- Open Access
Large area micropatterning of cells on polydimethylsiloxane surfaces
© Moustafa et al.; licensee BioMed Central Ltd. 2014
- Received: 8 August 2014
- Accepted: 30 September 2014
- Published: 24 October 2014
Precise spatial control and patterning of cells is an important area of research with numerous applications in tissue engineering, as well as advancing an understanding of fundamental cellular processes. Poly (dimethyl siloxane) (PDMS) has long been used as a flexible, biocompatible substrate for cell culture with tunable mechanical characteristics. However, fabrication of suitable physico-chemical barriers for cells on PDMS substrates over large areas is still a challenge.
Here, we present an improved technique which integrates photolithography and cell culture on PDMS substrates wherein the barriers to cell adhesion are formed using the photo-activated graft polymerization of polyethylene glycol diacrylate (PEG-DA). PDMS substrates with varying stiffness were prepared by varying the base to crosslinker ratio from 5:1 to 20:1. All substrates show controlled cell attachment confined to fibronectin coated PDMS microchannels with a resistance to non-specific adhesion provided by the covalently immobilized, hydrophilic PEG-DA.
Using photolithography, it is possible to form patterns of high resolution stable at 37°C over 2 weeks, and microstructural complexity over large areas of a few cm2. As a robust and scalable patterning method, this technique showing homogenous and stable cell adhesion and growth over macroscales can bring microfabrication a step closer to mass production for biomedical applications.
- Cell micropatterning
- Poly (dimethyl siloxane)
- Poly (ethylene glycol)
The development of methods to spatially direct cell growth in two and three dimensions is a fundamental challenge for in vitro research and simulating in vivo cellular microenvironments[1, 2]. Beyond applications in tissue engineering and microarray technologies, precisely controlling the location of cells has potential in furthering our understanding of fundamental cellular processes[3–5]. Precise regulation of cell response and fate can reveal insights into intercellular interactions and cues. By integrating microfabrication strategies, it is possible to form efficiently controlled cell cultures, or lead to hierarchical organization as tissues and organs. Typically, spatial control has been achieved by creating well-defined physical or biochemical barriers, or cell-adhesive regions to encourage specific attachment. Conversely, controlling non-specific adsorption in regions where cell growth is not desired provides a similar effect. Nevertheless, maintaining microscale precision and uniformity over large areas (cm), a characteristic of live tissues, remains a challenge for translation to application.
Various synthetic and natural materials are used as substrates for cell growth and differentiation. As an alternative to rigid polystyrene and glass surfaces, poly (dimethylsiloxane) (PDMS) is a versatile polymer that has been widely used as an elastic, stretchable, cellular substrate in the form of microfluidic channels, microwells, and micro- and nano-pillars[3, 9, 10]. PDMS possesses unique advantages including optical transparency, biocompatibility, flexibility, tunable mechanical properties, oxygen permeability, durability and low cost. However, the surface of PDMS is highly hydrophobic (contact angle ~105°) which tends to result in the non-specific adsorption of proteins and other biomolecules required for cell attachment and growth[12, 13]. Surface modification is therefore required for effective spatial regulation of cells. Typically, cells have been grown on PDMS substrates without specific spatial control[11, 14]. On the other hand, micropatterning strategies on PDMS have usually involved microcontact printing (μCP) to form high resolution cell-adhesive patterns[15, 16]. Surface modification using plasma oxidation of PDMS to increase hydrophilicity followed by surface functionalization has also been reported. However, these physisorption approaches are typically non-covalent in nature, confined to small areas (mm), or do not present adequate physical barriers for cellular growth, making them short-lived. Forming physico-chemical barriers can confine cells to adhesive regions, while allowing growth over extended periods of time and over large areas. Adapting photolithography to form geometrically distinct barriers for specific cell attachment provides the ability to easily fabricate high resolution patterns over large areas[5, 19].
