© Copyright 2004 American Chemical Society
APRIL 13, 2004 VOLUME 20, NUMBER 8
Letters Coupling of Individual Polyelectrolyte Capsules onto Patterned Substrates Marc Nolte and Andreas Fery* Max-Planck-Institute of Colloids and Interfaces, Department of Interfaces, 14424 Potsdam, Germany Received November 14, 2003. In Final Form: February 13, 2004 In this paper, we show that it is possible to direct the adhesion of polyelectrolyte microcapsules to patterned substrates. The patterned substrate contains regions of like and oppositely charged polyelectrolyte coatings: Adhesion of microcapsules is blocked by regions of the substrate containing like charges, while we find strong adhesion on oppositely charged regions. The approach is completely based on self-assembly and can be performed under ambient laboratory conditions. We demonstrate that it is thus possible to isolate individual microcapsules, with enclosed volumes on the order of femtoliters, and create arrays. This strategy could find applications in the field of combinatorial chemistry or sensing techniques where the capsules can serve as reaction volumes or containers for sensing agents, respectively.
The coupling of micron-sized reaction containers to specific positions on a substrate is interesting for development of methods in combinatorial chemistry, where the containers could serve as reaction volumes. In addition, this self-assembly technique could be applied in sensor applications where an indicator material could be stored inside the containers. We show here for the first time successful coupling of polyelectrolyte capsules to substrates that have been patterned using soft lithography. All of the processes, the production of the capsules, the surface coating, and the coupling of the two, are based on self-assembly. Using such a method, one is able to avoid both costly techniques requiring vacuum equipment and cleanroom facilities. Both containers and surface coatings can be made from a variety of materials including biocompatible components,1 rendering the principle highly versatile. All the steps for coupling the microcapsules to the substrate can be carried out in water. This is useful when it comes to filling the capsules or bioapplications where drying or even vacuum exposure might be harmful to the system. * To whom correspondence should be addressed. E-mail:
[email protected]. Fax: +49 (0)331 567-9202. (1) Decher, G. Polyelectrolyte Multilayers, an overview. In Multilayer Thin Films; Decher, G., Schlenoff, J. B., Eds.; Wiley-VCH: Weinheim, 2003; pp 1-46.
Substrate preparation and production of the microcapsules used here are based on the layer by layer (LBL) technique that was developed by Decher and co-workers.1,2 This technology allows one to coat surfaces with ultrathin layers made from various polymeric and other materials by alternating adsorption of positive and negative polyelectrolytes from aqueous solutions; see Decher Multilayer Thin Films3 for an overview. Donath and co-workers first demonstrated that hollow microcapsules can be produced using LBL. Using this technique, they showed that the multilayer coating is stable after dissolution of the colloidal core.4 Important features of these polyelectrolyte multilayer capsules in the context of this contribution are that they can be filled with various materials5-8 and used as microreactors9,10 and that their permeability properties (2) Decher, G.; Schmitt, J. Prog. Colloid Polym. Sci. 1992, 89, 160. (3) Decher, G.; Schlenoff, J. B. Multilayer Thin Films; Wiley-VCH: Weinheim, 2003. (4) Donath, E.; Sukhorukov, G. B.; Caruso, F.; Davis, S. A.; Mo¨hwald, H. Angew. Chem., Int. Ed. 1998, 37 (16), 2202-2205. (5) Tiourina, O. P.; Antipov, A. A.; Sukhorukov, G. B.; Larionova, N. L.; Lvov, Y.; Mo¨hwald, H. Macromol. Biosci. 2001, 1 (5), 209-214. (6) Lvov, Y.; Antipov, A. A.; Mamedov, A.; Mo¨hwald, H.; Sukhorukov, G. B. Nano Lett. 2001, 1 (3), 125-128. (7) Radtchenko, I. L.; Sukhorukov, G. B.; Mo¨hwald, H. Colloids Surf., A 2002, 202 (2-3), 127-133. (8) Sukhorukov, G.; Da¨hne, L.; Hartmann, J.; Donath, E.; Mo¨hwald, H. Adv. Mater. 2000, 12 (2), 112-115.
10.1021/la036144j CCC: $27.50 © 2004 American Chemical Society Published on Web 03/19/2004
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Figure 2. Fluorescence micrograph of a printed rhodaminelabeled PAH structure (quadratic nonprinted areas surrounded by a printed PAH layer).
