7402
Langmuir 2001, 17, 7402-7405
A Simple Method for Biocompatible Polymer Based Spatially Controlled Adsorption of Blood Plasma Proteins to a Surface William Inglis,† Giles H. W. Sanders,‡ Philip M. Williams,*,† Martyn C. Davies,† Clive J. Roberts,† and Saul J. B. Tendler† Laboratory of Biophysics and Surface Analysis, School of Pharmaceutical Sciences, The University of Nottingham, University Park, Nottingham NG7 2RD, U.K., and Chemistry Department, Zeneca/SmithKline Beecham Centre for Analytical Sciences, Imperial College of Science, Technology and Medicine, London SW7 2AY, U.K. Received April 5, 2001. In Final Form: July 30, 2001 Using the soft lithographic technique, microcontact printing, we demonstrate an example of how a biocompatible polymer can be easily patterned upon another polymer substrate, creating a surface with two spatially different properties. A poly(dimethylsiloxane) mold allowed the patterning of a negative replica of an E-PROM microchip, forming a spatially defined pattern with a period of approximately 1 µm. The amphiphilic biocompatible polymer, poly(lactic acid)-poly(ethylene glycol) (PLA-PEG), was used as the “ink” in order to block off areas of a hydrophobic, polystyrene (PS) substrate. This created a surface with two properties: 2-3 µm strips of PLA-PEG polymer, which resists protein adsorption, divided by micrometer strips of PS. The ability of the patterning technique to provide a true heterogeneous surface was analyzed using atomic force microscopy, while fluorescence microscopy provided a high-contrast method by which to trace the position of specific molecules. We believe this technique to be an elegant demonstration of how the properties of polymers can be exploited in order to arrange molecules at a surface, avoiding the more difficult use of gold, self-assembly, or self-assembled monolayers.
Introduction The continual drive toward the miniaturization of scientific techniques has resulted in an expansion of technologies that challenge conventional methods for creating microscopic structures and patterns. This field is loosely defined as microfabrication. Well-established techniques such as UV lithography, electron beam fabrication, and XY lithography are becoming limited by the fundamental barriers of diffraction and the harsh conditions they must be carried out in.1 This has prompted an increase in investigations into the use of biological and organic materials to form new generation electrical devices. In conjunction with the development of these possibilities has come the ability to miniaturize chemical sensing.2,3 Microfabrication technologies are also of use to cell biologists, in that the ability to spatially control cell adhesion may lead to new tissue engineering technology.4,5 In addition to these, the ability to control molecular scale combinatorial chemistry and molecular assembly has been enhanced by the development of new microfabrication techniques. The successes demonstrated in these fields suggest that micro- and nanoscale technologies will become of far greater significance in the near future. * To whom correspondence should be addressed. E-mail:
[email protected]. † The University of Nottingham. ‡ Imperial College of Science, Technology and Medicine. (1) Xia, Y.; Whitesides, G. M. Angew. Chem., Int. Ed. 1999, 37, 550575. (2) Sanders, G. H. W.; Manz, A. TrAC, Trends Anal. Chem. 2000, 19, 364-378. (3) Zlatanova, J.; Bavykin, S.; Mirzabekov, A. Biophys. J. 1999, 76, A456-A456. (4) (a) Folch, A.; Toner, M. Biotechnol. Prog. 1998, 14, 388-392. (b) Xia, Y. N.; Tien, J.; Qin, D.; Whitesides, G. M. Langmuir 1996, 12, 4033-4038. (5) Ghosh, P.; Amirpour, M. L.; Lackowski, W. M.; Pishko, M. V.; Crooks, R. M. Angew. Chem., Int. Ed. 1999, 38, 1592-1595.
A possible advancement of microfabrication may be sought in soft lithography, the generic name for a family of techniques that use an elastomer to form a high-quality negative when cured upon an etched master template.6,7 This mold can then be used in a variety of methods to transfer solution of molecules or modify surfaces to a negative relief of the pattern. Examples of these techniques include microfluidic networking (µFN),8-10 micromolding in capillaries,11,12 and microcontact printing (µCP).13,14 Xia et al. provide an in depth discussion upon most of these techniques.15 Of the many useful elements of these methods, perhaps the most striking are the simplicity and versatility of their application. µFN, for example, relies solely upon capillary force to pull reagents through a channel, reacting with molecules on the surface. Moreover, soft lithography can be achieved upon nonplanar surfaces16,17 and can be used to provide foundations for threedimensional assembly of surfaces.18,19 (6) (a) Kumar, A.; Whitesides, G. M. Appl. Phys. Lett. 1993, 63, 20022004. (b) Xu, S.; Miller, S.; Laibinis, P. E.; Liu, G. Y. Langmuir 1999, 15, 7244-7251. (c) Zhao, X. M.; Wilbur, J. L.; Whitesides, G. M. Langmuir 1996, 12, 5504-5504. (7) Delamarche, E.; Schmid, H.; Michel, B.; Biebuyck, H. Adv. Mater. 1997, 9, 741-746. (8) Patel, N.; Sanders, G. H. W.; Shakesheff, K. M.; Cannizzaro, S. M.; Davies, M. C.; Langer, R.; Roberts, C. J.; Tendler, S. J. B.; Williams, P. M. Langmuir 1999, 15, 7252-7257. (9) Biebuyck, H. A.; Larsen, N. B.; Delamarche, E.; Michel, B. IBM J. Res. Dev. 1997, 41, 159-170. (10) Delamarche, E.; Bernard, A.; Schmid, H.; Michel, B.; Biebuyck, H. Science 1997, 276, 779-781. (11) Kim, E.; Xia, Y. N.; Whitesides, G. M. Nature 1995, 376, 581584. (12) Xu, J. D.; Locascio, L.; Gaitan, M.; Lee, C. S. Anal. Chem. 2000, 72, 1930-1933. (13) Bernard, A.; Renault, J. P.; Michel, B.; Bosshard, H. R.; Delamarche, E. Adv. Mater. 2000, 12, 1067-1070. (14) Wilbur, J. L.; Kumar, A.; Biebuyck, H. A.; Kim, E.; Whitesides, G. M. Nanotechnology 1996, 7, 452-457. (15) Xia, Y. N.; Whitesides, G. M. Annu. Rev. Mater. Sci. 1998, 28, 153-184.
