Design of comb-like poly(2-methyl-2-oxazoline) and its rapid co-deposition with dopamine for the study of antifouling properties
Ye Han Yan, Muhammad Atif, Ren Yong Liu, Hai Kun Zhu & Li Juan Chen
KEYWORDS
Antifouling; dopamine; poly(2-methyl-2-oxazoline); capillary electrophoresis
1. Introduction
Biofouling significantly compromises a material’s surface performance and causes numerous problems such as bacterial infection, protein adsorption, and membrane fouling [1,2]. Surface modification by protein-resistant polymers is a common strategy to endow a material surface with an antifouling property [3,4]. Much work has been dedicated to building up hydrophilic and uncharged polymer brush surfaces, which creates highly hydrated surfaces and prevents protein adsorption onto material surfa- ces due to their outstanding water-trapping property [5]. The most commonly used protein-resistant polymers are poly(ethylene glycol) (PEG) [6,7], zwitterionic poly- mers [8,9], peptidomimetic polymers [10,11], and so on [12,13]. One of the more attractive candidates for such applications, poly(2-methyl-2-oxazoline) (PMOXA), is protein-resistant polymer because of its ability to sustain the formation of hydrogen- bonded water networks, and the formation of a hydration layer can prevent adsorp- tion of protein on the surface of materials [14,15]. The synthesis of PMOXA can be performed by living cationic ring-opening polymerization (CROP), which allows for the design of architectures of PMOXA by incorporating functional terminal groups and initiators [16]. Numerous strategies have been developed to prepare PMOXA coatings to incorporate hydrophilicity and antifouling properties in material surfaces [17–21], and the anchoring of hydrophilic PMOXA on a material’s surface is receiv- ing much attention.
The use of dopamine/polydopamine (PDA) as an anchor offers a promising route to fabrication of antifouling coatings of polymers because of its excellent adhesive property [22]. Oxidative polymerization of dopamine can form polydopamine coat- ings, although the details of the mechanism remain an active area of investigation. Functional polymers can tightly cover a material’s surface with the assistance of dopa- mine/PDA by the interaction of covalent/non-covalent. Xiang et al. [23] prepared PMOXA antifouling coatings by a two-step method based on PDA coating. They syn- thesized linear PMOXA with an amino end group, and grafted it onto a PDA coating to form PMOXA coatings; the entire coating preparation process took 48 h. A one- step co-deposition method based on dopamine to prepare antifouling coatings is attractive due to the simpler process. Zhang et al. [24] synthesized star-shaped PMOXA based on hyperbranched poly(ethylenimine) and PMOXA, followed by co- deposition with dopamine to prepare PMOXA-based antifouling coatings using differ- ent co-deposition times, and the fastest coating process took two hours. Dopamine/ PDA as anchors to fabricate antifouling coatings always need a long time. Therefore, a number of oxidants have been developed to speed up the self-polymerization of dopamine [25]. Zhang et al. [26] reported on a strategy to accelerate dopamine self- polymerization using H2O2 and CuSO4 as the trigger, and CuSO4/H2O2-triggered PDA coatings have been proved to have excellent stability and high uniformity [27]. It provides an effective way to quickly prepare antifouling polymer coatings covered on a material’s surface.
In this work, the comb-like PMOXA copolymer with amino groups, PMA, was
first designed and synthesized, followed by co-deposition with dopamine triggered by CuSO4/H2O2. The thickness, hydrophilicity, and surface chemical composition of the prepared coatings were characterized by ellipsometry, water contact angle (WCA) measurements, and X-ray photoelectron spectroscopy (XPS). Anti-protein adsorption and anti-platelet adhesion measurements were performed to evaluate the antifouling properties of the PMA-based coatings. Finally, the method of rapidly preparing PMA coatings triggered by CuSO4/H2O2 was used to modify capillary surfaces, and to sep- arate a mixture of egg white and alkaline proteins by capillary electrophoresis.
