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Evaluation of the Structure of Polydiacetylene Monolayers Using Fluorescence Microscopy and Scanning Tunneling Microscopy Troy E. Wilson,tJ D. Frank Ogletree,$ Miquel B. Salmeron,*and Mark D. Bednarski'PtJ Department of Chemistry, University of California at Berkeley, and the Center for Advanced Materials, Material Sciences Division, Lawrence Berkeley Laboratory, Berkeley, California 94720 Received January 2, 1992. In Final Form: August 6, 1992 This communication describes a structural study of polydiacetylene monolayers supported on highlyoriented pyrolytic graphite (HOPG).Both scanning tunneling microscopy (STM) and scanning laser confocal fluorescence microscopy were used to evaluate monolayer structure at the micrometer level. These studies demonstrate that extreme tunneling conditions (10 PA, 10 V) may be required to obtain images that are consistent with those of high-resolution fluorescence microscopy results. STM imaging using lower values of gap resistancedamaged or removed the organic monolayer from the graphite surface. The correlationbetween opticaland scanned microscopies clearly indicatesthe utility of STM for studying the structure of polydiacetylene films. Introduction Scanning tunneling microscopy (STM) provides an intriguing perspective on the microscopic structure of biological and organic materials.l Recent STM investigations have focused on the structures of nucleic a~ids,~-5 proteins,H eelf-assembledand Langmuir-Blodgett mono1ayers,+l1 and liquid crystalline overlayers.l2-14 We are developing polydiacetylene monolayer films to control biological adhesion and required an understanding of the microscopic structure of these materials.l5J8 This paper reports the use of STM and confocal fluorescence microscopy to investigate the structure of polydiacetylene monolayers.17J8 These studies demonstrate that under extreme tunneling conditions (10 PA, 10 V), STM can be used to image polydiacetylene films and these images are consistent with high-resolution confocal fluorescence micrographs.
Experimental Section Scanning Tunneling Microscopy. The STM instrument is similar to a double-tube design reported by Lyding.lg The STM was controlled using electronics developed at Lawrence Berkeley Laboratory and the data were acquired using a PC compatible computer equipped with a 12-bit 150-kHz A/D converter. Imaging was performed in ambient and all images were obtained in the topographic (constant current) mode of operation using tips mechanically cut from 1mm diameter Pt40% Rh alloy wire. Typical tunneling parameters were bias voltage of 10 V (sample negative) and a tunneling current of 10 PA. Images were recorded using 256 points per line and 128lines per image and the images presented are unprocessed. Confocal Fluorescence Microscopy. High-resolutionlaser scanningconfocalfluorescencemicrographswere obtained using a Zeiss LSM-10 with a 633/543 He-Ne laser.
Results and Interpretation The polydiacetylene fiims (Figure 1A)were constructed from N-(2-hydroxylethyl)-10,12-pentacdynamide(Wt Department of Chemistry, University of California, Berkeley, CA 94720. DA). Monolayers of EAPDA were formed a t an air-water t Center for Advanced Materials, Lawrence Berkeley Laboratory, interface, polymerized, and transferred to HOPG using Berkeley, CA 94720. methods described elsewhere.l5 We have previously (1) Hansma, P. K.; Eliigs, V. B.; Marti, 0.;Bracker, C. E. Science studied polymerized EAPDA monolayers supported on 1988,242, 209. (2) Driscoll, R. J.; Youngquist, M. G.; Baldeschwieler, J. D. Nature silicon and glass surfaces with the techniques of X-ray 1990,346, 294. photoelectron spectroscopy (XPS) and contact-angle (3) Dunlap, D.; Bustamante, C. Nature 1989, 342, 204. measurements and have confirmed that these films form (4) Arscott,P. G.; Lee, G.; Bloomfield, V. A.;Evans,D. F. Nature 1989, 339,484. monolayers with the ethanolamine moieties directed (5) Beeb, T. P.; Wilson, T. E.;Ogletree, D. F.; Katz, J. E.; Balhorn, toward the ambient i n t e r f a ~ e For . ~ ~this ~ ~ study, ~ highR.; Salmeron, M. B.; Siekhaus, W. J. Science 1989,243, 370. resolution optical micrographs were acquired using laser (6) Hauseling, L.; Michel, B.; Ringsdorf, H.; Rohrer, H. Angew. Chem., scanning confocal fluorescence microscopy and the STM Int. Ed. Engl. 1991,30, 569. (7) McMaster, T. J.; Carr, H.; Miles, M. J.; Cairns, P.; Morris, V. J. images were obtained using an instrument that has been Vac. Sei. Techol. A 1990,8, 648. described previously and was modified to operate at large (8) Guckenberger, R.; Wiegrabe, W.; Hillerbrand, A.; Hartmann, T.; gap-resistance values. All imaging was performed using Wang, 2.;Baumeister, W. Ultramicroscopy 1989, 31, 327. (9) Widrig, C. A.; Alvea, C. A.; Porter, M. D. J. Am. Chem. SOC.1991, EAPDA monolayers supported on HOPG and the exact 113, 2805. image dimensions and acquisition parameters are de(10) Sotobayashi, H.; Schilling,T.; Tesche,B. Langmuir 1990,6,1246. scribed in the figure caption. (ll)Heckl,W.M.;Kallury,K.M.R.;Thompson,M.;Gerber,C.;Horber, J. K. H.; Binnig, G.Langmuir 1989, 5, 1433. (12) Smith, D. P. E.; Horber, J. K. H.; Binnig, G.; Nejoh, H. Nature
1990,344,641. (13) Smith, D. P. E.; Horber, J. K. H.; Gerber, Ch.; Binnig, G. Science 1989,245, 43. (14) Foster, J. S.; Frommer, J. E. Nature 1988,333, 542. (15) Wilson, T. E.; Bednarski, M. B. Langmuir 1992,8, 2361. (16) Mastandrea, M.; Wilson, T. E.; Bednarski, M. B. J.Mater. Educ. 1989, 1 1 , 529. (17) Highly oriented pyrolytic graphite (grade ZYA and ZYB) was
graciously donated by A. W. Moore, Union Carbide Corp., Cleveland, OH.