10.1021/la010511e CCC: $20.00 © 2001 American Chemical Society Published on Web 10/17/2001
Method for Adsorption of Blood Plasma Proteins
Of the many applications of soft lithography, one of the most successful has been the creation of surfaces patterned with biological ligands upon many different substrates such as glass, silicon, and polystyrene (PS).20 This is of particular interest due to its ability to create well-defined arrays of multiple molecules, which can be used as a basis for applications such as cell patterning, fabrication of biosensors, and combinatorial screening. The easiest and most successful examples of using soft lithography for creating surfaces of patterned proteins are those of µFN and µCP. In the case of µFN, the capillary based technique has been used to create multiple arrays of molecules upon a surface to check for interactions between antigens and antibodies,21,22 while others have demonstrated the ability to control cell growth on substrates with a view to tissue engineering.23 There are, however, some disadvantages to µFN. The most difficult part of the technique is maintaining the contact between the mold channels and the surface to create the capillary. This creates an extra step in the form of plasma etching. Also, the molds cannot be used with solvents or reactants that swell, dissolve, or degrade the channels. Furthermore, the technique is subject to reactant depletion, where the concentration of molecules in the solution is decreased at the end of the channel due to deposition on the sides of the plasma desorption mass spectrometry (PDMS) capilliary.24 In this work, the authors have foregone the use of µFN in favor of µCP. This is principally due to described disadvantages. However, using direct inking of the µCP stamp with protein solution has been avoided due to the fragile nature of biological molecules.21 Ideally, methods that maintain the functionality of the biological materials should be used. Previously, this has been achieved using self-assembled monolayers (SAMs) of alkanethiols with modified functional groups on gold (which is expensive and difficult) to create areas of protein resistance or adsorption.25,26 The aim of this work is therefore to demonstrate a simple, rapid, alternative method of patterning proteins to µFN, to avoid the disadvantages of direct inking of stamps with protein solutions, and to demonstrate how surfaces can be spatially defined into areas of protein resistance and adsorption without the use of SAMs and gold. µCP of a biocompatible polymer poly(lactic acid)-poly(ethylene glycol) (PLA-PEG) therefore was chosen as the alternative, to create highly defined areas of protein resistance upon a hydrophobic polymer substrate. PLAPEG has been demonstrated to inhibit cell proliferation, (16) Jackman, R. J.; Brittain, S. T.; Adams, A.; Prentiss, M. G.; Whitesides, G. M. Science 1998, 280, 2089-2091. (17) Rogers, J. A.; Meier, M.; Dodabalapur, A.; Laskowski, E. J.; Cappuzzo, M. A. Appl. Phys. Lett. 1999, 74, 3257-3259. (18) Jeon, N. L.; Choi, I. S.; Whitesides, G. M.; Kim, N. Y.; Laibinis, P. E.; Harada, Y.; Finnie, K. R.; Girolami, G. S.; Nuzzo, R. G. Appl. Phys. Lett. 1999, 75, 4201-4203. (19) Lackowski, W. M.; Ghosh, P.; Amirpour, M. L.; Pishko, M. V.; Crooks, R. M. Abstr. Pap. Am. Chem. Soc. 1999, 218, 162-COLL. (20) Bernard, A.; Delamarche, E.; Schmid, H.; Michel, B.; Bosshard, H. R.; Biebuyck, H. Langmuir 1998, 14, 2225-2229. (21) Bernard, A.; Michel, B.; Delamarche, E. Anal. Chem. 2001, 73, 8-12. (22) Patel, N.; Padera, R.; Sanders, G. H. W.; Cannizzaro, S. M.; Davies, M. C.; Langer, R.; Roberts, C. J.; Tendler, S. J. B.; Williams, P. M.; Shakesheff, K. M. FASEB J. 1998, 12, 1447-1454. (23) Dike, L. E.; Chen, C. S.; Mrksich, M.; Tien, J.; Whitesides, G. M.; Ingber, D. E. In Vitro Cell. Dev. Biol: Anim. 1999, 35, 441-448. (24) Delamarche, E.; Bernard, A.; Schmid, H.; Bietsch, A.; Michel, B.; Biebuyck, H. J. Am. Chem. Soc. 1998, 120, 500-508. (25) Lopez, G. P.; Albers, M. W.; Schreiber, S. L.; Carroll, R.; Peralta, E.; Whitesides, G. M. J. Am. Chem. Soc. 1993, 115, 5877-5878. (26) Lucke, A.; Tessmar, J.; Schnell, E.; Schmeer, G.; Gopferich, A. Biomaterials 2000, 21, 2361-2370.
Langmuir, Vol. 17, No. 23, 2001 7403
and its protein resistant properties have been utilized in this work.27,28 The rapid nature of the µCP technique (