Surface modification using hydrophilic and neutrally charged polymers, in particular homo and hetero-functional polyethylene glycol (PEG) hydrogels, has been extensively used to repel non-specific protein adsorption and guide cell attachment[20–24]. Using PEG on PDMS therefore provides a means to precisely direct cell adhesion. However, covalently attaching PEG to PDMS has been difficult. Photo-induced grafting for surface modification of PDMS was first demonstrated using acrylic acid (PAA), acrylamide and polyethylene glycol methacrylate (PEG-MA) monomers. However, both PEG-MA and PAA are not optimal owing to surface charges and gradual loss of hydrophilicity resulting in eventual cell adhesion. Further, PEG-MA yields fragile patterning and cannot withstand physiological or microfluidic shear stresses. Photo-induced graft polymerization using polyethylene glycol diacrylate (PEG-DA) was used to micropattern PDMS, which is effective with long-lasting hydrophilic properties and stable patterns over 2 months. The micropatterned-PEGDA-grafted PDMS was applied to protein adsorption and cell adhesion. However, a reliable strategy to form stable, micro and macroscale patterns over large areas on different PDMS compositions is still challenging.
One of the advantages of using PDMS is its tunable mechanical nature. By controlling the ratio of monomer to crosslinker, the stiffness of the underlying substrate can be altered. This in turn has a great influence on the cell growth on the surface. For instance, our group and others have previously shown that human embryonic stem cell proliferation can be affected by varying the stiffness[28, 29]. Characteristically, exposure of cell-binding motifs differ based on the nanoscale surface stiffness, reflecting changes in cell behavior. In this work, we investigate the photopatterning of PEG-DA hydrogels that can be used on PDMS surfaces with different substrate stiffness. We show a facile strategy that allows the fabrication of stable, high resolution patterns for microfluidics and culture of cells over large areas (several cm). Areas covered by PEG-DA are used to prevent non-specific adhesion and confine the cells to spatially defined microstructural features. We demonstrate fibroblast attachment to these patterns and show homogenous and stable cell adhesion and growth over macroscales that can bring microfabrication a step closer to mass production over larger scales for biomedical applications.
Non-specific adsorption of proteins on surfaces is a common problem with various biomedical devices such as biosensors, microfluidic devices, and microarrays. Despite a host of favorable properties including flexibility, tunable mechanical properties and oxygen permeability, PDMS surfaces have required surface modification owing to a hydrophobic nature and such non-specific adsorption[11, 31]. A commonly used method for blocking the adsorption of proteins involves immobilizing hydrophilic and neutrally charged polymers to protect the surface. Strategies to prevent protein adsorption can also be used to spatially corral cells on to modified PDMS surfaces. In particular, immobilization of poly (ethylene glycol) (PEG) on surfaces has been widely adopted. Using acrylate functionalized PEGs further allows the integration of microfabrication via photolithography on such surfaces. In this work, micropatterns of PEG-DA were covalently attached to PDMS surfaces as a means to physically control the spatial positioning of cells at the microscale. Since the PEG regions are resistant to the non-specific adhesion of cells, cell growth is confined to exposed regions of the PDMS which in turn, can be functionalized as desired.
Microchannels of PEG-DA on PDMS surfaces
Optimization of micropatterning and stability
The overall strategy focused on two primary objectives – i) high fidelity of micro-architectures over a large (cm) area and, ii) stability of the covalently bound PEG microstructures to permit long-term cell culture. The covalent attachment of PEG to the underlying PDMS was developed using modifications of the methods described earlier[10, 35]. In order to form stable patterns over large areas, several parameters had to be optimized including intensity of the UV light source and time of exposure, which contribute to the amount of light energy transferred to the substrate. Over or under-development during photolithography can result in the formation of cloudy hydrogels, or microstructures that delaminate from the PDMS within a few hours. One successful modification used involves brief intervals between UV exposures that allows heat to dissipate and prevent possible thermal polymerization. In addition, the concentration of the photoinitiator benzophenone, and the chain transfer agent benzyl alcohol are also modulated. Benzyl alcohol aids in the diffusion of the reactive monomers to PDMS surface by decreasing the solution viscosity[31, 35]. This facilitates a stable attachment of the hydrogel to the underlying PDMS which can otherwise delaminate given the large areas. Hydrogels were examined for stability by incubating in a water bath at 37°C over a period of 2 weeks. No delamination was observed and the hydrogels remained intact with the channels maintaining their width. Samples were also observed to be stable over a month when stored dry and rehydrated, showing that this is a robust method. (Image shown in the Additional file1: Figure S1).