(18)The use of HOPG as a substrate for STM studies of biological and organic materials has been increasingly called into question. We wanted to compare our STM data with independent optical results to alleviate the risk of misinterpreting artifacts associated with the graphite surface. For a recent review, see Clemmer, C. R.; Beebe, T. P. Science 1991,261, 640, and references contained therein. (19)Lyding, J. W.; Skala, S.; Hubacek, J. S.; Brockenbrough, R.; Gammie, G. J. Microsc. 1988, 152, 371. (20) We have not included ellipsometric measurements in our analysis of the f i i thickness since the light-absorbing nature of the polydiacetylene monolayers renders these results unreliable.
0743-7463/9212408-2688$03.00/0 0 1992 American Chemical Society
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Figure 1. (A) Schematic structure of a polymerized EAPDA monolayer supported on graphite. (B) High-resolution laser scanning confocal fluorescence micrograph of an EAPDA film supported on HOPG. The magnification is 6oooOX and the scale indicates 10 pm. (C) An STM image (2.9 pm X 3.4 pm) of an EAPDA film supported on HOPG. The image was obtained using a mechanically cut Pt/Rh tip a t a 1 TQgap resistance (10 pA and 10 V, sample negative) and a tip velocity of 4.3 pm/s. (D)An STM image of the same area as Figure 2B (2.7 pm X 3.4 pm) acquired a t 13 GQ (10PA, 130 mV, sample negative). The film has been removed from the HOPG surface by reduction of the tip-sample separation (gap resistance) and the film structure is not recovered upon returning to the l-TI2 gap resistance. Note that the vertical scale for Figure 1C is 100 A and for Figure 1D 20 A.
Both optical microscopy (Figure 1B) and STM (Figure
1C)suggest that the structure of the monolayers is a closepacked array of polymer fibers. Furthermore, variations in the spacing and orientation of the polymerized fibers are resolved by the two techniques. For instance, both linear and radial arrangements of fibers appear to extend from nucleation sites which may result from variations in temperature or local lipid geometry during polymerization. In ordered monolayer films, the polymerization of amphiphiles usually leads to the formation of defects and we suspectthat the dark areas of the imagesare “holes”formed
during contraction of the monolayer upon polymerization.21-23 The STM image shows the structure and defects (e.g. “holes”) of the film at an order of magnitude greater resolution than the fluorescence micrographs. Single line scans obtained using the STM reveal that these depressionsare approximately30-40 A in depth, consistent with a monolayer structure. (21) Ringsdorf, H.; Schlarb, B.; Venzmer, J. Angew. Chem., Int. Ed. Engl. 1988,27,113. (22) Kuo, T.; OBrien, D. F. Longrnuir 1991, 7, W. (23) Dom, K.; Ringsdorf, H. Contemp. Top. Polym. Sci. 1984,5,73.
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It should be emphasized that extreme tunneling conditions were required to faithfully image the monolayer surface; the STM image shown in Figure 1C was acquired using a gap resistance of 1 Since polydiacetylene films are bound to graphite supports only by weak, cooperative van der Waals interactions, a relaxation of the tunneling conditions should irreversibly remove the film and expose the underlying HOPG support.25 Figure 1D was obtained by imaging at a reduced gap resistance (13 Gn) for the identical scan area as that in Figure 1C. Note that the graphite step is visible in both parts C and D of Figure 1,but the film structure shown in Figure 1C has been removed in Figure 1D. Furthermore, returning the gap resistance to the original value (1 Til) did not recover the film structure. A subsequent image expanded around Figure 1D revealed a square depression in the film surface approximately 40 A in depth surrounded by debris. (24) The gap resistance is defined as the magnitude of the bias voltage divided by the magnitude of the tunneling current. Gap resistance is a useful parameter for describing tipsample separation which increases monotonically as the gap resistance increases. (26) Wileon, T. E.; Murray, M. N.; Ogletrea, D. F.; Bednarski, M. D.; Cantor, C. R.; Salmeron, M. B. J. Vac. Sci. Technol. B 1991,9, 1171.
Letters In summary, these results demonstratethat at appropriate imaging conditions it is possible to both assess the structure of organic materials on HOPG and selectively modify the film structure using scanning tunneling microscopy.
Acknowledgment. This work was supported by the Diredor, Office of Energy Research, Basic Energy Science, Materials Science Division of the US. Department of Energy under contract number DE-AC03-76SF00098 and by the National Institutes of Health Award R29 GM4303702. TEW warmly thanks the Howard Hughes Medical Institute and the Fannie and John Hertz Foundation for Predoctoral Fellowships. MDB thanks the American Cancer Society for a Junior Faculty Award 1990-1993 JFRA-261 and the Eli Lilly Corporation for a Young Investigator Award. Supplementary Material Available: Additional fluorescence micrograph and STM images as well as an STM single line scan (2 pages). Ordering information is given on any current masthead page. Registry No. EAPDA, 143345-01-1; graphite, 7782-42-5.
