Spatially controlled cell adhesion via micropatterned surface modification of poly(dimethylsiloxane)

Spatial control of cell growth on surfaces can be achieved by the selective deposition of molecules that influence cell adhesion. The fabrication of such substrates often relies upon photolithography and requires complex surface chemistry to anchor adhesive and inhibitory molecules. The production of simple, cost-effective substrates for cell patterning would benefit numerous areas of bioanalytical research including tissue engineering and biosensor development. Poly(dimethylsiloxane) (PDMS) is routinely used as a biomedical implant material and as a substrate for microfluidic device fabrication; however, the low surface energy and hydrophobic nature of PDMS inhibits its bioactivity. We present a method for the surface modification of PDMS to promote localized cell adhesion and proliferation. Thin metal films are deposited onto PDMS through a physical mask in the presence of a gaseous plasma. This treatment generates topographical and chemical modifications of the polymer surface. Removal of the deposited metal exposes roughened PDMS regions enriched with hydrophilic oxygen-containing species. The morphology and chemical composition of the patterned substrates were assessed by optical and atomic force microscopies as well as X-ray photoelectron spectroscopy. We observed a direct correlation between the surface modification of PDMS and the micropatterned adhesion of fibroblast cells. This simple protocol generates inexpensive, single-component substrates capable of directing cell attachment and growth.     

Adhesion of microchannel-based complementary surfaces

We show that highly enhanced and selective adhesion can be achieved between surfaces patterned with complementary microchannel structures. An elastic material, poly(dimethylsiloxane) (PDMS), was used to fabricate such surfaces by molding into a silicon master with microchannel profiles patterned by photolithography. We carried out adhesion tests on both complementary and mismatched microchannel/micropillar surfaces. Adhesion, as measured by the energy release rate required to propagate an interfacial crack, can be enhanced by up to 40 times by complementary interfaces, compared to a flat control, and slightly enhanced for some special noncomplementary samples, despite the nearly negligible adhesion for other mismatched surfaces. For each complementary surface, we observe defects in the form of visible striations, where pillars fail to insert fully into the channels. The adhesion between complementary microchannel surfaces is enhanced by a combination of a crack-trapping mechanism and friction between a pillar and channel and is attenuated by the presence of defects.     

Contact instability of thin elastic films on patterned substrates

The free surface of a soft elastic film becomes unstable and forms an isotropic labyrinth pattern when a rigid flat plate is brought into adhesive contact with the film. These patterns have a characteristic wavelength, lambda approximately 3H, where H is the film thickness. We show that these random structures can be ordered, modulated, and aligned by depositing the elastic film (cross-linked polydimethylsiloxane) on a patterned substrate and by bringing the free surface of the film in increasing adhesive contact with a flat stamp. Interestingly, the influence of the substrate "bleeds" through the film to its free surface. It becomes possible to generate complex two-dimensional ordered structures such as an array of femtoliter beakers even by using a simple one-dimensional stripe patterned substrate when the instability wavelength, lambda approximately 3H, nearly matches the substrate pattern periodicity. The free surface morphology is modulated in situ by merely varying the stamp-surface separation distance. The free surface structures originating from the elastic contact instability can also be made permanent by the UV-ozone induced oxidation and stiffening.     

Covalent microcontact printing of proteins for cell patterning

We describe a straightforward approach to the covalent immobilization of cytophilic proteins by microcontact printing, which can be used to pattern cells on substrates. Cytophilic proteins are printed in micropatterns on reactive self-assembled monolayers by using imine chemistry. An aldehyde-terminated monolayer on glass or on gold was obtained by the reaction between an amino-terminated monolayer and terephthaldialdehyde. The aldehyde monolayer was employed as a substrate for the direct microcontact printing of bioengineered, collagen-like proteins by using an oxidized poly(dimethylsiloxane) (PDMS) stamp. After immobilization of the proteins into adhesive "islands", the remaining areas were blocked with amino-poly(ethylene glycol), which forms a layer that is resistant to cell adhesion. Human malignant carcinoma (HeLa) cells were seeded and incubated onto the patterned substrate. It was found that these cells adhere to and spread selectively on the protein islands, and avoid the poly(ethylene glycol) (PEG) zones. These findings illustrate the importance of microcontact printing as a method for positioning proteins at surfaces and demonstrate the scope of controlled surface chemistry to direct cell adhesion.     