Removal of benzophenone post-patterning
Effect of substrate stiffness
Cell behavior in terms of proliferation, spreading and attachment can be regulated by altering the stiffness of the substrate. Acting as the in vitro extra cellular matrix (ECM), the mechanical properties influence the chemical and physical cues responsible for cell fate. Diverse cell types vary in terms of adhesion and proliferation to changes in stiffness of the substrate. For instance, neural progenitor cells were found to favor neuron and astrocyte differentiation on softer surfaces, but oligodendrocyte differentiation on stiffer substrates. This indicates that such change in the mechanical properties of the substrate can also influence lineage specification. One of the primary advantages of PDMS over substrates such as glass and polystyrene is its tunable mechanical nature and flexibility. However, to date, most studies involving changes in stiffness of PDMS used un-patterned surfaces[11, 28, 30]. Here, in order to optimize cell micropatterning for different cell types, underlying substrate stiffness was further tested as an additional cue.
Elastic moduli and stiffness of PDMS with varying base to curing agent ratios
6.10 ± 0.11
1.90 ± 0.04
2.95 ± 0.05
1.35 ± 0.02
1.38 ± 0.05
1.10 ± 0.04
Cell culture on patterned PDMS surfaces
Earlier, a higher adhesion density of 3T3 mouse fibroblasts on un-patterned 20:1 PDMS than the stiffer 5:1 and 10:1 surfaces had been reported. Despite being less stiff at the macroscale, 20:1 PDMS is more pliable at the nanoscale resulting in exposed cell-binding motifs. On the other hand, a different study found transformed fibroblasts to grow at similar rates on PDMS irrespective of stiffness. In our experiments, expectedly, the morphology and slow proliferation of the adhered cells is an indication that substrate stiffness is an important contributor in micropatterned samples as well. This is observed in Figure 3 which shows consistent fidelity of the features across samples but lower cell density. Both 5:1 and 20:1 PDMS maintained precise cell adhesion to the channels but at lower densities compared to the 10:1 samples (Figure 5c, d). Favoring 10:1 PDMS over 20:1 supports the claim that fibroblasts may migrate preferentially to stiffer substrates and exhibit stronger traction forces. Similarly, the cell proliferation on stiffer 5:1 PDMS was higher in comparison to 20:1 PDMS. Overall this could be attributed to the optimal cross-linking density of 10:1 PDMS and the higher amount of un-crosslinked components when increasing the cross-linker or based beyond the normal ratio. These components can be either mobile affecting the nutrients in growth media or stationary on the PDMS surface influencing cell attachment and growth. However, since softer substrates are desirable for certain cultures, it is important to note that overall, the formation of high-resolution PEG microstructures on PDMS of different stiffness demonstrates that this strategy can be adapted to substrates of tunable mechanical properties to control cell growth.
The integration of microfabrication and cell culture can result in efficient and directed cellular responses with uniform cell patterning over large length scales. In this study, we demonstrate a technique for the fabrication of stable and long-lasting PEG-DA hydrogels that are photografted on PDMS of varying stiffness. The resultant micropatterns can be formed at a high resolution down to 5–10 μm. The patterns are robust over a period of weeks and maintain function in cell culture medium, specifically resisting cell adhesion and directing cell growth. By increasing benzyl alcohol concentration in the PEG-DA monomer, decreasing benzophenone treatment and using intermittent UV exposure, both hydrogel attachment and resolution were optimized. Such measures further lead to specific, high density protein and cell adhesion on PDMS that was uniformly micropatterned up to an area of 2–4 cm2. The effect of altering the mechanical properties of the patterned substrate, through stiffness, on cell behavior was examined on 5:1, 10:1 and 20:1 PDMS samples. Controlled cell adhesion and proliferation was achieved for all three ratios with the cells migrating specifically to the PDMS channels surrounded by PEG-DA. 10:1 PDMS, which is very commonly used, was also shown to be a versatile and flexible surface in terms of growing fibroblasts on micropatterned regions. The ability to further functionalize and tune the PDMS surface and its underlying stiffness, provides a method to tailor the attachment and culture of various cells in different geometries. This study shows potential to further increase the micropatterning areas while reaching smaller widths on the microscale. More complex features can also be adapted that mimic the in vivo microenvironment and be used for directing cells into specific lineages and controlling their fate.