Figure 1. (A) Distribution of heights for printing of PAH on a PEI/PSS layer pair. (B) AFM image of a printed pattern (circular nonprinted areas exposing the PSS-terminated substrate surrounded by the printed PAH layer) and a corresponding cross section.
are sensitive to pH and salt concentration11-13 allowing for controlled release or uptake of molecules through the wall. Thus they are promising candidates as containers in the applications mentioned above, provided the problem of coupling the capsules to substrates can be mastered. While we have previously investigated the adhesion properties of polyelectrolyte microcapsules on homogeneous substrates,14 little attention has been devoted so far to the coupling of those systems to patterned substrates. The possibility to couple capsules to substrates by electron beam irradiation was shown by Erokhin.15 However, in (9) Antipov, A.; Shchukin, D.; Fedutik, Y.; Zanaveskina, I.; Klechkovskaya, V.; Sukhorukov, G.; Mo¨hwald, H. Macromol. Rapid Commun. 2003, 24 (3), 274-277. (10) Da¨hne, L.; Leporatti, S.; Donath, E.; Mo¨hwald, H. J. Am. Chem. Soc. 2001, 123, 5431-5436. (11) Antipov, A. A.; Sukhorukov, G. B.; Donath, E.; Mo¨hwald, H. J. Phys. Chem. B 2001, 105 (12), 2281-2284. (12) Antipov, A. A.; Sukhorukov, G. B.; Leporatti, S.; Radtchenko, I. L.; Donath, E.; Mo¨hwald, H. Colloids Surf., A 2002, 198 (Special Issue SI), 535-541. (13) Antipov, A. A.; Sukhorukov, G. B.; Mohwald, H. Langmuir 2003, 19 (6), 2444-2448. (14) Elsner, N.; Dubreuil, F.; Fery, A. Phys. Rev. E, in press.
this approach, processing steps in a vacuum are necessary: these usually destroy the capsules. Here we use surfaces coated by LBL that have been patterned by means of microcontact printing16,17 of oppositely charged polyelectrolytes to direct the capsule adhesion. This patterning approach has first been employed by Hammond and coworkers.18-22 To pattern the substrates, we choose the polymer on polymer stamping method first used by Jiang et al.22 The resulting pattern promotes directed capsule adhesion in specific regions of the substrate. Patterning the substrate is performed in the following manner: In a first step, glass substrates were coated by a homogeneous double layer of poly(ethyleneimine)/poly(sodium-4-styrenesulfonate) (PEI/ PSS). Subsequently, fluorescently labeled poly(allylaminehydrochloride) (PAH) was stamped onto the polymer layer. The height of the printed structures was measured with tapping mode atomic force microscopy (AFM) in air. The heights that were obtained were 2.5 ( 0.3 nm, which is on the order of typical thicknesses of polyelectrolyte layers formed by adsorption. Figure 1 shows the distribution of heights obtained on different spots of different substrates by AFM and a typical AFM image including a cross section. The thickness was the same for all stamped patterns. Since fluorescently labeled polyelectrolyte (rhodamineB-isothiocyanate labeled PAH) was used for printing, the pattern could be recorded using fluorescence microscopy as shown in Figure 2. While the thickness of the adsorbed (15) Berzina, T.; Erokhina, S.; Shchukin, D.; Sukhorukov, G.; Erokhin, V. Macromolecules 2003, 36 (17), 6493-6496. (16) Kumar, A.; Whitesides, G. M. Science 1994, 263 (5143), 60-62. (17) Kumar, A.; Biebuyck, H. A.; Whitesides, G. M. Langmuir 1994, 10 (5), 1498-1511. (18) Hammond, P. T.; Whitesides, G. M. Macromolecules 1995, 28 (22), 7569-7571. (19) James, C. D.; Davis, R. C.; Kam, L.; Craighead, H. G.; Isaacson, M.; Turner, J. N.; Shain, W. Langmuir 1998, 14 (4), 741-744. (20) Berg, M. C.; Choi, J.; Hammond, P. T.; Rubner, M. F. Langmuir 2003, 19 (6), 2231-2237. (21) Hammond, P. T. Chemistry Directed Deposition via Electrostatic and Secondary Interactions: A Nonlithographic Approach to Patterned Polyelectryte Multilayer Systems. In Multilayer Thin Films; Decher, G., Schlenoff, J. B., Eds.; Wiley VCH: Heidelberg, 2003; pp 271-298. (22) Jiang, X. P.; Hammond, P. T. Langmuir 2000, 16 (22), 85018509.