The effect of surface microtopography of poly(dimethylsiloxane) on protein adsorption, platelet and cell adhesion

Chemical homogeneous poly(dimethylsiloxane) (PDMS) surface with dot-like protrusion pattern was used to investigate the individual effect of surface microtopography on protein adsorption and subsequent biological responses. Fibrinogen (Fg) and fibronectin (Fn) were chosen as model proteins due to their effect on platelet and cell adhesion, respectively. Fg labeled with ¹²⁵I and fluorescein isothiocyanate (FITC) was used to study its adsorption on flat and patterned surfaces. Patterned surface has a 46% increase in the adsorption of Fg when compared with flat surface. However, the surface area of the patterned surface was only 8% larger than that of the flat surface. Therefore, the increase in the surface area was not the only factor responsible for the increase in protein adsorption. Clear fluorescent pattern was visualized on patterned surface, indicating that adsorbed Fg regularly distributed and adsorbed most on the flanks and valleys of the protrusions. Such distribution and local enrichment of Fg presumably caused the specific location of platelets adhered from platelet-rich plasma (PRP) and flowing whole blood (FWB) on patterned surface. Furthermore, the different combination of surface topography and pre-adsorbed Fn could influence the adhesion of L929 cells. The flat surface with pre-adsorbed Fn was the optimum substrate while the virgin patterned surface was the poor substrate in terms of L929 cells spread.     

Probing the Adhesion of Hepatocellular Carcinoma HepG2 and SK‐Hep‐1 Cells

Carcinoma cell differentiation stage is an important indicator of cell behavior. For example, cell mobility is much higher for poorly‐differentiated hepatocellular carcinoma SK‐Hep‐1 cells than for well‐differentiated HepG2 cells. In this study, we have cultured HepG2 and SK‐Hep‐1 cells on chemically patterned polydimethylsiloxane (PDMS) substrates to observe differences in the adhesion properties and cell structure that occur due to the patterns. Both cell lines showed a preference for the hydrophobic regions on the patterned PDMS surface with SK‐Hep‐1 cells achieving a higher density than HepG2 for the same cell‐count solutions. Further, SK‐Hep‐1 cells adopted the square or hexagonal shape of the surface patterns while HepG2 cells maintained their more rounded shape. AFM force measurement arrays were also performed on the cell surfaces to measure and map adhesion values between the tip and cell surface membrane. These results demonstrate that, in addition to cell shape and size, adhesion expression in hepatocellular carcinoma cells is differentiation stage dependent. Further, the ability of the SK‐Hep‐1 cells to adopt the shape of the substrate pattern indicates they are more structurally labile, which may contribute to their higher mobility.     

Surface energy modified chips for detection of conformational states and enzymatic activity in biomolecules

A novel patterning method for anchoring biomolecules and noncovalent assembled conjugated polyelectrolyte (CPE)/biomolecule complexes to a chip surface is presented. The surface energy of a hydrophilic substrate is modified using an elastomeric poly(dimethylsiloxane) (PDMS) stamp, containing a relief pattern. Modification takes place on the parts where the PDMS stamp is in conformal contact with the substrate and leaves low molecular weight PDMS residues on the surface resulting in a hydrophobic modification, and then biomolecules and CPE/biomolecule complexes are then adsorbed in a specific pattern. The method constitutes a discrimination system for different conformations in biomolecules using CPEs as reporters and the PDMS modified substrates as the discriminator. Detection of different conformations in two biomacromolecules, a synthetic peptide (JR2E) and a protein (calmodulin), reported by the CPE and resolved by fluorescence was demonstrated. Also, excellent enzyme activity in patterned CPE/horseradish peroxidase (HRP) enzyme was shown, demonstrating that this method can be used to pattern biomolecules with their activity retained. The method presented could be useful in various biochip applications, such as analyzing proteins and peptides in large-scale production, in making metabolic chips, and for making multi-microarrays.     

Microstructured silicone substrate for printable and stretchable metallic films

Stretchable electronics (i.e., hybrid inorganic or organic circuits integrated on elastomeric substrates) rely on elastic wiring. We present a technique for fabricating reversibly stretchable metallic films by printing silver-based ink onto microstructured silicone substrates. The wetting and pinning of the ink on the elastomer surface is adjusted and optimized by varying the geometry of micropillar arrays patterned on the silicone substrate. The resulting films exhibit high electrical conductivity (～11?000 S/cm) and can stretch reversibly to 20% strain over 1000 times without failing electrically. The stretchability of the ≥200 nm thick metallic film relies on engineered strain relief in the printed film on patterned PDMS.     

