Julie Gold

Control of nanoparticle film structure for colloidal lithography

Colloidal lithography utilises nanoparticles’ ability to self-organise on surfaces, which make them suitable as lithographic masks for the production of nano sized surface features. Adsorption under the influence of electrostatic particle–particle interactions results in ordered structures with the particles separated an average distance described by the random sequential adsorption model (RSA). Large areas (cm2) with a high density of nanoparticles can be covered, which is very useful for application areas like biosensors, biomaterials and catalysis, where large numbers of nano sized features are often required. Feature size, shape and spacing can be systematically varied. In this work methods to control the deposition of films consisting of polystyrene particles on flat oxidised titanium surfaces for particle sizes between 20 and 500 nm and coverages of 0–0.45 are demonstrated and discussed. Experimental difficulties encountered were aggregation of particles at high coverage/large particles and a low coverage limit at low ionic strengths. Experimental solutions to overcome these limitations, maintaining the fast parallel processing advantage of colloidal lithography, are presented. They include heating to stabilize initial (i.e. after adsorption) particle arrangements, use of spacer silica particles, and plasma etching to reduce particle sizes.

Optically driven micromachine elements

We report on a proof of principle demonstration of an optically driven micromachine element. Optical angular momentum is transferred from a circularly polarized laser beam to a birefringent particle confined in an optical tweezers trap. The optical torque causes the particle to spin at up to 350 Hz, and this torque is harnessed to drive an optically trapped microfabricated structure. We describe a photolithographic method for producing the microstructures and show how a light driven motor could be used in a micromachine system.

An in vivo study of bone response to implants topographically modified by laser micromachining.

Dental implants topographically modified by laser ablation of periodic arrays of micron-sized craters, were studied in a two-part laboratory investigation. The patterned and control (turned) implants were inserted in rabbit femur and tibia. After 12 weeks the fixation in the bone was evaluated mechanically or by histomorphometry (all threads along the implant and the three best consecutive threads were analysed). In the pilot study no difference was found with respect to bone-to-implant contact and peak removal torque. Significantly more bone was found for the control implants when measuring the bone area inside the threads in the tibia. In the second part of the study, the pattern was improved and significantly more bone-to-implant contact was found for the laser-machined implants. The second part of the study also demonstrated significantly greater peak removal torque values in the tibia with the test implants than the control implants.

Quantitative assessment of the response of primary derived human osteoblasts and macrophages to a range of nanotopography surfaces in a single culture model in vitro

The effect of nanotopography on a range of Ti oxide surfaces was determined. Flat Ti, 3%, 19%, 30% and 43% topography densities of 110 nm high hemispherical protrusions were cultured in contact with primary derived human macrophages and osteoblasts in single culture models. Prior to introduction of the test substrate the phenotype and optimum conditions for in vitro cell culture were established. The cellular response was investigated and quantified by assessments of cytoskeletal development and orientation, viable cell adhesion, cytokine production and release and RT-PCR analysis of osteogenic markers. The tested nanotopographies did not have a statistically significant effect on viable cell adhesion and subsequent cytoskeletal formation. Surface chemistry was the dominant factor as established via incorporation of a tissue culture polystyrene, TCPS, control. The topography surfaces induced a release of chemotactic macrophage activation agents at 1 day in conjunction with stress fibre formation and a subsequent fibronectin network formation. Osteoblasts migrated away from the topography surfaces to the exposed TCPS within the wells during the 7-day period.

Nanostructured model biomaterial surfaces prepared by colloidal lithography

Adsorption of 110 nm negatively charged polystyrene particles on positively charged titanium surfaces was studied for the purpose of making surfaces with controlled nanotopography. Surfaces with 9–26 % saturation coverages of evenly spaced particles were produced by varying the salt concentration in the particle solution (0.01–1 mM NaCl). This is a quick and relatively simple method to make uniform nanostructures over large surface areas (~cm2). The produced surfaces are used for in vitro cell studies of how sub-micrometre topography influences cell adhesion and function.