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Figure 3. Scheme: Charge-controlled adhesion of microcapsules on surfaces structured by microcontact printing. Micrograph: Selective deposition of PE shells on structured surfaces. The structure has the same size as the shell diameter. The structure was printed with weakly labeled PAH, while the shells were strongly labeled; thus structure and shells are visible in the fluorescence image. The capsules are preferentially adsorbing to the nonprinted areas that are exposing the oppositely charged PSS.
layer can be measured by AFM, the fluorescence micrographs show the homogeneity of the patterning over larger areas. In previous work on capsule adhesion on homogeneous surfaces,14 we have found that PSS-terminated polyelectrolyte capsules adhere onto surfaces coated with PEI at pH 7, while no adhesion was found for the same capsules on bare glass or surfaces coated with PSS. Adhesion to oppositely charged surfaces such as those used here results in circular adhesion areas with radii that depended on the wall thickness of the capsules. The adhesion energy of PSS-terminated capsules on PEI was estimated to be in the range of 0.1 mJ/m2. The interactions in this case are expected to be comparable to the one investigated here, since one polyelectrolyte is the same (PSS) and the other has similar linear charge densities and amines as ionizable groups like PEI (PAH). For capsules with the same wall thickness as used here, adhesion disk radii on the order of 30-50% of the capsule radius are expected. This would correspond to a total adhesion energy around 10-14 J/capsule. Thus, the adhesion energy is about 7 orders of magnitude bigger than the thermal energy kBT at room temperature and spontaneous desorption can be ruled out. It is not straightforward to estimate peel-off forces from the adhesion energy as it is possible in the case of massive spherical particles. However, independent measurements of Cordeiro using a flow chamber showed that PAH/PSS capsules of higher wall thickness (and thus lower adhesion area) on PEI surfaces could withstand surface shear stresses of 12 N/m2, corresponding to peeloff forces around 160 nN, without detaching.23 We have found, using a micromanipulator, that capsules cannot be peeled off the surface but rather break leaving the adhering part behind (data not shown). Thus the anchoring is strong enough for application in fluidic systems where shear stresses and peel-off forces due to flow are orders of magnitude below those values. Consequently, we used the same conditions for directing the adhesion on the patterned substrates, as indicated in Figure 3. For the selective adhesion of polyelectrolyte capsules, a droplet of a suspension of the capsules in pH(23) Cordeio, A. L. T. Ph.D. Thesis, Universidade do Porto, Porto, 2003.
neutral water was put on the structured surfaces. The surface was carefully rinsed after 20 min of waiting and kept under water before observation using fluorescence microscopy. Figure 3 shows a typical picture of labeled shells adsorbed onto patterned substrates which clearly show that shell adhesion can be directed and that shells can be immobilized in a way that isolates them individually. The coverage in this experiment was not complete (around 40%), but we expect that full coverage can be reached by increasing the capsule concentration. In no case have we observed desorption of capsules or detachment due to flow. In the present case, we have reached a capsule density of 5 × 103 mm-2; full coverage would yield densities of 1.3 × 104 mm-2. Using microcontact printing, patterns of micron period can be created and recently it was shown by Stamou and co-workers that even smaller periods can be created and vesicles can be immobilized on them with densities of 106 vesicles/mm2. This would be the technological limit of our technique as well.24 Individual polyelectrolyte capsules have typical enclosed volumes of the order of femtoliters and can be filled with various agents.5-8,25 They are mechanically robust26 and can be produced with uniform and well-defined size. The coupling of these containers to flat substrates should open new perspectives for analyzing or influencing the interior using surface-sensitive methods. As well, the capsules can be adsorbed onto microelectrodes, which makes them accessible for electrochemical experiments. All of these techniques can be carried out with individual capsules. Following this line, the coupling is also a first step toward integrating these systems into chip architectures where they have great potential as cheap well-defined reaction containers for future application in combinatorial chemistry on the micron and submicron scale. First experiments with filled microcapsules along this line are under way in our laboratory. (24) Stamou, D.; Duschl, C.; Delamarche, E.; Vogel, H. Angew. Chem., Int. Ed. 2003, 42 (45), 5580-5583. (25) Mo¨hwald, H.; Donath, E.; Sukhorukov, G. Smart Capsules. In Multilayer Thin Films; Decher, G., Schlenoff, J. B., Eds.; Wiley VCH: New York, 2003; pp 363-392. (26) Dubreuil, F.; Elsner, N.; Fery, A. Europhys. J. E 2003, 12 (2), 215-221.
