Antifouling Surface Coating Using Droplet-Based SI-ARGET ATRP of

May 22, 2019 - Daewha Hong* ... however, their air-tightness may cause inconvenience to users. ... Hyeongeun Kang, Wonwoo Jeong, and Daewha Hong*...
0 downloads 0 Views 2MB Size
Download PDF
Article Cite This: Langmuir 2019, 35, 7744−7750

pubs.acs.org/Langmuir

Antifouling Surface Coating Using Droplet-Based SI-ARGET ATRP of Carboxybetaine under Open-Air Conditions Hyeongeun Kang, Wonwoo Jeong, and Daewha Hong* Department of Chemistry, Chemistry Institute of Functional Materials, Pusan National University, Busan 46241, South Korea

Downloaded via UNIV OF SOUTHERN INDIANA on July 29, 2019 at 12:11:50 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

S Supporting Information *

ABSTRACT: The formation of a dense zwitterionic brush through surface-initiated atom transfer radical polymerization (SI-ATRP) is a typical graft-from approach used to achieve antifouling surfaces with high fidelity; however, their air-tightness may cause inconvenience to users. In this context, activator regenerated by electron transfer (ARGET) ATRP is emerging as an alternative surface-coating tool because limited amount of air is allowed to form a dense polymer brush. However, the degree of air tolerance that can ensure a thick polymer brush has not been clearly defined, limiting its practical usage under ambient-air conditions. In this study, we investigated the SI-ARGET ATRP of carboxybetaine (CB) by changing the air conditions, along with the air-related parameters, such as the concentration of the reducing agent, the volume of the polymerization solution (PS), or the solvent composition, and correlated their effects with the poly(CB) thickness. Based on the optimized reaction conditions, a poly(CB) brush with reliable thickness was feasibly formed even under open-air conditions without a degassing step. In addition, a microliter droplet (∼100 μL) of PS was sufficient to proceed with the SI-ARGET ATRP for the covering of a poly(CB) brush on the surface area of interest. By applying an optimized SI-ARGET ATRP of CB, antifouling was feasibly achieved in the surface region of interest using an array to form a large surface area under fully exposed air conditions. In other words, optimized SI-ARGET ATRP enabled the formation of a thick poly(CB) brush on the surfaces of various dimensions under open-air conditions.

■

INTRODUCTION The grafting of a polymer brush with hydrophilic materials on a substrate is an effective and classical method for repressing the nonspecific binding of proteins or cells, and is an important issue in the development of biosensors, medical devices, and immunodiagnostic assays.1,2 Among the various coating methods, the formation of a zwitterionic polymer brush composed of phosphorylcholine, sulfobetaine, or carboxybetaine (CB) using surface-initiated atom transfer radical polymerization (SI-ATRP) is an attractive approach to obtain an excellent antifouling performance because the formation of a strong hydration layer around a dense zwitterionic brush can inhibit the access and subsequent adsorption of a bioentity with high fidelity.3,4 However, the successful formation of a polymer brush using classical SI-ATRP is only valid under airtight conditions, which requires a cumbersome degassing step with specialized equipment.5 This fact itself brings about inconvenience to users and narrows the scope of its applicability to small and flat surfaces, which allow deoxygenation to be managed under a isolated system. The recent development of activator regenerated by electron transfer (ARGET) ATRP has alleviated the air-tight issues of conventional ATRP.6−8 In this case, excessive amount of reducing agent, represented by ascorbic acid, continuously regenerates an active Cu(I) species from an inactive Cu(II) species and allows polymerization under limited amount of air. © 2019 American Chemical Society

This strategy has also been transformed in the version of surface-initiated polymerization to coated surfaces through the formation of a polymer brush, and their degassing-free condition itself enables the coating of large substrates, as well as the interfaces among nanoparticles, proteins, or living cells for their own application.9−12 However, SI-ARGET ATRP is still regarded as an “air-tolerant” coating method, conceding that it is not fully compatible with an infinite amount of air. In this sense, SI-ARGET ATRP is mainly conducted under a closed system in which the amount of air is predetermined by the headspace volume within a sealed jar. However, the detailed amount of allowed air during SIARGET ATRP has not been fully described, and it is therefore virtually impossible to compare or estimate the degree of air tolerance to ensure the application of a polymer brush using SI-ARGET ATRP. We reasoned that a strong correlation exists between a zwitterionic polymer brush formation on a surface versus the amount of air, and the degree of air tolerance that can allow a reliable polymer brush to be applied can be enhanced, for example, under open-air conditions, based on optimized polymerization conditions. Although the enhanced air Received: March 20, 2019 Revised: May 15, 2019 Published: May 22, 2019 7744