Poly (dimethyl siloxane) (PDMS) prepolymer and curing agent (Sylgard 184) were obtained from Dow Corning (Midland, MI). PEG-DA (number average molecular weight, 575) and albumin-fluorescein isothiocyanate conjugate were purchased from Sigma-Aldrich (St. Louis, MO). Benzophenone, acetone, Benzyl alcohol, Sodium periodate (NaIO4), methanol, paraformaldehyde powder and Phosphate buffered saline (PBS) solution (10X) were obtained from Fisher Scientific (Fair Lawn, NJ) and used as received. 3T3 mice fibroblasts and human dermal fibroblasts were used to test cell adhesion and cell growth. 4′, 6-diamidino-2-phenylindole (DAPI) and Alexa Fluor Phalloidin 488 (Life Technologies, Grand Island, NY) were used for staining. A bright-field reflective chrome photomask was designed using CleWin and custom fabricated to form large area grids consisting of 50 and 25 μm lines.
UV photografting of PEG-DA on PDMS
To prepare the PDMS substrates of varying stiffness, the mass ratio of base to curing agent was varied to form (in order of decreasing stiffness) 5:1, 10:1 and 20:1 samples. 12.5 g of pre-polymer was mixed with 1.25 g, 0.625 g and 2.5 g of curing agent respectively and added to a 60 mm plastic petridish. After overnight curing at 62°C, the PDMS was peeled off and diced into squares (~1-4 cm2). PDMS slabs were then immersed in a 10 wt. % benzophenone solution in acetone for 2 minutes. The samples were rinsed with methanol and air dried.
1 ml of 40wt% PEG-DA solution was prepared by dissolving 400 μl of PEG-DA, 10 μl of 100mM NaIO4 (1 mM) and 50 μl of benzyl alcohol (5 wt. %) in water. 65 μl of the reaction solution was cast on the PDMS surfaces. PEG-DA was photopolymerized through a photomask using a 365 nm, 500 mW/cm2 light source (OmniCure S1000, Lumen Dynamics). PEG-DA behaves as a negative photoresist in the presence of UV light and is crosslinked owing to the photoinitiator benzophenone [32, 45]. The schematic for this reaction is shown in the (Additional file 1: Figure S4). The exposure conditions were optimized by varying intensity and time of exposure to obtain large patterned areas. Figure 1 shows a schematic of the steps involved in obtaining a stable, micropatterned PEG-DA hydrogel on PDMS. Micropatterned PDMS samples were stored in 1x PBS and sterilized by UV exposure for 30 minutes (<0.01 W/cm2) prior to cell studies. To promote cell attachment, samples were subsequently immersed in 3 ml of 6 μg/ml fibronectin solution for one hour prior to cell culture.
Cell culture on micropatterned samples
3T3 mice fibroblasts were cultured at 37°C in 5% humidified environment in Dulbecco’s modified eagle’s medium (DMEM) with 10% fetal bovine serum. On reaching 90% confluence in four days, the cells were trypsinized (0.25% trypsin solution) and passaged at a density of 5×104 cells per PDMS sample. Human dermal fibroblasts (HDFs) were maintained in minimum essential medium supplemented with 2 mM l-glutamine, 1% penicillin/streptomycin, 15% fetal bovine serum, non-essential and essential amino acids, sodium pyruvate and vitamins. Cells were cultured on the 5:1 and 20:1 samples at the same density. For cell staining, PDMS samples were each covered with 1 mg/ml of albumin-fluorescein isothiocyanate (FITC-BSA) conjugate for 30 minutes before being examined under a fluorescence microscope (Nikon ECLIPSE TE2000-U). Following 6 days of cell culture, the samples were fixed by adding 4% paraformaldehyde solution for 30 minutes. Phalloidin 488 was added as a cytoskeletal stain and DAPI as a nuclear stain for 30 minutes each. The samples were washed 3 times for 5 minutes each using PBS wash buffer following each of the fixing and staining steps.
Nanomechanical measurements of PDMS substrates
Mechanical properties of crosslinked films of PDMS were measured using AFM-based nanoindentation (MFP-3D, Asylum Research, Santa Barbara, CA). All samples were indented using an AC160 TS cantilever (Olympus Research, Tokyo, Japan) with nominal spring constants varying from 30–40 N/m. The actual spring constants were determined prior to each experiment using the thermal fluctuation method on a hard mica surface. Different PDMS samples were indented in air with ~30 indents at different areas on the surface using constant force mode (100 nN). The Young’s modulus and stiffness were obtained via the Oliver-Pharr model in Igor Pro 6.22 A (Wavemetrics Inc., OR) .