2. Methods
2.1. Materials
2-methyl-2-oxazoline (MOXA) was purchased from TCI (Shanghai, China), dried over calcium hydride (CaH2), and distilled before use. Methacrylic acid (MAA) was distilled with a small amount of hydroquinone under reduced pressure before use. Isopropanol (IPA) and acetonitrile (ACN) were dried over CaH2 and distilled before use. 2,2’-Azobis(2-methylpropionitrile) (AIBN) was recrystallized from methanol. 2-amino ethyl methacrylate hydrochloride (AEMA) was obtained from Aladdin (Shanghai, China). Methyl trifluoro methane sulfonate (MeOTf) was bought from TCI (Shanghai, China). Dopamine hydrochloride and triethylamine (TEA) were purchased from Sigma-Aldrich (St. Louis, Mo., USA). ACN, IPA, MAA, tris-(hydroxymethyl) amino methane (Tris), and phosphate were bought from Sinopharm Chemical Reagents (Shanghai, China). All proteins used in this paper were bought from Sigma- Aldrich (St. Louis, Mo., USA), FITC-BSA was prepared by the reported method [28], and all details are listed in the supplementary materials. Silicon (111) substrates with a natural oxidized layer coating were purchased from Zhejiang Crystal Photoelectric Technology Co. (Zhejiang, China). The fused silica capillaries were obtained from Yongnian Optical Fiber Co. (Hebei, China).
2.2. Synthesis of PMA
The synthesized routes of PMOXA-MA and PMA are presented in Figure 1. Poly(2- methyl-2-oxazoline)-methacrylate (PMOXA-MA) macromonomer must first synthe- size [29]. PMOXA-MA with a degree of polymerization (DP) of 10 or 20 is called PMOXA10-MA or PMOXA20-MA, respectively. The copolymerization of PMOXA and AEMA was performed using free radical polymerization, with AIBN as an initi- ator. The feed molar ratio of PMOXA to AEMA was fixed at 1:2, with details pre- sented in supplementary materials. PMOXA10-r-AEMA and PMOXA20-r-AEMA are called PM10A and PM20A, respectively, for convenience.
2.3. Surface modification
Glass slides/silicon wafers and gold chips were cleaned for 15 min in ethanol, soni- cated for 15 min in deionized water, rinsed with deionized water, and dried with a stream of nitrogen. To prepare PMA-based coatings with dopamine co-deposition triggered by CuSO4/H2O2, as a typical example, dopamine, PM10A, H2O2 (19.6 mM), and CuSO4 (5 mM) were well dissolved in a Tris-HCl buffer (50 mM, pH 8.5) for the preparation of reaction solutions. The concentration of dopamine was adjusted to 2 mg/mL, while the mass ratio of the dopamine/PM10A was fixed at 1:1. The bare glass substrates were dipped in the reaction solutions and shaken well in an air oscil- lator at 25 ◦C for 10–60 min. Finally, the samples were washed with distilled water and dried over a stream of nitrogen. Following the above procedure, PM10A, PM20A, and PM10A mixed with PM20A based coatings (referred to as mixed PMOXA-based coatings) were prepared, respectively. For comparison, the traditional co-deposition method under atmospheric conditions without CuSO4/H2O2 were used to prepare PM10A coatings, and marked as TPM10A-based coatings. A schematic illustration of the above prepared coatings is shown in Figure 2.
2.4. Capillary coating
The bare fused-silica capillaries were first open 0.2 cm optical window for UV-vis detector, and pretreated for 30 min by flushing with 1.0 M NaOH and 1.0 M HCl, and then by deionized water for 5 min. PM10A, dopamine, CuSO4, and H2O2 were all dis- solved in a buffer solution of Tris-HCl (50 mM, pH 8.5), and then injected into pre- treated capillaries, which were sealed for 60 min at 30 ◦C on both ends. Finally, they were flushed with deionized water to remove the contents of residual materials, fol- lowed by an air stream for 15 min, to obtain a PM10A-based coating for capillaries.
2.5. Characterization
2.5.1. NMR spectroscopy
1H-NMR spectra of samples were recorded by a Bruker DMX-300 spectrometer (Bruker, Germany) at 300 MHz. The spectrum was obtained using deuterated water (DHO) as a solvent at room temperature.
2.5.2. Ellipsometry
A variable angle spectroscopic ellipsometer (M-2000, Woollam Co., Inc., Lincoln, Neb.) was used to measure coating thickness, which was conducted in the spectral range of 370-1000 nm at angles of incidence of 65◦ and 75◦. CompleteEASE 4.81 soft- ware was used to analyze all data. The optical constants (extinction coefficient, refractive index) of the Si (n ¼ 3.865, k ¼ 0.020) and the SiO2 (n ¼ 1.465, k ¼ 0) were used to measure the layer thickness of SiO2 on freshly washed silicon surfaces. As described by the Cauchy model, each polymer layer was shown as a slab of uniform thickness having sharp interfaces and the optical properties.