Super-hydrophobic, highly adhesive, polydimethylsiloxane (PDMS) surfaces

Super-hydrophobic surfaces have been fabricated by casting polydimethylsiloxane (PDMS) on a textured substrate of known surface topography, and were characterized using contact angle, atomic force microscopy, surface free energy calculations, and adhesion measurements. The resulting PDMS has a micro-textured surface with a static contact angle of 153.5° and a hysteresis of 27° when using de-ionized water. Unlike many super-hydrophobic materials, the textured PDMS is highly adhesive, allowing water drops as large as 25.0 μL to be inverted. This high adhesion, super-hydrophobic behavior is an illustration of the "petal effect". This rapid, reproducible technique has promising applications in transport and analysis of microvolume samples.     

Micro-perforated elastomeric poly(dimethylsiloxane) mask fabricated using high-aspect-ratio micro-pillar arrays for spatially defined surface modification: an unconventional method for establishing a microarray platform

Herein, we introduce the fabrication of a micro-perforated elastomeric poly(dimethylsiloxane) (PDMS) mask and employ it for spatially defined surface modification. To fabricate the micro-perforated PDMS mask, high-aspect-ratio micro-pillar arrays having millimeter scale height were first fabricated via direct photopolymerization using a thiol–ene-based UV-curable adhesive. Square pillars (500 × 500 μm) and 200 μm circular pillars with 5 and 12.5 in the aspect ratios, respectively, were successfully fabricated with high pattern fidelity, reaching 2.5mm in height. Next, using the micro-pillar-array platform as a master mold, PDMS prepolymer was cast and polymerized to form an elastomeric PDMS mask having micro-perforation arrays. Alternating hydrophilic and hydrophobic surfaces were successfully obtained by oxidizing PDMS-covered Si wafer with corona discharge. Spatially defined chemical functionalities obtained by selective oxidation and subsequent silanization were clearly distinguished via colorimetric detection methods employing ninhydrin and toluidine reagents. The micro-perforated elastomeric PDMS mask enables selective modification of a surface without utilizing photoreactive chemicals and a photomask. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

基于水滴模板法的微纳复合超疏水结构制备的研究

利用粒子辅助水滴模板法的实施获得规则蜂窝状图案化多孔结构模板,并进一步利用聚二甲基硅氧烷(PDMS)复制转移技术获得表面具有微米尺寸蜂窝状突起阵列的反向图案化结构.以这种图案化突起结构作为微米尺寸所提供的微米级粗糙度为基础,设计了2种的简单的二次纳米结构的引入过程,最终实现了微米级阵列和纳米级粗糙度的复合.第一种方法借助银镜反应来实现纳米银结构的化学沉积,最终在PDMS阵列表面获得了致密的纳米银颗粒沉积层,并成功获得了表面接触角达166度的超疏水性质.第二种方法利用了聚电解质/二氧化硅粒子层层静电自组装的方法引入纳米结构,结果在仅仅进行了2个组装循环的条件下即可获得超疏水性质的表面复合结构.通过简单的实验设计试图提供一种基于水滴模板法的微纳复合超疏水结构的普适性制备方法.     

In situ estimation of the chemical and mechanical contributions in local adhesion force measurement with AFM: the specific case of polymers

The atomic force microscope (AFM) was used to perform surface force measurements in contact mode to investigate surface properties of model systems at the nanoscale. Two types of model systems were considered. The first one was composed of a rigid substrate (silicon plates) which was chemically modified by molecular self-assembling (SAMs) to display different surface properties (hydroxyl, amine, methyl and ester functional groups). The second system consists of model polymer networks (cross-linked polydimethylsiloxane or PDMS) of variable mechanical properties, whose surfaces were chemically modified with the same groups as before with silicon substrates. The comparison of the force curves obtained from the two model systems shows that the viscoelastic or mechanical contribution dominates in the adhesion on polymer substrates. Finally, a relationship, which expresses the separation energy at a local scale as a function of the energy dissipated within the contact zone, on one hand and the surface properties of the polymer on the other, was proposed.     

