Using Self-Polymerized Dopamine to Modify the Antifouling Property

LETTER pubs.acs.org/Langmuir

Using Self-Polymerized Dopamine to Modify the Antifouling Property of Oligo(ethylene glycol) Self-Assembled Monolayers and Its Application in Cell Patterning Kang Sun,†,§ Lusheng Song,‡,§ Yunyan Xie,† Dingbin Liu,† Dong Wang,† Zhuo Wang,*,† Wanshun Ma,† Jinsong Zhu,‡ and Xingyu Jiang*,† †

Key Lab for Biological Effects of Nanomaterials and Nanosafety and ‡Laboratory for Nanomanufacture and Applications, National Center for Nanoscience and Technology, 11 Beiyitiao, ZhongGuanCun, Beijing, China 100190

bS Supporting Information ABSTRACT: We report a one-step, mild method to modify antifouling oligo(ethylene glycol)-terminated self-assembled monolayers. We demonstrate for the first time that self-polymerized dopamine, previously reported as an underwater adhesive, can be patterned on typical antifouling surfaces by microfluidic patterning or microcontact printing. The patterns can be applied in spatiotemporal cell patterning.

C

ell patterning has long been an important issue in fundamental cell biology, tissue engineering, drug screening, and so forth.1
5 In cell patterning, oligo(ethylene glycol) selfassembled monolayers (OEG SAMs) have been widely used to serve as antifouling surfaces that resist cell adhesion.6,7 Among current various methods to create patterns,8
12 a set of studies focus on making modifications on homogeneous antifouling surfaces. Examples include local damage of SAMs by generating active chemical species using electrochemical microelectrodes, selective desorption of SAM molecules by applying potentials in microfluidic channels, deep UV irradiation, and local desorption using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.13
19 The methods mentioned above can construct patterns of desired quality though; they rely on complex equipment, which is not always available in common biological laboratories. Herein, we develop a convenient and mild method for OEG SAM modification by employing dopamine. In basic solutions, selfpolymerized dopamine can automatically adhere to a wide range of substrates, behaving as an underwater adhesive.20
24 Despite the large number of types of surfaces that have been demonstrated to be coated with polymerized dopamine (polydopamine), OEG SAMs have not been examined. The examination of interaction between adhesive polydopamine and antifouling OEG SAM would be meaningful, as they represent opposite molecular behaviors. As an initial study, we showed that self-polydopamine could overcome the antifouling property of OEG SAMs, and we applied this phenomenon in patterning cells in combination with soft lithography techniques. We first studied if polydopamine film can form on OEG SAMs in the same condition as reported for other types of surfaces.20 We prepared OEG SAMs by immersing freshly evaporated gold substrates in an aqueous solution of hexa(ethylene glycol) r 2011 American Chemical Society

undecanethiol (EG6OH). We immersed OEG SAMs in a dopamine aqueous solution. We set the reaction condition as previously reported, that is, 2 mg/mL dopamine in 10 mM trisHCl buffer, pH 8.5. To prevent false positive adhesion arising from the precipitation of polydopamine on the surface, we placed the substrate carrying the SAM facing down. After 12 h, we rinsed the SAM in distilled water and dried it with compressed air. Scanning electron microscopy (SEM) of the SAM indicated the formation of nanoclusters (d < 150 nm) on the surface (Figure 1a inset). The nanoclusters grew in both number and size with the increase of immersion time, finally leading to the formation of a continuous film (see the Supporting Information). We characterized the fouled SAM using X-ray photoelectron spectroscopy (XPS). An obvious N1s peak appeared, confirming that nanoclusters were polydopamine (Figure 1a). To check if dopamine monomers can adsorb on OEG SAMs, we directly dissolved dopamine in distilled water (pH∼7) in 2 mg/mL and immersed OEG SAMs in the solution for 12 h. Dopamine did not polymerize in neutral distilled water as the solution remained transparent and colorless. XPS characterization showed no nitrogen signal (Figure 1b). This result suggests that the coating of polydopamine on OEG SAMs is not initiated by adsorption of monomers, but is likely to be caused by dopamine radicals or oligomers.25 We studied the dynamics of dopamine coating on OEG SAMs using surface plasmon resonance (SPR). We passed a dopamine solution in a microfluidic channel above OEG SAMs and recorded SPR signals for 1 h. For comparison, we monitored the adsorption of bovine serum albumin (BSA) on SAMs formed with undecanethiol Received: February 28, 2011 Revised: April 20, 2011 Published: April 26, 2011 5709

dx.doi.org/10.1021/la2012099 | Langmuir 2011, 27, 5709–5712

Langmuir

LETTER

Figure 1. (a) XPS characterization of OEG SAM immersed in self-polymerizing dopamine solution, confirming the adsorbent to be polydopamine. Note the N1s peak at 400 eV. Inset: SEM image of the surface with accumulated polymerized dopamine for 12 h. (b) XPS measurement of OEG SAM immersed in dopamine monomers for 12 h.