2.5.3. X-ray photoelectron spectroscopy (XPS)
XPS analysis of material surface chemical composition was performed on an X-ray photoelectron spectrometer (VG ESCALAB MK II, VG Scientific Instruments, England) using a 1486.6 eV Al (Ka) X-ray source.
2.5.4. Water contact angle measurements (WCA)
Measurements of static water contact angles were performed at ambient temperature with a CA system (SL200KB, USA KINO Industry Co., Ltd, Boston, USA) by water droplets of 2 mL, which were injected to the surface by a microliter syringe; values were recorded after 5 sec. Each measurement was performed at three points on the sample surface, and these results are presented as the mean ± standard devi- ation (n ¼ 3).
2.6. Evaluation of antifouling properties
2.6.1. Evaluation of protein adsorption by fluorescence imaging
Due to the strong adsorption tendency of BSA to the material’s surface, FITC-BSA was selected as a model protein to evaluate protein adsorption [30]. Samples were incubated for 2 h at 37 ◦C in a solution of FITC-BSA with a concentration of 0.5 mg/ mL (pH 7.4, 10 mM, PBS) in a dark room, followed by washing with PBS and deion- ized water to remove weakly bound FITC-BSA. The fluorescence images of all sam- ples were taken using an Olympus IX81 inverted microscope (Olympus, USA). The quantitative analysis of the adsorbed amount was performed using Image J software, based on the color intensity of fluorescence images, and the results are shown as mean ± standard deviation (n ¼ 3).
2.6.2. Surface plasmon resonance (SPR)
A Biacore (Uppsala, Sweden) T200 instrument was used to monitor protein adsorp- tion on surfaces. The SPR sensor chips before and after modification were mounted on the sample holders, docked, and then primed with PBS buffer for 6 min; the flow rate was set at 10 mL·min—1. The sensor was first rinsed with PBS buffer for 300 sec to obtain the baseline signal, and 0.1 mg·mL—1 protein solution was then flowed through four independent channels for 240 sec, followed by flushing with PBS buffer for another 300 sec. The response signal from the SPR measurement is given in response units (RU). The amount of protein absorbed on surfaces was quantified by measuring the change in RU value (DRU) in the buffer baselines before and after protein adsorption. In this study, 10 RU equals a change of approximately 1.0 ng·cm—2 in surface protein concentration on the chip surface [24], the results are shown as mean ± standard deviation (n ¼ 3).
2.6.3. Platelet adhesion
Platelet adhesion measurements were carried out according to the standard protocol [31]. Briefly, fresh whole blood (donated by healthy volunteers), was trans- ferred to a 5.0 mL vacuum tube using 109 mM sodium citrate as an anti-coagulant, and the whole blood was centrifuged for 15 min at 1200 rpm to obtain platelet-rich plasma. Samples were incubated for 2 h at 37 ◦C in platelet-rich plasma. After flushing three times with the PBS solution, the adhered platelets were fixed by 2.5% glutaral- dehyde for 30 min, followed by more flushing with the PBS solution, and dehydration by 25%, 50%, 75%, 90%, and 100% ethanol solutions for 20 min each. Gold sputtering of the samples was performed in a vacuum and observed by scanning electron microscopy (Sirion200, FEI, USA) at þ20 kV.
2.6.4. Protein separation by CE
A Beckman P/ACE MDQ capillary electrophoresis system with a diode array detector (Beckman Coulter Instruments, USA) was used to separate protein by capillary elec- trophoresis (CE). PMOXA-coated capillaries were first rinsed for 5 min with a buffer solution to maintain a balanced pH, and then protein mixture samples (ribonuclease A; cytochrome c; lysozyme; and a-chymotrypsinogen A, 0.2 mg/mL) were injected for 5 s with 3447.5 Pa. A hen egg was chosen as the analytical sample and pretreated as follows. The egg yolk and white were separated. The egg white was diluted with 20 mM Tris buffer (pH 7.40) at a ratio of 1:20, and filtered through a 0.22 lm milli- pore membrane filter [32].