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Article

Langmuir

ascorbic acid, 58 μM CuBr2, and 350 μM bpy. After reacting for 3 h, the surface was rinsed with phosphate-buffered saline (PBS) and water, and dried with a stream of air. For a large surface area, an initiator-functionalized gold wafer (50 cm2) was immersed in PS whose component concentrations were identical, yet differed in the total PS volume (10 mL). The conditions for a large surface area are as follows: 625 mM CB, 2.94 mM ascorbic acid, 58 μM CuBr2, and 350 μM bpy; reacted for 3 h. The SI-ARGET ATRP with the MeOH/ water co-solvent was performed with a slight modification in that the stock solution of CuBr2/bpy was dissolved in MeOH instead of water, and the CB monomer was dissolved in an appropriate ratio of MeOH and water. Others were performed identically. SI-ARGET ATRP Using Droplet-Based Method. An aqueousbased PS solution was prepared in advance, in a manner identical to that described in the previous section. A silicon isolator was compressed onto the initiator presenting surfaces, forming a roundshaped well (diameter of 9 mm, depth of 1.7 mm). An aliquot of the as-prepared PS (200 or 100 μL) was then applied to the well for 3 h. After polymerization, the solution was removed from the well to terminate the SI-ARGET ATRP. The silicon isolator was then detached from the surfaces, followed by rinsing with PBS and water, and dried with a stream of air. To prepare the patterned poly(CB), an identical process was performed on an initiator-patterned gold surface (refer to the Supporting Information describing the microcontact printing (μCP)). Typical SI-ATRP on Gold Surfaces. Typical SI-ATRP of CB was performed based on a previous report.19 In contrast to the air-tolerant SI-ARGET ATRP, all of the experiments were performed under airtight conditions. For this, MeOH and water were nitrogen-purged for 2 h before use. In a glass tube, 7.4 mg of CuBr, 48.2 mg of bpy, and 500 mg of CB were added and degassed under vacuum and purged with nitrogen. To this solid mixture, 2 mL of water and 1.33 mL of MeOH were added. The resulting solution was stirred for 30 min to complete the dissolution of the solid mixture. The total volume of the solution was measured to be ∼3.5 mL. The initiator-coated gold surfaces were placed in another glass tube and then degassed under vacuum and purged with nitrogen. The solution mixture was then transferred to the glass tube containing gold surfaces to initiate typical SI-ATRP for 3 h. After the reaction, the gold surfaces were rinsed with PBS and water and dried with a stream of air. Typical SI-ATRP of CB under air was identically performed without any degassing process. Polymerization under air was performed under a closed system as well as an open system.

tolerance of SI-ARGET ATRP will be beneficial to the coating of real-life substrates with large surface areas, it may also be inversely advantageous for miniaturization in which nonfouling is highly required, including biosensors, diagnostic devices, or the simple pattern generation of proteins.13−15 Because the formation of a polymer brush within a small surface area is sufficient to fulfill the aforementioned application, a costeffective SI-ARGET ATRP can be achieved by reducing the reaction volume itself, which contains a catalyst, a ligand, and even synthesized monomers. However, reducing the volume of polymerization solution (PS) not only decreases the absolute amount of monomers used to build up a polymer brush, it also shortens the solution layer. These circumstances are more susceptible to oxygen diffusion because it is proportional to the square of the solution layer thickness.16 Taken together, enhancing the air tolerance of the SI-ARGET ATRP is of particular importance for the development of a practical and cost-effective coating by applying a minimum PS volume that can cover the surfaces. In other words, simply introducing a thin layer of PS can enable the formation of a thick polymer brush in open air. Controlled radical polymerization under an open-air system has recently been gradually illustrated through a limited scope of materials, although not for zwitterion materials, which can lead to antifouling surfaces.17,18 In this work, we report the SI-ARGET ATRP of CB that can form a thick poly(CB) brush under a completely open system, and demonstrate that such a formation is possible by introducing a 100 μL droplet of PS on a small surface area (0.3 cm2).