Figure 2. SPR characterization of adsorption of BSA on CH3 and OEG SAM, and coating of self-polymerizing dopamine on OEG SAM.

(abbreviated as “CH3”) or EG6OH (Figure 2). For BSA, the plateau in the process of the adsorption of BSA on CH3 indicates the saturation of adsorption. The amount of adsorption on EG6OH is about 2% of that on CH3. In contrast, dopamine coating shows a linear profile, which reflects a continuous adsorption, or accumulation of polydopamine on the surface. The result is in accordance with SEM results (see the Supporting Information). We further showed that dopamine can modify OEG SAMs when it was not in a process of polymerization. We stopped the reaction of a dopamine solution that had been polymerized for 8 h by adjusting pH to be acid (∼ 3), and immersed OEG SAMs in the solution for 5 h. Polydopamine was visible under SEM characterization, but it was largely reduced compared to the substrate immersed in polymerizing condition. Cell experiments showed that cells could adhere on such surfaces (see the Supporting Information). To check if polydopamine can directly transfer onto OEG SAMs besides accumulation from solution, we prepared a poly(dimethylsiloxane) (PDMS) stamp with patterns and coated it with polydopamine overnight. We performed microcontact

Figure 3. (a) Printed polydopamine on OEG SAM observed under optical microscopy, (b) magnified SEM image of printed polydopamine on OEG SAM. (c, d) Patterned NIH 3T3 fibroblast cells on polydopamine-printed OEG SAM and microfluidic-patterned polydopamine on OEG SAM.

printing (μCP) of polydopamine on OEG SAMs. Polydopamine can transfer onto the SAM with high fidelity (Figure 3a). These patterns of printed polydopamine remained stable after rinsing. We characterized the morphology of polydopamine on OEG SAMs after printing by SEM (Figure 3b). We found flattened polydopamine sheets on OEG SAMs, suggesting that polydopamine is a soft and sticky material, which can be reshaped under compression. The adhesion of polydopamine on antifouling surfaces made it suitable for cell patterning. We confined NIH 3T3 fibroblast cells to the polydopamine patterns (Figure 3c, d). The patterns were achieved by either microcontact printing (μCP) or microfluidic patterning (μFP). For μFP, we placed plasma-treated PDMS channels on an OEG SAM and filled them with a polymerizing 5710

dx.doi.org/10.1021/la2012099 |Langmuir 2011, 27, 5709–5712

Langmuir

Figure 4. Cathodic potential (
1.2 V, 30s) on the surface initiated the release of NIH 3T3 fibroblast cells on predefined areas by patterned polydopamine on OEG SAM. Cells were restricted on the patterns before voltage pulse (a) and migrated into prerestricted areas after 7 h (b).

dopamine solution. We left the solution overnight and washed the substrate three times in distilled water after peeling off PDMS. We seeded cells on these patterns and found that polydopaminepatterned OEG SAM was very effective in confining cells. Cell patterns could maintain as long as over 1 week (see the Supporting Information). The long maintenance of patterns further proves the stable modification of polydopamine on OEG SAMs as they can sustain cell movements, which is comparable to mixed SAMs employing alkanethiol to confine cells.26 We also show that this patterning technique can be adapted to other manipulations on surfaces, such as electrochemical desorption.4,27,28 After we cultured cells with polydopaminepatterned OEG SAM overnight, we applied a cathodic potential (
1.2 V, 30s) on the surface and released cells for free movements in real time. After 7 h, cells migrated out of the patterns (Figure 4). The stable binding of polydopamine on OEG SAMs provides a new material for studying its interaction with antifouling surfaces. Although the mechanism of antifouling property of OEG SAMs has been extensively studied,29
31 there have been reports suggesting that OEG SAMs do not repulse all molecules. Leckband and Sheth reported that poly(ethylene glycol) (PEG) had attractive interactions with proteins that were pressed into PEG polymers. Jiang and Cao found reduced resistance of OEG SAM when placed in blood plasma, and Chen et al. directly printed protein on OEG SAMs.32
34 Compared to blood plasma, dopamine solution is relatively simple in composition. The degree of polymerization can be tuned by adjusting the pH values of the solution. We believe that further investigation of interactions between polydopamine and OEG SAM would provide informative clues for studying strong adhesive ability of polydopamine. To show that polydopamine is unique even among synthetic polymers, we performed experiments employing polyaniline and polypyrrole as ink in μCP. Both polymers contain amine groups and can be modified on PDMS stamps in their polymerization state. We found that neither polymer could be well-transferred onto OEG SAM and neither could support cell adhesion (see the Supporting Information). Polydopamine has an especially stronger interaction with OEG SAM compared to the two polyamine in our test. Our finding also provides a new way to modify ethyleneglycol-grafted surfaces. Once surfaces are modified with homogeneous ethylene glycol groups (such as OEG SAM), it is difficult to make further modification that can attach molecules firmly on the surface. Usually researchers have to destroy ethylene glycol groups and create sites for reaction or adsorption of molecules of interest.13
19 Alternatively, chemical synthesis is needed to add functional groups onto ethylene glycol molecules.35
37 In our method, altering inactive OEG SAMs is achieved by simple