3. Results and discussion
3.1. Synthesis of PMOXA-MA and PMA
In this work, the comb-like copolymer PMA was synthesized by PMOXA-MA ran- dom co-polymerization with an AEMA monomer. The initial step was to synthe- size PMOXA-MA with a DP of 10 and 20 [28,29]. The successful end functionalization of PMOXA was confirmed by the 1H-NMR spectrum, as shown in Figure 3(a). It can clearly see that two vinylic protons at 5.62 and 6.10 ppm (da, CH2) came from the methacrylate end group of PMOXA-MA, and the methylene protons at 4.29 ppm (dc, -O-CH2-) next to the ester functionality, by the way DHO solvent peak was appeared at d ¼ 4.70 ppm. The comparison of peak integral areas derived from the vinylic protons at 5.62 and 6.10 ppm (da, ¼CH2) with the peak of a-terminal methyl group at 3.05 ppm (df, -CH3) shows the degree of functionaliza- tion, and the result is 64.5% for PM10A and 94.5% for PM20A. The DP of the PMOXA-MA macromonomer was also confirmed by the integral comparison of the PMOXA-MA pendant methyl group signal around 2.05 ppm (de, -CH3) with the peak of the a-terminal methyl group at 3.05 ppm (df, -CH3), and the result is close to the design value (supplementary materials, Figure S1). Figure 3(b) shows the 1H-NMR spectrum of the PMA; the resonance peaks at 3.34 ppm (dd, -O-CH2- CH2-) and 4.25 ppm (de, -O-CH2-CH2-) are ascribed to the proton of AEMA. The resonance peaks at 2.07 ppm (db, -CH3) and 3.45 ppm (dc, -CH2-) are attributed to the PMOXA segment. Obviously, after copolymerization, the methacrylate end group in PMOXA-MA had almost all disappeared, and the backbone signals of PMA around 0.5 to 1.5 ppm (dfþg, -CH2- and -CH3) appeared. The copolymer compositions were calculated by 1.5 dd/da and determined by 1H-NMR, as pre- sented in Table 1 and supplementary materials, Figure S2. The ratios of PMOXA to AEMA from obtained copolymers are close to the monomer feed ratios, which means the PMA copolymers are successful.
3.2. Characterization of PMA-based coatings
The thickness of PMA-based coatings was evaluated with an ellipsometer, as shown in Figure 4. Obviously, the thickness increased with the increment of coating depos- ition time in both traditional and CuSO4/H2O2-accelerated methods. Among them, dopamine mixed with PM10A, co-deposition triggered by CuSO4/H2O2 presented the fastest growth rate in the first 10 min, and achieved a thickness value of 9.0 ± 0.1 nm. By further prolonging the co-deposition time, the thickness slowly increased, and the thickness of the coating could reach about 13.0 ± 0.2 nm within 60 min. By contrast, TPM10A presented a slow growth rate, and the thickness of the coating reached about 2.0 ± 0.1 nm during the first 60 min by the traditional method without CuSO4/H2O2. Oxygen plays a key role in the dopamine self-polymerization process, in an aerobic environment, dopamine must undergo oxidation, rearrangement, structural trans- formation, and deposition, which is a slow process [33]. Under the accelerated condi- tion, Cu2þand H2O2 produce reactive oxygen species (ROS), including O2·-and HO2·, as well as OH·. These radicals play an important role in the dopamine self-polymer- ization process and greatly improve the deposition rate of polydopamine coatings. Some work has shown that a small amount of CuSO4/H2O2 can enormously enhance the deposition rate of PDA coatings on various substrates [27]. Compared to the coating thickness formed from PMOXA with different chain lengths, it is easy to find that the short-chain PMOXA10 presents a much faster deposition rate in the same time, probably because the short chain is more likely to be fixed to the substrate material surfaces by a polydopamine anchor, and then to present a thicker coating. Based on the above results, we have investigated dopamine with a PM10A co- deposition accelerated system by a surface chemical composition study using XPS. Figure 5(a) presents XPS wide-scan spectra of dopamine mixed with PM10A depos- ited with deposition times of 40 min and 60 min. Comparing the results, the strong signals of Si as well as O were presented on the bare glass wafers, while no N signal was detected. The presence of a C signal on the bare substrate surface may come from chemicals during the manufacturing process. A significant increase in C 1 s and N 1 s signals was observed at the cost of a decrease in Si 2p, Si 2 s, and O 1 s signals for PMA-coated glass substrates.