Control of the ZnO nanowires nucleation site using microfluidic channels

We report on the growth of uniquely shaped ZnO nanowires with high surface area and patterned over large areas by using a poly(dimethylsiloxane) (PDMS) microfluidic channel technique. The synthesis uses first a patterned seed template fabricated by zinc acetate solution flowing though a microfluidic channel and then growth of ZnO nanowire at the seed using thermal chemical vapor deposition on a silicon substrate. Variations the ZnO nanowire by seed pattern formed within the microfluidic channel were also observed for different substrates and concentrations of the zinc acetate solution. The photocurrent properties of the patterned ZnO nanowires with high surface area, due to their unique shape, were also investigated. These specialized shapes and patterning technique increase the possibility of realizing one-dimensional nanostructure devices such as sensors and optoelectric devices.     

Adhesion of bioinspired micropatterned surfaces: effects of pillar radius, aspect ratio, and preload

Inspired by biological attachment systems, micropatterned elastomeric surfaces with pillars of different heights (between 2.5 and 80 microm) and radii (between 2.5 and 25 microm) were fabricated. Their adhesion properties were systematically tested and compared with flat controls. Micropatterned surfaces with aspect ratios above 0.5 were found to be more compliant than flat surfaces. The adhesion significantly increases with decreasing pillar radius and increasing aspect ratio of the patterned features. A preload dependence of the adhesion force has been identified and demonstrated to be crucial for obtaining adhesives with tunable adherence.     

Instabilities and pattern miniaturization in confined and free elastic-viscous bilayers

We present an analysis of the instabilities engendered by van der Waals forces in bilayer systems composed of a soft elastic film (<10 microm) and a thin (<100 nm) viscous liquid film. We consider two configurations of such systems: (a) Confined bilayers, where the bilayer is sandwiched between two rigid substrates, and (b) free bilayers, where the viscous film is sandwiched between a rigid substrate and the elastic film. Linear stability analysis shows that the time and length scales of the instabilities can be tuned over a very wide range by changing the film thickness and the material properties such as shear modulus, surface tension, and viscosity. In particular, very short wavelengths comparable to the film thickness can be obtained in bilayers, which is in contrast to the instability wavelengths in single viscous and elastic films. It is also shown that the instabilities at the interfaces of the free bilayers are initiated via an in-phase "bending" mode rather than out-of-phase "squeezing" mode. The amplitudes of deformations at both the elastic-air and elastic-viscous interfaces become more similar as the elastic film thickness decreases and its modulus increases. These findings may have potential applications in the self-organized patterning of soft materials.     

Mechanism of adhesion of polyurethane/polymethacrylate simultaneous interpenetrating networks adhesives to polymer substrates

Polyether-based polyurethane/poly (methyl methacrylate-co-ethyleneglycol dimethacrylate) interpenetrating polymer networks [PU/P (MMA–co–EGDMA)-IPNs] were synthesized and used as adhesives to adhere vulcanized natural rubber (NR) and soft polyvinyl chloride (PVC). The structure and morphology of the IPN adhesives in bulk and near the adhesive/substrate interfaces were investigated. A new mechanism of adhesion called conjugate interpenetration of networks across interfaces, which is suitable for IPN adhesives and polymer substrates, was put forward. According to this mechanism, while forming simultaneous interpenetrating networks in the adhesive, the monomers in the IPN adhesive can permeate polymer substrates and polymerize in situ to form gradient IPNs, thereby producing conjugate three-component IPNs near the adhesive/substrate interfaces. It is the conjugate interpenetration of the networks across the interfaces that strengthens interfacial combination remarkably and results in high bond strength of IPN adhesives. © 1994 John Wiley & Sons, Inc.     