■

EXPERIMENTAL SECTION

SI-ARGET ATRP on Gold Surfaces. The gold surfaces (2 cm2) were rinsed with water and ethanol and then treated with plasma for 5 min before the formation of a self-assembled monolayer (SAM). The SAM of the initiator was prepared by immersing the rinsed gold surfaces in an ethanolic solution of the initiator (0.1 mM) for 12 h. For the mixed SAMs, the gold surfaces were immersed into an ethanolic solution of 0.1 mM thiols (total concentration) containing 11-mercapto-1-undecanol and an initiator. The surfaces were then rinsed with ethanol and dried with a stream of air. The initiatorcoated surfaces were then used for the subsequent SI-ARGET ATRP. Throughout SI-ARGET ATRP, no deoxygenation step was applied. First, the SI-ARGET ATRP was conducted in a pure water system as follows. Owing to the extremely low amount of catalyst/ligand (∼ppm) applied in this system, an aqueous stock solution containing 6.92 mg CuBr2 and 29.02 mg bpy in 50 mL water was prepared in advance. In a glass vial, 500 mg CB monomer was dissolved in 2.87 mL water and an as-prepared 0.33 mL CuBr2/bpy stock solution was added. When investigating the effect of monomer concentration on the SI-ARGET ATRP, different amounts of CB were dissolved in this step. After stirring for 20 min, the solution mixture was transferred to a Petri dish (reaction chamber: diameter = 3.5 cm, depth = 1.75 cm) containing the initiator-coated surfaces. Subsequently, 0.13 mL water dissolving different amounts of ascorbic acid (0.4−18.2 mg) was added to begin SI-ARGET ATRP. Overall, the total volume of PS was measured to be ∼3.5 mL. The total volume of the PS was higher than that of the solvent due to the excessive amount of monomer. When a small amount of CB monomer was dissolved, additional water was added, resulting in a PS volume of 3.5 mL. To obtain a closed system containing limited amount of air, the reaction chamber was directly sealed after the addition of ascorbic acid. In our closed system, the headspace volume was measured to be 12.5 mL, which corresponds to ∼0.1 mmol of O2 under ambient condition. To obtain an open system, the chamber was not sealed to face infinite amount of external air. If the amounts of applied CB monomer and ascorbic acid are 500 and 1.81 mg, respectively, the overall the SI-ARGET ATRP conditions will be as follows, by calculation: 625 mM CB, 2.94 mM

■

RESULTS AND DISCUSSION Figure 1 shows a schematic of our study estimating the air tolerance of SI-AREGT ATRP in terms of a polymer brush formation. We investigate the poly(CB) thickness by changing the SI-ARGET ATRP conditions, including the amount of air, concentration of ascorbic acid, volume of PS, or solvent composition, which have been loosely studied but are important parameters for the surface coating. In addition, we correlated the poly(CB) thickness by varying the concentration of the monomer and the density of the initiator on the surfaces, which have been regarded as important parameters for controlled radical polymerization.20,21 Among them, we start with the air effect on the SI-ARGET ATRP because the expression of “air tolerance” or “oxygen tolerance” has not been consistently defined across the related literature.22 For example, a certain system was claimed to be air tolerant if a reaction can be achieved within a sealed jar containing a predetermined amount of air but only after the solvents are degassed in advance. Others have emphasized the degassingfree process itself but have not discovered whether a polymer brush formation is still available when the sealed jar is open, which faces a virtually infinite amount of air. In our system, we clearly defined the “air conditions” as follows: no need for a 7745

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Article

Langmuir

further supported by measuring the water contact angle (WCA): overall, the formation of the poly(CB) brush resulted in a hydrophilic surface. In our system, the headspace volume of the reaction vessel was ∼12.5 mL, which corresponds to ∼0.1 mmol of O2.6 Our new finding is that the SI-ARGET ATRP of CB under an open system is highly dependent on the water composition of the solvent. Entries 1 and 2, which contain 40/20% (v/v) of MeOH, respectively, failed to form poly(CB), whereas entry 3 still formed poly(CB), indicating that the SI-ARGET ATRP open to air is still valid under a pure aqueous system. This was further confirmed by direct comparison between the closed system and open system under various reaction conditions. With a pure aqueous solvent, the closed system and the open system showed similar tendency in their poly(CB) thickness, regardless of air conditions (Figure S1). We assumed that the enhanced air tolerance of the SI-ARGET ATRP under a pure aqueous solvent can be attributed to the combinatory effects of oxygen solubility in the solvents and the kinetics of the polymerization. Preliminary studies reported that the oxygen solubility of water under ambient conditions is 5 times lower than that of MeOH.22 In other words, a pure aqueous system can be less vulnerable against degassing-free conditions than a water/ MeOH co-solvent system. Additionally, a fast polymerization rate under aqueous condition can be a minor factor to overcome oxygen sensitivity before unwanted radical termination by dissolved oxygen. Based on our comparative studies, the initial rate of film growth was fastest for pure aqueous solvent, yet became slower as the water content of the solvent decreased. Their difference became even polarized under open system, resulting in only pure aqueous solvent allowing a thick poly(CB) brush (Figure S2). As a control, we also conducted typical SI-ATRP of CB under nitrogen (air-tight condition) as well as air conditions in a closed system and an open system. As expected, poly(CB) was formed via SI-ATRP only under air-tight conditions (10 nm) yet failed to form poly(CB) under both closed and open systems where air existed (Table S1). Based on our comparative study, we focused on the reliable operation of SI-ARGET ATRP of CB under an open system (entry 3) and further investigated the film formation by adjusting other experimental parameters, including the ascorbic acid concentration, PS volume, monomer concentration, and amount of initiator on the surfaces. All of the SI-ARGET ATRP experiments from now were conducted under pure aqueous solvent and open-air system. First, we varied the initial concentration of ascorbic acid, which acts as a chemical reservoir for purging dissolved oxygen within the PS. As the concentration of ascorbic acid increases, the thickness of a poly(CB) brush increases to up to 38 nm but decreases when the concentration exceeds 2.94 mM (Figure 2a). Based on our experiments, the excess amount of ascorbic