The XPS high-resolution N 1 s and C 1 s scans were investigated, as shown in Figure 5(b). The C 1 s spectra split into three peaks of C–C/C–H ( 284.79 eV), C–N/ C–O ( 285.95 eV), and C O ( 287.99 eV) [34]. The N 1 s core-level spectra also split into three peaks corresponding to C-N ( 399.49 eV), N-C O ( 400.17 eV), and C-NH3þ ( 401.19 eV) [24,35]. By contrast, after coating, one extra peak N- C O appeared on the PMA coatings from PMOXA, and another additional peak C- NH3þ, resulting from polydopamine and AEMA segments. This means that PMA with dopamine can anchor effectively onto glass substrate triggered by CuSO4/H2O2 within 60 min. The surface chemical compositions of five types of glass substrate surfaces were studied by XPS, with results as shown in Table 2. The surface of bare glass consisted of Si and O derived from surface oxides, and after coating, the intensity of N 1 s sig- nals increased because the PMA copolymer had anchored onto the surfaces, assisted by dopamine. At the same time, the intensity of Si 2 s signals significantly decreased after coating. The silicon signal coverage after PM10A coating was the best, which was in line with the obtained results of coating thickness. To study the hydrophilicity of PMA coatings, WCA measurement was carried out because it is the most commonly used method to assess the hydrophilicity of material surfaces. Pictures of water droplets are presented in Figure S3, and, as listed in Table 2, the bare glass surface has a static contact angle of about 68◦. After coating, the modified surfaces were found to consistently decrease to 44◦, 45◦, and 48◦, respectively, for the PM10A, mixed PMOXA, and PM20A based coatings. By contrast, TPM10A based coatings presents a larger static contact angle of about 55◦ due to the slow deposition rate under the traditional method, and less PMOXA covered on the substrate. In conclusion, the existence of the PMA based coatings leads to an appar- ent increase in hydrophilicity, and these results are in line with the results of XPS and thickness of coatings.
3.3. Antifouling performance of PMA-based surfaces
Based on the WCA results, fluorescence tests were first carried out to evaluate the surfa- ce’s antifouling property. The model protein was chosen as FITC-BSA because BSA has strong adsorption characteristics on material surfaces [36]. The adsorption of FITC-BSA on the material’s surface was observed by fluorescence microscopy, as shown in Figure 6. It can be seen that the PMA-coated surfaces present low fluorescence, but the bare glass surface showed extremely strong fluorescence. To more accurately compare the protein-resistant effect on the material’s surface, the relative quantitative analysis of Results from fluorescence microscopy: (a) bare glass surface; (b) TPM10A-based coating 60 min; (c) PM10A-based coating 40 min; (d) PM10A-based coating 60 min; (e) mixed PMOXA-based coating 60 min; (f) PM20A-based coating 60 min. Bar graph on right presents the relative fluores- cence intensity of these test samples the corresponding images is provided in Figure 6 using fluorescence intensities. Obviously, the coatings from the accelerated system showed a better anti-protein adsorption effect at the same coating deposition time, and the adsorption capacity of FITC-BSA could be reduced to 15% of the original adsorption capacity. By contrast, PM10A with a short PMOXA chain was easily anchored onto substrates by polydop- amine, and presented much better anti-protein adsorption ability. As Figure 6(d) shows, the adsorption capacity of FITC-BSA can be reduced to 5%, and these results are in line with the previous thickness and XPS results. To test and compare the protein-resistant properties of the PMA-based coatings, TPM10A-based coatings, PM10A-based coatings, mixed PMOXA-based coatings, and PM20A-based coatings were used to modify gold chip sensors, respectively, and five model proteins were chosen for SPR study: BSA (66 kDa, pI 4.8), lysozyme (14.3 kDa, pI ¼ 11.1), cytochrome c (13.0 kDa, pI ¼ 10.2), ribonuclease A (13.7 kDa, pI ¼ 9.3) and a-chymotrypsinogen A (25.7 kDa, pI 9.2).