LETTER

printing or immersion. The application of this method in patterning cells has several advantages. First, all reactions take place in mild conditions, which is convenient in common laboratories. Second, the materials are easy to prepare. And finally, the procedure is rather straightforward; substrates are patterned with polydopamine by one step of μCP or μFP. Thus, this method is a robust one to directly modify polydopamine on ethylene-glycol-terminated surfaces for controlling cell behavior, without multiple steps, severe reaction conditions, or extra equipment. In conclusion, we found that polydopamine could overcome the antifouling property of OEG SAMs. We realized patterning polydopamine on OEG SAM surfaces by μCP or μFP. Cells can be easily patterned using polydopamine-patterned surfaces. The combination of powerful polydopamine and antifouling surfaces provides a convenient, mild, and easily accessible method for controlling cells both spatially and temporally.

’ ASSOCIATED CONTENT

bS

Supporting Information. Figures SI1
SI4; method for microcontact printing of polyaniline and polypyrrole. This material is available free of charge via the Internet at http:// pubs.acs.org.

’ AUTHOR INFORMATION Corresponding Author

*E-mail: [email protected] (X.J.); [email protected] (Z.W.). Fax: þ86-10-82545631. Telephone: þ86-10-82545558. Author Contributions §

These authors contributed equally to this work.

’ ACKNOWLEDGMENT We thank the Human Frontier Science Program, the National Science Foundation of China (90813032 , 50902025, and 20890020), the Ministry of Science and Technology (2007CB714502, 2009CB930001, and 2011CB933201), and the Chinese Academy of Sciences (KJCX2-YW-M15) for funding. ’ REFERENCES (1) Mahmud, G.; Campbell, C. J.; Bishop, K. J. M.; Komarova, Y. A.; Chaga, O.; Soh, S.; Huda, S.; Kandere-Grzybowska, K.; Grzybowski, B. A. Nat. Phys. 2009, 5, 606–612. (2) Nakanishi, J.; Takarada, T.; Yamaguchi, K.; Maeda, M. Anal. Sci. 2008, 24, 67–72. (3) Tuleuova, N.; Lee, J. Y.; Lee, J.; Ramanculov, E.; Zern, M. A.; Revzin, A. Biomaterials 2010, 31, 9221–9231. (4) Jiang, X. Y.; Ferrigno, R.; Mrksich, M.; Whitesides, G. M. J. Am. Chem. Soc. 2003, 125, 2366–2367. (5) Hou, S. Y.; Burton, E. A.; Wu, R. L.; Luk, Y. Y.; Ren, D. C. Chem. Commun. 2009, 10, 1207–1209. (6) Whitesides, G. M.; Ostuni, E.; Takayama, S.; Jiang, X. Y.; Ingber, D. E. Annu. Rev. Biomed. Eng. 2001, 3, 335–373. (7) Prime, K. L.; Whitesides, G. M. J. Am. Chem. Soc. 1993, 115, 10714–10721. (8) Suh, K. Y.; Park, M. C.; Kim, P. Adv. Funct. Mater. 2009, 19, 2699–2712. (9) Falconnet, D.; Csucs, G.; Grandin, H. M.; Textor, M. Biomaterials 2006, 27, 3044–3063. (10) Ostuni, E.; Kane, R.; Chen, C. S.; Ingber, D. E.; Whitesides, G. M. Langmuir 2000, 16, 7811–7819. 5711