Figure 1. Schematic representation investigating the air tolerance of SI-ARGET ATRP. The film thickness on the surface was measured by changing the SI-ARGET ATRP parameters, including the air conditions, solvent, ascorbic acid concentration, PS volume, monomer concentration, and initiator density.

degassing step during the entire process of SI-ARGET ATRP, including the preparation of all of the reagents and reaction vessels. To demonstrate the “enhanced” air tolerance of SIARGET ATRP, the polymerization was conducted under an open-air system by taking off the lid. Then, in the view of the polymer brush formation, we claimed that the given SIARGET ATRP is air-tolerant if it can result in an acceptable polymer brush thicker than 10 nm because such a thickness has been reported to exhibit an effective protein resistance.23 For a direct comparison to evaluate the air tolerance, the SI-ARGET ATRP was attempted under a sealed or open jar, which is referred to as a closed or an open system, respectively (Table 1). In our study, the closed system is a positive control because a limited amount of air is known to allow SI-ARGET ATRP. Initially, a solvent composed of water and MeOH was selected to guarantee the high solubility of ascorbic acid and the efficient reducing capability in generating an active Cu(I) species from an inactive Cu(II) species.9,24 Because the surface tension of MeOH was lower than that of water, the initial wetting of PS on the initiator-coated gold surfaces was highly dependent on the ratio of MeOH and water: high contents of water in PS resulted in low wetting. Although the initial wetting of PS was significantly different depending on the composition of MeOH and water, conditions corresponding to all entries (1−3) in the table ensured the formation of a polymer brush (≥15 nm) under our positive control (a closed system), thereby confirming that the SI-ARGET ATRP is typically capable under a limited amount of air. These were Table 1. SI-ARGET ATRP of CB in the Presence of Aira

Thickness (nm)

WCA of coated film (deg)

Entry

Initial wetting of PS (deg)

Solvent

Closed

Open

Closed

Open

1 2 3

35 ± 2.8 50 ± 1.7 64 ± 2.3

MeOH/water = 6:4 (v/v) MeOH/water = 8:2 (v/v) water

15 ± 0.4 17 ± 5.2 23 ± 5.4

1 ± 0.2 5 ± 2.8 27 ± 3.3

7.4 ± 1.6 7.3 ± 2.2 7.1 ± 1.0

37.4 ± 2.8 28.4 ± 5.7 5.9 ± 1.0

a Other polymerization conditions were fixed (625 mM CB, 5.88 mM ascorbic acid, 58 μM CuBr2, and 350 μM bpy; reacted for 3 h). The reaction was conducted in a Petri dish (diameter of 3.5 cm, depth of 1.75 cm), and the total volume of the polymerization solution (PS) was 3.5 mL. In this case, the headspace volume of the reaction was ∼12.5 mL, which corresponds to ∼0.1 mmol of O2.

7746

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Article

Langmuir

Figure 2. Film thickness of poly(CB) and its antifouling capability. The SI-ARGET ATRP conditions were fixed (625 mM monomer, 58 μM CuBr2, and 350 μM bpy; reacted for 3 h). (a) Measurement of poly(CB) thickness by changing the ascorbic acid concentration and volume of the polymerization condition. (b) Relative fibrinogen fouling based on ELISA. In this case, the ascorbic acid concentration was fixed (2.94 mM) with a different volume of PS. Each fouling was compared with an uncoated gold surface (control).

Figure 3. Effect of monomer concentration and initiator amount for growing poly(CB) brush. The other parameters for SI-ARGET ATRP were fixed (58 μM CuBr2 and 350 μM bpy; reacted for 3 h). (a) Film thickness of poly(CB) formed by various concentrations of monomer and ascorbic acid. (b) Film thickness of poly(CB) formed by changing the amount of initiator on the surfaces. In this case, mixed SAMs of 11-mercapto-1undecanol and the initiator, differing in composition, were coated on the gold surfaces, and 625 mM CB and 2.94 mM ascorbic acid were used.