This research was supported by a grant from the National Science Foundation (CBET – 1144611). Photomasks were custom fabricated in the Wright Virginia Microelectronics Center in the VCU School of Engineering. The authors thank Congzhou Wang for help with nanoindentation experiments and Dr. Xuejun Wen for the 3T3 mouse fibroblasts used in this study.
- Wang L, Sun B, Ziemer KS, Barabino GA, Carrier RL: Chemical and physical modifications to poly (dimethylsiloxane) surfaces affect adhesion of Caco-2 cells. J Biomed Mater Res A 2009, 93A: 1260-1271.Google Scholar
- Walker GM, Zeringue HC, Beebe DJ: Microenvironment design considerations for cellular scale studies. Lab Chip 2004, 4: 91-97. 10.1039/b311214dView ArticleGoogle Scholar
- Choi JH, Lee H, Jin HK, Bae J-s, Kim GM: Micropatterning of neural stem cells and Purkinje neurons using a polydimethylsiloxane (PDMS) stencil. Lab Chip 2012, 12: 5045-5050. 10.1039/c2lc40764gView ArticleGoogle Scholar
- Goudar VS, Suran S, Varma MM: Photoresist functionalisation method for high-density protein microarrays using photolithography. Micro Nano Lett 2012, 7: 549-553. 10.1049/mnl.2012.0336View ArticleGoogle Scholar
- Ross AM, Lahann J: Surface engineering the cellular microenvironment via patterning and gradients. J Polym Sci Pol Phys 2013, 51: 775-794. 10.1002/polb.23275View ArticleGoogle Scholar
- Folch A, Toner M: Microengineering of cellular interactions. Annu Rev Biomed Eng 2000, 2: 227-256. 10.1146/annurev.bioeng.2.1.227View ArticleGoogle Scholar
- Weibel DB, DiLuzio WR, Whitesides GM: Microfabrication meets microbiology. Nat Rev Microbiol 2007, 5: 209-218. 10.1038/nrmicro1616View ArticleGoogle Scholar
- Ross AM, Jiang ZX, Bastmeyer M, Lahann J: Physical aspects of cell culture substrates: topography, roughness, and elasticity. Small 2012, 8: 336-355. 10.1002/smll.201100934View ArticleGoogle Scholar
- Chen W, Lam RHW, Fu J: Photolithographic surface micromachining of polydimethylsiloxane (PDMS). Lab Chip 2012, 12: 391-395. 10.1039/c1lc20721kView ArticleGoogle Scholar
- Sugiura S, Edahiro JI, Sumaru K, Kanamori T: Surface modification of polydimethylsiloxane with photo-grafted poly (ethylene glycol) for micropatterned protein adsorption and cell adhesion. Colloid Surf B 2008, 63: 301-305. 10.1016/j.colsurfb.2007.12.013View ArticleGoogle Scholar
- Lee JN, Jiang X, Ryan D, Whitesides GM: Compatibility of mammalian cells on surfaces of poly (dimethylsiloxane). Langmuir 2004, 20: 11684-11691. 10.1021/la048562+View ArticleGoogle Scholar
- Park JY, Ahn D, Choi YY, Hwang CM, Takayama S, Lee SH, Lee S-H: Surface chemistry modification of PDMS elastomers with boiling water improves cellular adhesion. Sensor Actuat B Chem 2012, 173: 765-771.View ArticleGoogle Scholar
- Wu MH: Simple poly (dimethylsiloxane) surface modification to control cell adhesion. Surf Interface Anal 2009, 41: 11-16. 10.1002/sia.2964View ArticleGoogle Scholar
- Zhang WJ, Choi DS, Nguyen YH, Chang J, Qin LD: Studying cancer stem cell dynamics on PDMS surfaces for microfluidics device design. Sci Rep 2013, 3: 2322.Google Scholar
- Sugaya S, Kakegawa S, Fukushima S, Yamada M, Seki M: Micropatterning of hydrogels on locally hydrophilized regions on PDMS by stepwise solution dipping and in situ gelation. Langmuir 2012, 28: 14073-14080. 10.1021/la3014706View ArticleGoogle Scholar