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Experimental Section Poly(sodium-4-styrenesulfonate) (PSS, Mw ) 70 000 g/mol), poly(allylamine-hydrochloride) (PAH, Mw ) 70 000 g/mol), and branched poly(ethyleneimine) (PEI, Mw ) 25 000 g/mol) were purchased from Aldrich. Rhodamine-B-isothiocyanate was purchased from Fluka. Dispersions of melamine formaldehyde (MF) particles with a diameter of 5.1 ((0.09) µm (monodisperse, 10% in volume fraction) were purchased from Microparticles GmbH (Berlin, Germany). All chemicals were used without further purification except for the PSS, which was dialyzed against water (Mw cutoff ) 14 000 g/mol; Millipore, Schwalbach, Germany) and lyophilized prior to use. For all experiments and for all cleaning steps, deionized water from a Purelab Plus UV/UF (Elga LabWater, Germany) with a resistance of 18.2 MΩ cm-1 was used. Rhodamine-B-isothiocyanate labeled PAH (RBITC-PAH) was prepared as described by Richter et al.27 based on Ibarz et al.28 Fluorescence microscopy measurements were carried out on a Zeiss Axiovert 200 (Zeiss, Germany) for transparent samples and on a Zeiss Axioscope 2 plus for nontransparent samples. As objectives, on the Axiovert a LD A-Plan 20× and on the Axioscope a Plan Apochromat 20× were used. The light source was a Hgvapor lamp. The excitation and the emission were filtered between 515-565 nm and 450-490 nm, respectively. A Zeiss AxiocamHR high-resolution monochromatic camera was used. The capsules were prepared according to Donath et al.29 MF particles were coated by alternating adsorption of positively and negatively charged polyelectrolytes (PEs) from aqueous solution (even number of layers). (PE concentration, 1 mg/mL; 0.5 M NaCl). Subsequently, the MF particles were dissolved in 0.1 M HCl solution. Completeness of particle removal and thickness of the (27) Richter, B.; Kirstein, S. J. Chem. Phys. 1999, 111 (11), 51915200. (28) Ibarz, G.; Da¨hne, L.; Donath, E.; Mo¨hwald, H. Adv. Mater., in print. (29) Donath, E.; Sukhorukov, G. B.; Caruso, F.; Davis, S. A.; Mo¨hwald, H. Angew. Chem., Int. Ed. 1998, 37 (16), 2202-2205.
Letters capsule walls were determined by AFM imaging of capsules dried on mica using a Nanoscope IIIa AFM in Tapping Mode. The dry thickness measured by scanning force microscopy after dissolving the core was 2.8 ( 0.3 nm per layer pair. Capsules were prepared with 5 layer pairs before dissolving the core. After dissolution, one layer of PSS and one layer of RBITC-PAH were added. Coverslips (Menzel-Glaser, Braunschweig, Germany) and the silicon wafers (Wacker, Burghausen, Germany; orientation 100, naturally oxidized) were cleaned by the RCA method30 prior to coating. For the coating, the slides were put in a solution of 0.5 mg/mL PEI without added salt for 20 min, rinsed carefully, and immersed into a 1 mg/mL solution of PSS containing 0.5 M NaCl. The surfaces were stored under water before use. The silicon master used for producing microcontact-printing stamps was obtained from GeSim (Grosserkmannsdorf, Germany) and hydrophobized using heptadecafluoro-1,1,2,2-tetrahydrodecyldimethylchlorsilan (ABCR, Germany). PDMS Base Silicon Elastomer Sylgard 184 and the curing agent (Dow Corning Midland, MI) were mixed in a 10:1 ratio, respectively, and degassed in a vacuum. The masters were covered with the prepolymer, degassed again, and cured for 12 h at 60 °C. The stamps were cut out and hydrophilized31 in an air-plasma (Harrick PDC-32G-2; pressure, 2 × 10-1 mbar; 25 s). Before the stamping was carried out, a 1 mg/mL solution of RBITC-PAH with an ionic strength of 0.5 M (NaCl) was spread on the stamp. The pH of the solution was 8.7. After 15 min, the stamps were briefly rinsed with water and excess solution was removed in a stream of nitrogen. The inked stamps were brought into contact with dried, coated silicon wafers or coverslips for 20 min and then removed. The structured surfaces were carefully rinsed with water and stored under water until use.
Acknowledgment. We thank Helmuth Mo¨hwald and Ana Cordeiro for stimulating discussions. LA036144J (30) Kern, W.; Puotinen, D. A. RCA Rev. 1970, 31, 187-206. (31) Jiang, X. P.; Zheng, H. P.; Gourdin, S.; Hammond, P. T. Langmuir 2002, 18 (7), 2607-2615.