an ellipsometric analysis (Figure 2a). Overall, the poly(CB) thickness was decreased compared with the submerging system but still resulted in a thick and reliable poly(CB) layer close to 20 nm under an optimized ascorbic acid concentration (5.88 mM). During the droplet-based SI-ARGET ATRP, the droplets of PS did not dry under ambient condition or humid environments, and their difference did not significantly affect poly(CB) formation. In addition, the concentration of ascorbic acid that can result in the thickest poly(CB) slightly increased as the volume of polymerization decreased. In this case, the diffusion and accessibility of the dissolved oxygen can be the dominant factors owing to the short solution layer between the initiator-functionalized surfaces and the external air/solution interfaces: for an immerging system, the solution layer is measured to be ∼4.5 mm, whereas for a 100 μL droplet system, it is ∼1.5 mm. Within a small microliter scale, it can be concluded that an oversupply of ascorbic acid has a positive effect on the formation of poly(CB). Within this dimension, an oversupply of ascorbic acid can place more weight on the purging of oxygen that steadily permeates into the reaction system, rather than causing a fast uncontrolled polymerization, which was observed in an immerging system. Overall, our comparative study indicates that even a 100 μL volume of PS is sufficient to form a poly(CB) brush on a small surface area (0.3 cm2) under open-air conditions without any instrumentalbased deoxygenation step. Then, protein adsorption on the poly(CB)-coated region was estimated using enzyme-linked

acid did not ensure a thick poly(CB) brush, which is consistent with previous studies: an immoderate amount of ascorbic acid instead interferes with the equilibrium during the early stage of SI-ARGET ATRP, causing rapid and uncontrolled polymerization.9,16 Next, we attempted to reduce the volume of PS to the droplet scale without changing the concentration of each component. Officially, SI-ARGET ATRP is described as a costeffective coating method because it requires an extremely low concentration of catalyst (∼ppm) compared to conventional SI-ATRP. However, a large amount of synthesized monomer is often required with a high concentration, and for a complete immersion of a small surface area, the solution volume is frequently overspent. Therefore, it is desirable to reduce the absolute volume of PS to the minimum level for the actual development of a cost-effective coating method. Reducing the volume, however, results in a thin solution layer between the initiator-functionalized surfaces and solution/air interfaces whose circumstances can be more susceptible to oxygen diffusion owing to their short distance. We anticipate that our enhanced air tolerance will remain functional within a microliter droplet scale. In this sense, the overall volume of PS was reduced from 3500 to 200 μL or 100 μL, which did not immerse the surface but covered it in a droplet form whose surface contact area proceeded with SI-ARGET ATRP for the brush formation. Moreover, a droplet was fully exposed to open-air conditions. After our droplet-based SI-ARGET ATRP was applied, the thickness of poly(CB) was measured through 7747

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Article

Langmuir

step.15 In this case, 100 μL of PS was identically applied to initiator-patterned gold surfaces formed using microcontact printing (μCP).27 The resultant surface was then immersed in a PBS solution containing albumin−FITC (0.1 mg/mL in PBS) for 1 h to induce protein adsorption on the non-poly(CB) brush region. After washing with PBS, the entire surface was analyzed using a fluorescence microscope (Figure 4b). As expected, a remarkable contrast in a grid form was observed, indicating that the poly(CB) brush repressed the nonspecific binding of the protein, and, therefore, the adhesion occurred on a square (grid) region where a poly(CB) brush is absent. Because our developed SI-ARGET ATRP is capable under open-air conditions with a small reaction volume, it can be flexibly applied to the surface region of interest. In other words, a regioselective poly(CB) brush in an array form can be established by simply dropping PS on a certain surface area. In this case, a highly ordered microwell was provided by a silicon isolator (diameter of 9 mm, depth of 1.7 mm) on the initiatorfunctionalized surfaces, and the droplet-based SI-ARGET ATRP of CB was identically performed on each well by applying a 100 μL volume of PS. The dark circle region was feasibly observed in appearance, indicating the regioselective poly(CB) brush formation (Figure 5a), and the WCA on each

immunosorbent assays (ELISAs). As a model protein, fibrinogen was selected because it is known to be sticky on a broad range of surfaces.25 In this case, fibrinogen adsorption on each poly(CB) brush formed under a different solution volume, including the immerging (3500 μL) and droplet (200 and 100 μL) systems, was compared (625 mM monomer, 2.94 mM ascorbic acid, 58 μM CuBr2, and 350 μM bpy; reacted for 3 h). As a result, all of the poly(CB)-coated surfaces resulted in an antifouling performance with high fidelity: The relative fouling of all of the poly(CB)-coated surfaces whose formation differs in the volume of PS was measured to be ∼5% with respect to the uncoated surfaces (Figure 2b). Finally, we investigated the effects of monomer concentration and the amount of initiator, which are known to be critical for the formation of polymer brushes of different thickness.20,21 First, the thickness of the poly(CB) film was investigated by varying the CB concentration as well as the ascorbic acid concentration (Figure 3a). The experimental results indicate that a certain amount of CB concentration is inevitably required to enable the formation of a thick poly(CB) brush. Concentrations below 300 mM CB were not effective for the formation of a thick poly(CB) brush regardless of the ascorbic acid concentration, while higher concentrations were successful in forming a thick poly(CB) (25−40 nm) and were highly dependent on the ascorbic acid concentration. In addition, we formed mixed SAMs composed of different ratios of 11-mercapto-1-undecanol and initiator on the gold surfaces. As expected, an increase in the initiator amount on the surfaces resulted in a thicker poly(CB) brush, which is consistent with the typical SI-ATRP.21 Because a small volume of PS resulted in a poly(CB) brush with a reliable antifouling performance, we used the droplet method to generate protein patterns on the surfaces (Figure 4a). Protein pattern formation on the surfaces provides a useful tool for developing biosensors, drug screening platforms, and diagnostic devices.26 Previously, the regioselective immobilization of an initiator followed by the typical SI-ATRP of an antifouling polymer was attempted to form a protein pattern on the surfaces; however, this required a large amount of PS and involved a cumbersome degassing