The effect of electrostatics on the contact mechanics of adherent phospholipid vesicles

Adhesion of cells on biomaterial surface is resulted from the complex interplay of specific recognitions and colloidal interactions. Thus understanding the role of electrostatic interactions in bioadhesion may help to elucidate the physiochemical basis of cell signaling pathway on therapeutic devices. In this report, high-resolution reflection interference contrast microscopy, cross-polarized light microscopy and contact mechanics modeling are applied to probe the equilibrium adhesion of giant phospholipid vesicles on 3-amino-propyl-triethoxy-silane coated glass. Simultaneously, the effects of vesicle wall thickness, pH, osmotic stress and surface chemistry on the electrostatic interactions at the membrane–substrate interface are evaluated. The results show that both unilamellar vesicles (ULV) and multilamellar vesicles (MLV) strongly adhere on the cationic substrates at neutral pH. In the presence of electrostatic interactions, ULV is slightly deformed on the substrate as the dimension of its adhesive–cohesive zone is only 6–10% higher than the theoretical value of a rigid sphere with the same mid-plane diameter. The variances of contact angle and capillary length at different locations surrounding MLV are ten times higher than those of ULV. The adhesion energy of ULV with mid-plane diameter of 45 and 20 μm is determined as 3.8×10⁻¹² and 8.6×10⁻¹² J/m², respectively, from the truncated sphere model. Moreover, the increase of osmotic stress induces irregular pattern in ULV's adhesion disc and raises the adhesion energy by 10-fold. Finally, the reduction of pH further enhances the electrostatic attractions/repulsions between vesicle surface and cationic or anionic substrates and leads to an increase of adhesion strength.     

A Simple Method for Fabricating Patterned Curvilinear Microstructures in Poly(dimethylsiloxane) by Selective Wetting

The fabrication of patterned microstructures in poly(dimethylsiloxane) (PDMS) is a prerequisite for soft lithography. Herein, curvilinear surface relief microstructures in PDMS are fabricated through a simple three‐stage approach combining microcontact printing (μCP), selective surface wetting/dewetting and replica molding (REM). First, using an original PDMS stamp (first‐generation stamp) with linear relief features, a chemical pattern on gold substrate is generated by μCP using hexadecanethiol (HDT) as an ink. Then, by a dip‐coating process, an ordered polyethylene glycol (PEG) polymer‐dot array forms on the HDT‐patterned gold substrate. Finally, based on a REM process, the PEG‐dot array on gold substrate is used to fabricate a second‐generation PDMS stamp with microcavity array, and the second‐generation PDMS stamp is used to generate third‐generation PDMS stamp with microbump array. These fabricated new‐generation stamps are utilized in μCP and in micromolding in capillaries (MIMIC), allowing the generation of surface micropatterns which cannot be obtained using the original PDMS stamp. The method will be useful in producing new‐generation PDMS stamps, especially for those who want to use soft lithography in their studies but have no access to the microfabrication facilities.     

Microcontact printing of monodiamond nanoparticles: an effective route to patterned diamond structure fabrication

By combining microcontact printing with a nanodiamond seeding technique, a precise micrometer-sized chemical vapor deposition (CVD) diamond pattern have been obtained. On the basis of the guidance of basic theoretical calculations, monodisperse detonation nanodiamonds (DNDs) were chosen as an "ink" material and oxidized poly(dimethylsiloxane) (PDMS) was selected to serve as a stamp because it features a higher interaction energy with the DNDs compared to that of the original PDMS. The adsorption kinetics shows an approximately exponential law with a maximum surface DND density of 3.4 × 10(10) cm(-2) after 20 min. To achieve a high transfer ratio of DNDs from the PDMS stamp to a silicon surface, a thin layer of poly(methyl methacrylate) (PMMA) was spin coated onto the substrates. A microwave plasma chemical vapor deposition system was used to synthesize the CVD diamond on the seeded substrate areas. Precise diamond patterns with a low expansion ratio (3.6%) were successfully prepared after 1.5 h of deposition. Further increases in the deposition time typically lead to a high expansion rate (～0.8 μm/h). The general pattern shape, however, did not show any significant change. Compared with conventional diamond pattern deposition methods, the technique described here offers the advantages of being simple, inexpensive, damage-free, and highly compatible, rendering it attractive for a broad variety of industrial applications.     

Topography printing to locally control wettability

This paper reports a new patterning method, which utilizes NaOH to facilitate the irreversible binding between the PDMS stamp and substrates and subsequent cohesive mechanical failure to transfer the PDMS patterns. Our method shows high substrate tolerance and can be used to "print" various PDMS geometries on a wide range of surfaces, including Si100, glass, gold, polymers, and patterned SU8 photoresist. Using this technique, we are able to locally change the wettability of substrate surfaces by printing well-defined PDMS architectures on the patterned SU8 photoresist. It is possible to generate differential wetting and dewetting properties in microchannels and in the PDMS printed area, respectively.