dx.doi.org/10.1021/la2012099 |Langmuir 2011, 27, 5709–5712

Langmuir

LETTER

(11) Singh, A. V.; Lenardi, C.; Gailite, L.; Gianfelice, A.; Milani, P. J. Micromech. Microeng. 2009, 19, 115028. (12) Yan, H.; Zhao, Y. T.; Qiu, C.; Wu, H. K. Sens. Actuators, B 2008, 132, 20–25. (13) Ducker, R. E.; Janusz, S.; Sun, S. Q.; Leggett, G. J. J. Am. Chem. Soc. 2007, 129, 14842–14843. (14) Kim, Y. K.; Ryoo, S. R.; Kwack, S. J.; Min, D. H. Angew. Chem., Int. Ed. 2009, 48, 3507–3511. (15) Zhao, C.; Witte, I.; Wittstock, G. Angew. Chem., Int. Ed. 2006, 45, 5469–5471. (16) Li, Y.; Yuan, B.; Ji, H.; Han, D.; Chen, S. Q.; Tian, F.; Jiang, X. Y. Angew. Chem., Int. Ed. 2007, 46, 1094–1096. (17) Kaji, H.; Kawashima, T.; Nishizawa, M. Langmuir 2006, 22, 10784–10787. (18) Azioune, A.; Storch, M.; Bornens, M.; Thery, M.; Piel, M. Lab Chip 2009, 9, 1640–1642. (19) Suh, K. Y.; Seong, J.; Khademhosseini, A.; Laibinis, P. E.; Langer, R. Biomaterials 2004, 25, 557–563. (20) Lee, H.; Dellatore, S. M.; Miller, W. M.; Messersmith, P. B. Science 2007, 318, 426–430. (21) Ku, S. H.; Lee, J. S.; Park, C. B. Langmuir 2010, 26, 15104–15108. (22) Kang, S. M.; You, I.; Cho, W. K.; Shon, H. K.; Lee, T. G.; Choi, I. S.; Karp, J. M.; Lee, H. Angew. Chem., Int. Ed. 2010, 49, 9401–9404. (23) Yang, S. H.; Kang, S. M.; Lee, K.-B.; Chung, T. D.; Lee, H.; Choi, I. S. J. Am. Chem. Soc. 2011, 133, 2795–2797. (24) Kang, S. M.; Park, S.; Kim, D.; Park, S. Y.; Ruoff, R. S.; Lee, H. Adv. Funct. Mater. 2011, 21, 108–112. (25) Bernsmann, F.; Ponche, A.; Ringwald, C.; Hemmerle, J.; Raya, J.; Bechinger, B.; Voegel, J. C.; Schaaf, P.; Ball, V. J. Phys. Chem. C 2009, 113, 8234–8242. (26) Jiang, X. Y.; Bruzewicz, D. A.; Thant, M. M.; Whitesides, G. M. Anal. Chem. 2004, 76, 6116–6121. (27) Yousaf, M. N.; Houseman, B. T.; Mrksich, M. Proc. Natl. Acad. Sci. U.S.A. 2001, 98, 5992–5996. (28) Yeo, W. S.; Yousaf, M. N.; Mrksich, M. J. Am. Chem. Soc. 2003, 125, 14994–14995. (29) Ostuni, E.; Chapman, R. G.; Holmlin, R. E.; Takayama, S.; Whitesides, G. M. Langmuir 2001, 17, 5605–5620. (30) Herrwerth, S.; Eck, W.; Reinhardt, S.; Grunze, M. J. Am. Chem. Soc. 2003, 125, 9359–9366. (31) Chen, S. F.; Zheng, J.; Li, L. Y.; Jiang, S. Y. J. Am. Chem. Soc. 2005, 127, 14473–14478. (32) Sheth, S. R.; Leckband, D. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 8399–8404. (33) Jiang, S. Y.; Cao, Z. Q. Adv. Mater. 2010, 22, 920–932. (34) Tan, J. L.; Tien, J.; Chen, C. S. Langmuir 2002, 18, 519–523. (35) Liu, D. B.; Xie, Y. Y.; Shao, H. W.; Jiang, X. Y. Angew. Chem., Int. Ed. 2009, 48, 4406–4408. (36) Yeo, W. S.; Mrksich, M. Langmuir 2006, 22, 10816–10820. (37) Yuan, S. J.; Wan, D.; Liang, B.; Pehkonen, S. O.; Ting, Y. P.; Neoh, K. G.; Kang, E. T. Langmuir 2011, 27, 2761–2774.

5712

dx.doi.org/10.1021/la2012099 |Langmuir 2011, 27, 5709–5712