Figure 5. Regioselective formation of poly(CB). (a) Formation of poly(CB) in an array form. Dark circle patterns observed by optical camera indicate poly(CB)-coated region. (b) Water-capturing property of poly(CB). Stream of water on tilted surfaces resulted in site-selective hydration layer on poly(CB)-coated region. The scale bar is 1 cm.

surface area changed from ∼74° (initiator-coated surface) to ∼6°, providing hydrophilicity on the coated region.28 When a stream of water was randomly introduced onto the tilted surfaces, the water droplets selectively remained on the poly(CB)-coated region, indicating that an effective hydration layer was formed in this region (Figure 5b). A similar coating procedure was conducted on a large surface area (10 cm in diameter) by applying a minimum volume of PS (10 mL) that can cover a 50 cm2 surface. After polymerization, the initiator-functionalized surface region that was in contact with the PS turned dark, indicating the formation of a poly(CB) brush (Figure 6a). The thickness of the poly(CB) on the entire surface was measured to be 23 nm with a small standard deviation of 2.8 nm, indicating that the SI-ARGET ATRP of CB resulted in a uniform and reliable poly(CB) film across the large surface area. A strong hydration layer was consistently observed over the entire surface when a stream of water was introduced onto the poly(CB)-coated

Figure 4. Pattern generation of protein using droplet-based SIARGET ATRP of CB. (a) Procedure for pattern generation of poly(CB) and albumin−fluorescein isothiocyanate (FITC) on gold surfaces. A droplet of 100 μL PS solution (625 mM CB, 2.94 mM ascorbic acid, 58 μM CuBr2, and 350 μM bpy; reacted for 3 h) was applied to grow the patterned poly(CB) brush. After forming the patterned poly(CB) brush, albumin−FITC (0.1 mg/mL in PBS) solution was introduced onto the surface for 1 h to induce protein patterns. (b) Fluorescence microscopy image of patterned albumin− FITC. The scale bar is 100 μm. 7748

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Langmuir

■

Article

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Daewha Hong: 0000-0002-2339-2188 Notes

The authors declare no competing financial interest.

■ ■

ACKNOWLEDGMENTS This research was supported by the National Research Foundation of Korea (NRF-2018R1C1B6005624).

Figure 6. Optical image of large surfaces coated with poly(CB) brush. (a) The dark circle represents the formation of poly(CB). (b) The hydration layer was selectively formed on the poly(CB)-coated region. The scale bar is 2 cm. (c) Relative fibrinogen fouling between uncoated surface and poly(CB)-coated surface. The fibrinogen fouling was measured by ELISA.

wafer (Figure 6b). The antifouling capability of a poly(CB)coated film on a large surface area was also evaluated by ELISA (Figure 6c). Fibrinogen adsorption of the poly(CB)-coated surface was measured to be ∼3% compared to that of the uncoated one, indicating that the antifouling performance on large surfaces was consistently achieved with the use of minimum PS volume.

■

CONCLUSIONS In summary, we developed a SI-ARGET ATRP of CB that can form a poly(CB) brush under open-air conditions with a minimum reaction volume. Considering that the practical surface coating achieves low fouling, we optimized the SIARGET ATRP to meet convenient and inexpensive conditions under open-air conditions. The developed coating method was compatible with various surface dimensions from a microliter array to a large surface area, forming a reliable poly(CB) brush. Because our coating system is fully compatible with an aqueous solution and open air, we expected it to be feasibly combined with an external device, such as a microarray or fluidics for their use in immunodiagnostic or flow chemistry, respectively.29,30 In addition, we expected that its aqueous-based conditions with a parts per million level of a catalyst can be beneficial to the engineering of living cell surfaces in the development of tissue therapy, cell-based sensors, and implantation.31