Typical SPR sensorgrams in Figure 7 show that the gold surface adsorbed large amounts of proteins. By contrast, TPM10A-based coatings present better protein-resistant properties than gold surfaces due to the existence of PMOXA; however, it is far less effective than PM10A-based coat- ings. Obviously, very small amounts of protein (~95% reduction relative to gold surface) adsorbed on PM10A-based coatings, and the adsorption amount of five proteins were all less than 10 ng/cm2, indicating the outstanding properties of resisting protein adsorption which is dependent on the surface PMOXA chain densities. The comb-like PMOXA with adjustable PMOXA antifouling segment and -NH2 anchor segment could co- deposit with dopamine quickly, and the PMOXA chains covered on materials surface is higher than the traditional methods in the short co-deposition times. Platelet adhesion measurement is also a classical method to evaluate the anti-fouling ability of a material’s surface [37]. Therefore, the adsorption of platelets adhered to a material’s surface was also investigated and characterized by SEM images. Figure 8(a,b) show significant adhesion of blood platelets on the glass and TPM10A-coated surfaces. By contrast, as seen in Figure 8(c–f), the PMA-coated surfaces under accelerated condi- tions exhibit excellent resistance to platelet adhesion. It can be concluded that the PMA based coatings can effectively improve the anti-platelet adhesion property.
3.4. Application in CE for protein separation
CE as a powerful analytical tool has many advantages, such as high resolution, high separation efficiency, and less consumption of samples and reagents, so it is more con- venient for separation of proteins [38]. Alkaline proteins have a great tendency to adsorb on the inner surface of fused-silica capillaries by electrostatic interaction during CE analysis, resulting in band-broadening and poor repeatability [39]. To solve this problem, many antifouling polymer coatings are used to modify the capillary inner wall to resist protein adsorption. To further investigate its practical application, the co- deposition of PM10A with dopamine triggered by CuSO4/H2O2 was applied in the preparation of PMA based coatings modified capillaries, and was used for the separ- ation of a mixture of four alkaline models of proteins (lysozyme, a-chymotrypsinogen A, cytochrome c, and ribonuclease A) and egg white proteins. As shown in Figure 9, combined peaks and several broadenings were detected by the use of a bare fused-silica capillary because of the strong adsorption of proteins on the inner surface of the capil- lary. When using PMA-coated capillaries, a good baseline separation of proteins can be achieved due to the existence of the PMOXA antifouling part, whose peak identification is shown in Figure 9, and it presents a low relative standard deviation (RSD < 0.71%,, as seen in Table S1) of migration time in nine times of consecutive separation.
To evaluate the applicability of a PMA-coated capillary, a complex sample analysis is necessary. Egg white proteins were chosen for analysis, and, as shown in Figure 10, it is difficult to detect the egg white proteins when using a bare fused silica capillary in CE analysis, and only a small broad peak appeared in the first running program. Compared to the bare capillary, the three types of egg white proteins were detected successfully by CE, and the peak identification as the reported order [40]. After nine times of consecutive separation for egg white proteins, the migration times of the main peaks showed almost no change, which means the PMA-coated capillary exhibits good stability for pro- tein separation.
4. Conclusions
In this paper, a comb-like PMOXA was synthesized, and PMA based coatings were constructed by the rapid co-deposition of dopamine with PMA triggered by CuSO4/ H2O2. Compared to traditional methods, the co-deposition time was much reduced, to ~60 min, and the PMA based coatings thickness could achieve about 13 nm. Furthermore, the PMA-coated surfaces presented an excellent protein-repellent prop- erty and anti-platelet adhesion ability. The PMA based coatings were applied to capil- lary inner surface modification, and the PMA based coatings modified capillary could achieve alkaline model proteins mixture and egg white proteins baseline separation by CE, and exhibited excellent stability. To sum up, the practicability of this promis- ing PMA-based coatings could be used for further proteomics analysis.
Disclosure statement
No potential conflict of interest is reported by the authors.
Funding
This work was supported by the National Natural Science Foundation of China (no. 11804253, 21804100) and the Natural Science Foundation of Anhui (no. KJ2018A0416; 1808085QB44).
References
[1] Banerjee I, Pangule RC, Kane RS. Antifouling coatings: Recent developments in the design of surfaces that prevent fouling by proteins, bacteria, and marine organisms. Adv Mater. 2011;23(6):690–718.