- Beduer A, Vieu C, Arnauduc F, Sol J-C, Loubinoux I, Vaysse L: Engineering of adult human neural stem cells differentiation through surface micropatterning. Biomaterials 2011, 33: 504-514.View ArticleGoogle Scholar
- Bodas D, Khan-Malek C: Formation of more stable hydrophilic surfaces of PDMS by plasma and chemical treatments. Microelectron Eng 2006, 83: 1277-1279. 10.1016/j.mee.2006.01.195View ArticleGoogle Scholar
- de Silva M, Desai R, Odde D: Micro-patterning of animal cells on PDMS substrates in the presence of serum without use of adhesion inhibitors. Biomed Microdevices 2004, 6: 219-222.View ArticleGoogle Scholar
- Kane RS, Takayama S, Ostuni E, Ingber DE, Whitesides GM: Patterning proteins and cells using soft lithography. Biomaterials 1999, 20: 2363-2376. 10.1016/S0142-9612(99)00165-9View ArticleGoogle Scholar
- Kang K, Kang G, Lee BS, Choi IS, Nam Y: Generation of patterned neuronal networks on cell-repellant poly (oligo(ethylene glycol) methacrylate) films. Chem Asian J 2010, 5: 1804-1809. 10.1002/asia.200900761View ArticleGoogle Scholar
- Rogers CI, Pagaduan JV, Nordin GP, Woolley AT: Single-monomer formulation of polymerized polyethylene glycol diacrylate as a nonadsorptive material for microfluidics. Anal Chem 2011, 83: 6418-6425. 10.1021/ac201539hView ArticleGoogle Scholar
- Harris JM, Zalipsky S: Poly (ethylene glycol). Washington DC: American Chemical Society; 1997.View ArticleGoogle Scholar
- Koh W-G, Revzin A, Simonian A, Reeves T, Pishko M: Control of mammalian cell and bacteria adhesion on substrates micropatterned with poly (ethylene glycol) hydrogels. Biomed Microdevices 2003, 5: 11-19. 10.1023/A:1024455114745View ArticleGoogle Scholar
- Revzin A, Tompkins RG, Toner M: Surface engineering with poly (ethylene glycol) photolithography to create high-density cell arrays on glass. Langmuir 2003, 19: 9855-9862. 10.1021/la035129bView ArticleGoogle Scholar
- Hu SW, Ren XQ, Bachman M, Sims CE, Li GP, Allbritton N: Surface modification of poly (dimethylsiloxane) microfluidic devices by ultraviolet polymer grafting. Anal Chem 2002, 74: 4117-4123. 10.1021/ac025700wView ArticleGoogle Scholar
- Patrito N, McCague C, Norton PR, Petersen NO: Spatially controlled cell adhesion via micropatterned surface modification of poly (dimethylsiloxane). Langmuir 2007, 23: 715-719. 10.1021/la062007lView ArticleGoogle Scholar
- Stevens MM, George JH: Exploring and engineering the cell surface interface. Science 2005, 310: 1135-1138. 10.1126/science.1106587View ArticleGoogle Scholar
- Eroshenko N, Ramachandran R, Yadavalli VK, Rao RR: Effect of substrate stiffness on early human embryonic stem cell differentiation. J Biol Eng 2013, 7: 7. 10.1186/1754-1611-7-7View ArticleGoogle Scholar
- Engler AJ, Sen S, Sweeney HL, Discher DE: Matrix elasticity directs stem cell lineage specification. Cell 2006, 126: 677-689. 10.1016/j.cell.2006.06.044View ArticleGoogle Scholar
- Seo J-H, Sakai K, Yui N: Adsorption state of fibronectin on poly (dimethylsiloxane) surfaces with varied stiffness can dominate adhesion density of fibroblasts. Acta Biomater 2013, 9: 5493-5501. 10.1016/j.actbio.2012.10.015View ArticleGoogle Scholar
- Almutairi Z, Ren CL, Simon L: Evaluation of polydimethylsiloxane (PDMS) surface modification approaches for microfluidic applications. Colloid Surface A 2012, 415: 406-412.View ArticleGoogle Scholar
- Revzin A, Russell RJ, Yadavalli VK, Koh W-G, Deister C, Hile DD, Mellott MB, Pishko MV: Fabrication of poly (ethylene glycol) hydrogel microstructures using photolithography. Langmuir 2001, 17: 5440-5447. 10.1021/la010075wView ArticleGoogle Scholar