■

REFERENCES

(1) Liu, L.; Li, W.; Liu, Q. Recent Development of Antifouling Polymers: Structure, Evaluation, and Biomedical Applications in Nano/Micro-Structures. Wiley Interdiscip. Rev.: Nanomed. Nanobiotechnol. 2014, 6, 599−614. (2) Banerjee, I.; Pangule, R. C.; Kane, R. S. Antifouling Coatings: Recent Developments in the Design of Surfaces That Prevent Fouling by Proteins, Bacteria, and Marine Organisms. Adv. Mater. 2011, 23, 690−718. (3) Siegwart, D. J.; Oh, J. K.; Matyjaszewski, K. ATRP in the Design of Functional Materials for Biomedical Applications. Prog. Polym. Sci. 2012, 37, 18−37. (4) Ladd, J.; Zhang, Z.; Chen, S.; Hower, J. C.; Jiang, S. Zwitterionic Polymers Exhibiting High Resistance to Nonspecific Adsorption from Human Serum and Plasma. Biomacromolecules 2008, 9, 1357−1361. (5) Enciso, A. E.; Fu, L.; Russell, A. J.; Matyjaszewski, K. A Breathing Atom-Transfer Radical Polymerization: Fully Oxygen-Tolerant Polymerization Inspired by Aerobic Respiration of Cells. Angew. Chem., Int. Ed. 2018, 57, 933−936. (6) Matyjaszewski, K.; Dong, H.; Jakubowski, W.; Pietrasik, J.; Kusumo, A. Grafting from Surfaces for “Everyone”: ARGET ATRP in the Presence of Air. Langmuir 2007, 23, 4528−4531. (7) Lee, B. S.; Kim, H.; Choi, I. S.; Cho, W. K. Formation of Activation-Free, Selectively Bioconjugatable Poly(N-acryloxysuccinimide-co-oligoethylene glycol methyl ether methacrylate) Films by Surface-Initiated ARGET ATRP. J. Polym. Sci., Part A: Polym. Chem. 2017, 55, 329−337. (8) Lee, B. S.; Kim, J. Y.; Park, J. H.; Cho, W. K.; Choi, I. S. Comparative Study on Surface-Initiated ATRP and SI-ARGET ATRP of Oligo(Ethylene Glycol) Methacrylate on Gold. J. Nanosci. Nanotechnol. 2016, 16, 3106−3109. (9) Hong, D.; Hung, H.-C.; Wu, K.; Lin, X.; Sun, F.; Zhang, P.; Liu, S.; Cook, K. E.; Jiang, S. Achieving Ultralow Fouling under Ambient Conditions via Surface-Initiated ARGET ATRP of Carboxybetaine. ACS Appl. Mater. Interfaces 2017, 9, 9255−9259. (10) Forbes, D. C.; Creixell, M.; Frizzell, H.; Peppas, N. A. Polycationic Nanoparticles Synthesized Using ARGET ATRP for Drug Delivery. Eur. J. Pharm. Biopharm. 2013, 84, 472−478. (11) Averick, S. E.; Bazewicz, C. G.; Woodman, B. F.; Simakova, A.; Mehl, R. A.; Matyjaszewski, K. Protein-Polymer Hybrids: Conducting ARGET ATRP from a Genetically Encoded Cleavable ATRP Initiator. Eur. Polym. J. 2013, 49, 2919−2924. (12) Kim, J. Y.; Lee, B. S.; Choi, J.; Kim, B. J.; Choi, J. Y.; Kang, S. M.; Yang, S. H.; Choi, I. S. Cytocompatible Polymer Grafting from Individual Living Cells by Atom-Transfer Radical Polymerization. Angew. Chem., Int. Ed. 2016, 55, 15306−15309. (13) Yu, Q.; Zhang, Y.; Wang, H.; Brash, J.; Chen, H. Anti-Fouling Bioactive Surfaces. Acta Biomater. 2011, 7, 1550−1557. (14) Homola, J. Surface Plasmon Resonance Sensors for Detection of Chemical and Biological Species. Chem. Rev. 2008, 108, 462−493. (15) Kim, J.; Hong, D.; Jeong, S.; Kong, B.; Kang, S. M.; Kim, Y.-G.; Choi, I. S. Aryl Azide Based, Photochemical Patterning of Cyclic Olefin Copolymer Surfaces with Non-Biofouling Poly[(3(methacryloylamino)propyl)dimethyl(3-sulfopropyl)ammonium hydroxide]. Chem. - Asian J. 2011, 6, 363−366.