[2] Rana D, Matsuura T. Surface modifications for antifouling membranes. Chem Rev.
2010;110(4):2448–2471.
[3] Serrano A^ , Sterner O, Mieszkin S, et al. Nonfouling response of hydrophilic uncharged polymers. Adv Funct Mater. 2013;23(46):5706–5718.
[4] Lowe S, O’Brien-Simpson NM, Connal LA. Antibiofouling polymer interfaces: poly(- ethylene glycol) and other promising candidates. Polym Chem. 2015;6(2):198–212.
[5] Chen S, Li L, Zhao C, et al. Surface hydration: principles and applications toward low-
fouling/nonfouling biomaterials. Polymer 2010;51(23):5283–5293.
[6] Shi X, Tan L, Xing J, et al. Synthesis of hydroxyethylcellulose-g-methoxypoly (ethylene glycol) copolymer and its application for protein separation in CE. J Appl Polym Sci. 2012;128:1995–2002.
[7] Xing J, Tan L, Cao F, et al. Synthesis of star polymer poly(ethylene glycol)3–poly(N,N- dimethyl acrylamide) and its application in protein resistance and separation. J Appl Polym Sci. 2013;129(3):1179–1186.
[8] Chen L, Tan L, Liu S, et al. Surface modification by grafting of poly(SBMA-co-AEMA)-
g-PDA coating and its application in CE. J Biomater Sci Polym Ed. 2014;25(8):766–785.
[9] Cao F, Tan L, Xiang L, et al. Application of the copolymers containing sulfobetaine methacrylate in protein separation by capillary electrophoresis. J Biomater Sci Polym Ed. 2013;24(18):2058–2070.
[10] Tauhardt L, Kempe K, Gottschaldt M, et al. Poly(2-oxazoline) functionalized surfaces: from modification to application. Chem Soc Rev. 2013;42(20):7998–8011.
[11] Lorson T, Lu€btow MM, Wegener E, et al. Poly(2-oxazoline)s based biomaterials: a com- prehensive and critical update. Biomaterials 2018;178:204–280.
[12] Wang X, Yuan S, Shi D, et al. Integrated antifouling and bactericidal polymer mem- branes through bioinspired polydopamine/poly(N-vinyl pyrrolidone) coating. Appl Surf Sci. 2016;375:9–18.
[13] Tan L, Zheng X, Chen L, et al. Quality testing of human albumin by capillary electro- phoresis using thermally cross-linked poly(vinyl pyrrolidone)-coated fused-silica capil- lary. J Sep Sci. 2014;37(20):2974–2982.
[14] Konradi R, Acikgoz C, Textor M. Polyoxazolines for nonfouling surface coatings - a direct comparison to the gold standard PEG. Macromol Rapid Commun. 2012;33(19): 1663–1676.
[15] Morgese G, Benetti EM. Polyoxazoline biointerfaces by surface grafting. Eur Polym J. 2017;88:470–485.
[16] Wiesbrock F, Hoogenboom R, Leenen MAM, et al. Investigation of the living cationic ring-opening polymerization of 2-methyl-, 2-ethyl-, 2-nonyl-, and 2-phenyl-2-oxazoline in a single-mode microwave reactor. Macromolecules 2005;38(12):5025–5034.
[17] Zhu H, Chen L, Zhang C, et al. A new strategy to prepare poly(2-methyl-2-oxazoline)- based antifouling coatings: using uv-crosslinked copolymer anchors. Macromol Mater Eng. 2019;304(4):1800685.
[18] Mumtaz F, Chen C, Zhu H, et al. Controlled protein adsorption on PMOXA/PAA based coatings by thermally induced immobilization. Appl Surf Sci. 2018;439:148–159.
[19] Mao K, Du H, Bai L, et al. Poly(2-methyl-2-oxazoline) coating by thermally induced immobilization for determination of bovine lactoferrin in infant formula with capillary electrophoresis. Talanta 2017;168:230–239.
[20] Du H, Zhang C, Mao K, et al. A star-shaped poly(2-methyl-2-oxazoline)-based antifoul- ing coating: application in investigation of the interaction between acetaminophen and bovine serum albumin by frontal analysis capillary electrophoresis. Talanta 2017;170: 275–285.