- Whitesides GM, Ostuni E, Takayama S, Jiang X, Ingber DE: Soft lithography in biology and biochemistry. Ann Rev Biomed Eng 2001, 3: 335-373. 10.1146/annurev.bioeng.3.1.335View ArticleGoogle Scholar
- Ruiz SA, Chen CS: Microcontact printing: a tool to pattern. Soft Matter 2007, 3: 168-177. 10.1039/b613349eView ArticleGoogle Scholar
- Wang YL, Lai HH, Bachman M, Sims CE, Li GP, Allbritton NL: Covalent micropatterning of poly (dimethylsiloxane) by photografting through a mask. Anal Chem 2005, 77: 7539-7546. 10.1021/ac0509915View ArticleGoogle Scholar
- Jothimuthu P, Carroll A, Bhagat AAS, Lin G, Mark JE, Papautsky I: Photodefinable PDMS thin films for microfabrication applications. J Micromech Microeng 2009, 19: 045024. 10.1088/0960-1317/19/4/045024View ArticleGoogle Scholar
- Schlapak R, Pammer P, Armitage D, Zhu R, Hinterdorfer P, Vaupel M, Frühwirth T, Howorka S: Glass surfaces grafted with high-density poly (ethylene glycol) as substrates for DNA oligonucleotide microarrays. Langmuir 2005, 22: 277-285.View ArticleGoogle Scholar
- Anadon A: Toxicological evaluation of benzophenone. EFSA J 2009, 1104: 1-30.Google Scholar
- Bhagat AAS, Jothimuthu P, Papautsky I: Photodefinable polydimethylsiloxane (PDMS) for rapid lab-on-a-chip prototyping. Lab Chip 2007, 7: 1192-1197. 10.1039/b704946cView ArticleGoogle Scholar
- Leipzig ND, Shoichet MS: The effect of substrate stiffness on adult neural stem cell behavior. Biomaterials 2009, 30: 6867-6878. 10.1016/j.biomaterials.2009.09.002View ArticleGoogle Scholar
- Brown XQ, Ookawa K, Wong JY: Evaluation of polydimethylsiloxane scaffolds with physiologically-relevant elastic moduli: interplay of substrate mechanics and surface chemistry effects on vascular smooth muscle cell response. Biomaterials 2005, 26: 3123-3129. 10.1016/j.biomaterials.2004.08.009View ArticleGoogle Scholar
- Mata A, Fleischman AJ, Roy S: Characterization of polydimethylsiloxane (PDMS) properties for biomedical micro/nanosystems. Biomed Microdevices 2005, 7: 281-293. 10.1007/s10544-005-6070-2View ArticleGoogle Scholar
- Carrillo F, Gupta S, Balooch M, Marshall SJ, Marshall GW, Pruitt L, Puttlitz CM: Nanoindentation of polydimethylsiloxane elastomers: effect of crosslinking, work of adhesion, and fluid environment on elastic modulus. J Mater Res 2006, 21: 535-537. 10.1557/jmr.2005.0354eView ArticleGoogle Scholar
- Lo CM, Wang HB, Dembo M, Wang YL: Cell movement is guided by the rigidity of the substrate. Biophys J 2000, 79: 144-152. 10.1016/S0006-3495(00)76279-5View ArticleGoogle Scholar
- Decker C: Photoinitiated crosslinking polymerization. Prog Polym Sci 1996, 21: 593-650. 10.1016/0079-6700(95)00027-5View ArticleGoogle Scholar
- Abraham S, Riggs MJ, Nelson K, Lee V, Rao RR: Characterization of human fibroblast-derived extracellular matrix components for human pluripotent stem cell propagation. Acta Biomater 2010, 6: 4622-4633. 10.1016/j.actbio.2010.07.029View ArticleGoogle Scholar
- Hutter JL, Bechhoefer J: Calibration of atomic-force microscope tips. Rev Sci Instrum 1993, 64: 1868-1873. 10.1063/1.1143970View ArticleGoogle Scholar
- Oliver WC, Pharr GM: Measurement of hardness and elastic modulus by instrumented indentation: advances in understanding and refinements to methodology. J Mater Res 2004, 19: 3-20. 10.1557/jmr.2004.19.1.3View ArticleGoogle Scholar
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