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.langmuir.9b00822. Materials, experimental details including microcontact printing (μCP) for generating patterned initiator surfaces, enzyme-linked immunosorbent assay (ELISA) for protein fouling assay, comparison between the closed and open systems of the SI-ARGET ATRP, and results for the typical SI-ATRP under different gas conditions (PDF) 7749

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750

Article

Langmuir (16) Dunderdale, G. J.; Urata, C.; Miranda, D. F.; Hozumi, A. LargeScale and Environmentally Friendly Synthesis of pH-Responsive OilRepellent Polymer Brush Surface under Ambient Conditions. ACS Appl. Mater. Interfaces 2014, 6, 11864−11868. (17) Zhang, T.; Du, Y.; Kalbacova, J.; Schubel, R.; Rodriguez, R. D.; Chen, T.; Zahn, D. R. T.; Jordan, R. Wafer-Scale Synthesis of Defined Polymer Brushes under Ambient Conditions. Polym. Chem. 2015, 6, 8176−8183. (18) Zhang, T.; Du, Y.; Müller, F.; Amin, I.; Jordan, R. SurfaceInitiated Cu(0) Mediated Controlled Radical Polymerization (SICuCRP) Using a Copper Plate. Polym. Chem. 2015, 6, 2726−2733. (19) Brault, N. D.; Sundaram, H. S.; Li, Y.; Huang, C.-J.; Yu, Q.; Jiang, S. Dry Film Refractive Index as an Important Parameter for Ultra-Low Fouling Surface Coatings. Biomacromolecules 2012, 13, 589−593. (20) Lee, B. S.; Chi, Y. S.; Lee, K.-B.; Kim, Y.-G.; Choi, I. S. Functionalization of Poly(oligo(ethylene glycol) methacrylate) Films on Gold and Si/SiO2 for Immobilization of Proteins and Cells: SPR and QCM Studies. Biomacromolecues 2007, 8, 3922−3929. (21) Ma, H.; Wells, M.; Beebe, T. P.; Chilkoti, A. Surface-Initiated Atom Transfer Radical Polymerization of Oligo(ethylene glycol) Methyl Methacrylate from a Mixed Self-Assembled Monolayer on Gold. Adv. Funct. Mater. 2006, 16, 640−648. (22) Yeow, J.; Chapman, R.; Gormley, A. J.; Boyer, C. Up in the Air: Oxygen Tolerance in Controlled/Living Radical Polymerisation. Chem. Soc. Rev. 2018, 47, 4357−4387. (23) Fontes, C. M.; Achar, R. K.; Joh, D. Y.; Ozer, I.; Bhattacharjee, S.; Hucknall, A.; Chilkoti, A. Engineering the Surface Properties of a Zwitterionic Polymer Brush to Enable the Simple Fabrication of Inkjet-Printed Point-of-Care Immunoassays. Langmuir 2019, 35, 1379−1390. (24) Isaacs, N. S.; van Eldik, R. A Mechanistic Study of the Reduction of Quinones by Ascorbic Acid. J. Chem. Soc., Perkin Trans. 2 1997, 2, 1465−1468. (25) Cho, W. K.; Kong, B.; Choi, I. S. Highly Efficient NonBiofouling Coating of Zwitterionic Polymers: Poly((3(methacryloylamino)propyl)-dimethyl(3-sulfopropyl)ammonium hydroxide). Langmuir 2007, 23, 5678−5682. (26) Liu, W.; Li, Y.; Yang, B. Fabrication and Applications of the Protein Patterns. Sci. China Chem. 2013, 56, 1087−1100. (27) Shah, R. R.; Merreceyes, D.; Husemann, M.; Rees, I.; Abbott, N. L.; Hawker, C. J.; Hedrick, J. L. Using Atom Transfer Radical Polymerization to Amplify Monolayers of Initiators Patterned by Microcontact Printing into Polymer Brushes for Pattern Transfer. Macromolecules 2000, 33, 597−605. (28) Wu, C.-J.; Huang, C.-J.; Jiang, S.; Sheng, Y.-J.; Tsao, H.-K. Superhydrophilicity and Spontaneous Spreading on Zwitterionic Surfaces: Carboxybetaine and Sulfobetaine. RSC Adv. 2016, 6, 24827−24834. (29) Gormley, A. J.; Yeow, J.; Ng, G.; Conway, Ó .; Boyer, C.; Chapman, R. An Oxygen-Tolerant PET-RAFT Polymerization for Screening Structure-Activity Relationships. Angew. Chem., Int. Ed. 2018, 57, 1557−1562. (30) Chan, N.; Boutti, S.; Cunningham, M. F.; Hutchinson, R. A. Continuous Atom Transfer Radical Polymerization with Low Catalyst Concentration in a Tubular Reactor. Macromol. React. Eng. 2009, 3, 222−231. (31) Kim, B. J.; Cho, H.; Park, J. H.; Mano, J. F.; Choi, I. S. Strategic Advances in Formation of Cell-in-Shell Structures: From Syntheses to Applications. Adv. Mater. 2018, 30, No. 1706063.

7750

DOI: 10.1021/acs.langmuir.9b00822 Langmuir 2019, 35, 7744−7750