[21] Konradi R, Pidhatika B, Mu€hlebach A, et al. Poly-2-methyl-2-oxazoline: a peptide-like polymer for protein-repellent surfaces. Langmuir 2008;24(3):613–616.
[22] Ryu JH, Messersmith PB, Lee H. Polydopamine surface chemistry: a decade of discov- ery. ACS Appl Mater Interfaces. 2018;10(9):7523–7540.
[23] Xiang L, Chen L, Tan L, et al. The preparation of a novel polydopamine-graft-poly(2- methyl-2-oxazoline) protein-resistant coating and its applications in protein separation. Chin Chem Lett. 2013;24(7):597–600.
[24] Zhang C, Liu S, Tan L, et al. Star-shaped poly(2-methyl-2-oxazoline)-based films: rapid preparation and effects of polymer architecture on antifouling properties. J Mater Chem B. 2015;3(27):5615–5628.
[25] Liu Y, Ai K, Lu L. Polydopamine and its derivative materials: synthesis and promising applications in energy, environmental, and biomedical fields. Chem Rev. 2014;114(9): 5057–5115.
[26] Zhang C, Ou Y, Lei W, et al. CuSO4/H2O2-induced rapid deposition of polydopamine coatings with high uniformity and enhanced stability. Angew Chem Int Ed. 2016;55(9): 3054–3057.
[27] Zhang C, Li HN, Du Y, et al. CuSO4/H2O2-triggered polydopamine/poly(sulfobetaine methacrylate) coatings for antifouling membrane surfaces. Langmuir 2017;33(5): 1210–1216.
[28] Bai L, Tan L, Chen L, et al. Preparation and characterizations of poly(2-methyl-2- oxazoline) based antifouling coating by thermally induced immobilization. J Mater Chem B. 2014;2(44):7785–7794.
[29] Kobayashi S, Masuda E, Shoda S, et al. Synthesis of acryl- and methacryl-type macro- monomers and telechelics by utilizing living polymerization of 2-oxazolines. Macromolecules 1989;22(7):2878–2884.
[30] Downard AJ, Jackson SL, Tan E. Fluorescence microscopy study of protein adsorption at modified glassy carbon surfaces. Aust J Chem. 2005;58(4):275–279.
[31] Xin Z, Yan S, Ding J, et al. Surface modification of polypropylene nonwoven fabrics via covalent immobilization of nonionic sugar-based surfactants. Appl Surf Sci. 2014;300: 8–15.
[32] Hsieh Y, Chen T, Liu C. Capillary electrochromatographic separation of proteins on a column coated with titanium dioxide nanoparticles. Electrophoresis. 2006;27(21): 4288–4294.
[33] Dreyer DR, Miller DJ, Freeman BD, et al. Elucidating the structure of poly(dopamine). Langmuir 2012;28(15):6428–6435.
[34] Liu Y, Chang C, Sun T. Dopamine-assisted deposition of dextran for nonfouling appli- cations. Langmuir 2014;30(11):3118–3126.
[35] Yang Z, Tu Q, Zhu Y, et al. Mussel-inspired coating of polydopamine directs endothe- lial and smooth muscle cell fate for re-endothelialization of vascular devices. Adv Healthcare Mater. 2012;1(5):548–559.
[36] Sun X, Wang H, Wang Y, et al. Creation of antifouling microarrays by photopolymeri- zation of zwitterionic compounds for protein assay and cell patterning. Biosens Bioelectron. 2018;102:63–69.
[37] Zhu J, Marchant RE. Dendritic saccharide surfactant polymers as antifouling interface materials to reduce platelet adhesion. Biomacromolecules 2006;7(4):1036–1041.
[38] Voeten RLC, Ventouri IK, Haselberg R, et al. Capillary electrophoresis: trends and recent advances. Anal Chem. 2018;90(3):1464–1481.
[39] Lucy CA, MacDonald AM, Gulcev MD. Non-covalent capillary coatings for protein separations in capillary electrophoresis. J Chromatogr A. 2008;1184(1–2):81–105.
[40] Chen L, Zeng R, Xiang L, et al. Polydopamine-graft-PEG antifouling 2-Aminoethyl coating for quanti- tative analysis of food proteins by CE. Anal Methods. 2012;4(9):2852–2859.