan speeds and feedback conditions to minimize any consequences of creep. As discussed previously, the O-ring in our fluid cell attachment is sandwiched between the sample and the fluid cell attachment, often very tightly to form a reliable seal. This compression adds an additional force to the piezoelectric crystal and impedes its expansion as it scans during an image acquisition. Thus, for a given applied voltage the piezo material does not respond in accordance with factory calibration. The observed features appear larger in the fluid cell because the scanner is actually moving over a smaller scan area than is calculated by the calibration. This effect is not observed in the conventional setup, which lacks the extra force of the O-ring. The viton O-ring is stiff and, as expected, exerts a larger resistive force on the piezo and causes greater distortion of images than does the more compressible silicone O-ring. We have performed stiffness measurements employing a MTS Model 830 elastomer test system on both O-rings and found that the viton O-ring is 3-4 times more stiff
Figure 1. AFM images (15.0 × 15.0 µm2) of a holographic film grating in ambient conditions. The gray scale ranges from 0 nm (black) to 160 nm (white). Images are taken (a) without the fluid cell, (b) with a fluid cell with a silicone O-ring, and (c) with a fluid cell with a viton O-ring.
that the silicone O-ring. In addition, the viton O-ring exhibits a nonlinear compressibility, resulting in an increase in the stiffness with increases in the displacement that was not observed for the silicone O-ring. The higher stiffness measured for the viton O-ring, coupled with the larger lateral amplifications that are observed when using the viton O-ring, provides excellent support for our
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hypothesis of a force-induced distortion. Further, when imaging smaller areas, less than 2.5 × 2.5 µm2, using the viton O-ring, the amplification is often not as dramatic as that when imaging larger areas. This observation is again supported by our stiffness measurements, which show that the stiffness of the viton O-ring increases with increasing displacement. The distortions and amplifications observed when using the fluid cell attachment can be more subtle or can be easily overlooked when imaging a surface in which the topology is not regular, as in the case of an adsorbed polymer film. We have observed amplifications and distortions in AFM images from PS/PEO films adsorbed on an oxide-coated Si substrate. Figure 2a shows a 15.0 × 15.0 µm2 AFM image of a typical PS/PEO film obtained in air without use of the fluid cell. The image shows a fine network structure which we commonly observed for films adsorbed from solutions of this particular MW polymer in toluene. This network structure is fairly uniform across the entire scan area and under these feedback conditions can be reproducibly imaged over several scans without significant deformation. For comparison the same sample was imaged in the fluid cell in ambient conditions using the silicone (Figure 2b) and viton (Figure 2c) O-rings. Features are quantitatively compared in these images in the following manner. Using the Nanoscope software, three different cross sections are examined in each image, and four different features within each cross section are measured laterally. The twelve resulting feature sizes are then averaged. The average lateral sizes in the fluid cell images are compared to the images obtained without the fluid cell by calculating an amplification factor relative to the nonfluid cell image. For the image taken in the fluid cell with the silicone O-ring (Figure 2b) a network structure is again observed, but the features now appear approximately 1.25 times larger than the features in the image obtained without the fluid cell. The image taken in the fluid cell with the viton O-ring (Figure 2c) shows features 1.50 times larger than those without the fluid cell. Figure 2 is representative of the average magnification observed with the polymer samples. Smaller and larger magnifications have been observed depending on the degree of O-ring compression for a particular O-ring seating. In the extreme limit magnifications as large as 1.6 and 3.3 have been observed with the silicone and viton O-rings, respectively. We have developed a simple protocol which allows us to calibrate our images obtained in the fluid cell. The sample is imaged first without the fluid cell to calibrate feature size. The sample is then imaged in the fluid cell in air in order to compare observed feature sizes to those observed without the fluid cell for a particular O-ring seating. Then fluid can be added according to the needs of the particular experiment. Further comparison could also be performed after imaging in the fluid cell, by allowing the sample to dry and then reimaging in air in the dry cell. This latter comparison is only valid under conditions where the fluid does not change the film structure. Such a procedure has become routine for us when imaging polymer films with our fluid cell AFM attachment to ensure accurate lateral dimensions. Conclusions The images shown here clearly indicate anomalous size measurements when using a fluid cell in AFM imaging. Specifically, larger features (magnification) are observed when imaging in air with the fluid cell attachment than when imaging without the fluid cell. Magnification was
Figure 2. AFM images (15.0 × 15.0 µm2) of a PS/PEO copolymer film adsorbed onto SiO2/Si from toluene solution, imaged in ambient conditions. The gray scale ranges from 0 nm (black) to 10 nm (white). Images are taken (a) without the fluid cell, (b) with a fluid cell with a silicone O-ring, and (c) with a fluid cell with a viton O-ring.
observed in images of a nondeformable holographic grating as well as a diblock copolymer film. Use of both silicone and viton O-rings to seal the fluid cell to the sample caused noticeable distortions. We attribute these distortions to the additional force that the O-ring exerts on the piezoelectric scanner, impeding its unrestricted expansion and changing its performance relative to factory calibration. The more compressible silicone O-ring, standard for use
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with water, caused less distortion than the stiffer viton O-ring, necessary for organic solvents. The observed distortion resulting from this additional force is not reproducible in magnitude from experiment to experiment. Therefore, calibration prior to and possibly after fluid cell imaging of every sample adds little time to the experiment but provides valuable information that is necessary for reporting reliable lateral length scales.
Notes
Acknowledgment. The authors thankfully acknowledge Professor Joe King for assistance with the stiffness measurements and for useful discussions concerning O-ring compressibility. The authors would also like to acknowledge the support of the National Science Foundation (DMR 9623718) and the Department of Energy through a PECASE award. LA981334T