Design and microstructuring of PDMS surfaces for improved marine biofouling resistance

In this study room temperature vulcanized (RTV) silicone surfaces with designed surface microstructure and well-defined surface chemistry were prepared. Their resistance to marine macrofouling by barnacles Balanus improvisus was tested in field experiments for deducing optimal surface topography dimensions together with a better understanding of macrofouling mechanisms. Polydimethylsiloxane (PDMS) surfaces were microstructured by casting the PDMS pre-polymer on microfabricated molds. The master molds were made by utilizing photolithography and anisotropic etching of monocrystalline silicon wafers. Several iterative casting steps of PDMS and epoxy were used to produce large quantities of microstructured PDMS samples for field studies. The microstructured PDMS surface consisted of arrays of pyramids or riblets creating a surface arithmetic mean roughness ranging from 5 to 17 μm for different microstructure sizes and geometries, as determined by scanning electron microscopy. Chemophysical properties of the microstructured films were investigated by electron spectroscopy for chemical analysis, time-of-flight secondary ion mass spectroscopy and dynamic contact angle measurements. Films were chemically homogeneous down to the submicron level. Hydrophobicity and contact angle hysteresis increased with increased surface roughness. Field tests on the west coast of Sweden revealed that the microstructure containing the largest riblets (profile height 69 μm) reduced the settling of barnacles by 67%, whereas the smallest pyramids had no significant influence on settling compared to smooth PDMS surfaces. The effect of dimensions and geometry of the surface microstructures on the B. improvisus larvae settling is discussed.

Reversible Changes in Cell Morphology due to Cytoskeletal Rearrangements Measured in Real-Time by QCM-D

The mechanical properties and responses of cells to external stimuli (including drugs) are closely connected to important phenomena such as cell spreading, motility, activity, and potentially even differentiation. Here, reversible changes in the viscoelastic properties of surface-attached fibroblasts were induced by the cytoskeleton-perturbing agent cytochalasin D, and studied in real-time by the quartz crystal microbalance with dissipation (QCM-D) technique. QCM-D is a surface sensitive technique that measures changes in (dynamically coupled) mass and viscoelastic properties close to the sensor surface, within a distance into the cell that is usually only a fraction of its size. In this work, QCM-D was combined with light microscopy to study in situ cell attachment and spreading. Overtone-dependent changes of the QCM-D responses (frequency and dissipation shifts) were first recorded, as fibroblast cells attached to protein-coated sensors in a window equipped flow module. Then, as the cell layer had stabilised, morphological changes were induced in the cells by injecting cytochalasin D. This caused changes in the QCM-D signals that were reversible in the sense that they disappeared upon removal of cytochalasin D. These results are compared to other cell QCM-D studies. Our results stress the combination of QCM-D and light microscopy to help interpret QCM-D results obtained in cell assays and thus suggests a direction to develop the QCM-D technique as an even more useful tool for real-time cell studies.

Microfabricated force-sensitive elastic substrates for investigation of mechanical cell–substrate interactions

Mechanical cell–substrate interactions affect many aspects of cellular functions. In order to investigate these interactions, we have microfabricated force-sensitive cell adhesion substrates. A new design of the substrates has been proposed, where force detection is based on monitoring deflection of close-packed standing cantilevers constituting the surface. Such a force-sensitive substrate may be used both for modulating substrates of different elasticity as well as for measuring local mechanical forces from the cell adhesion contacts. The substrates with different cantilever geometry and spatial arrangement have been microfabricated in oxidized silicon wafers using photolithography and deep reactive ion etching. Scanning electron microscopy and atomic force microscopy have been used to measure the dimensions of the cantilevers and to estimate their spring constants. Cell culture experiments on the microfabricated substrates have been performed to test the applicability of the substrates in different experimental setups.

The importance of surface texture for bone integration of screw shaped implants: An in vivo study of implants patterned by photolithography

The aim of the present study was to investigate the influence of different properties inherent in surface topography on the integration of an implant in bone. Using a photolithography technique, a specific surface pattern was produced on the screw flanks of threaded titanium oral implants. Surface topography was qualitatively assessed by scanning electron microscopy (SEM) and a confocal laser scanning profilometer. Quantitative analysis with the confocal laser profilometer derived parameters for surface roughness and surface roughness together with waviness. The chemical composition of the implant surfaces was analyzed by Auger electron spectroscopy. The patterned and control (turned) implants were inserted in New Zealand White rabbits with a healing period of 3 months. Bone fixation was evaluated with resonance frequency analysis (RFA), peak removal torque analysis (RTQ), and by histomorphometry. No statistically significant differences were seen in the fixation, with respect to bone-to-implant contact, between the patterned and control implants.

A method to integrate patterned electrospun fibers with microfluidic systems to generate complex microenvironments for cell culture applications

The properties of a cells microenvironment are one of the main driving forces in cellular fate processes and phenotype expression invivo. The ability to create controlled cell microenvironments invitro becomes increasingly important for studying or controlling phenotype expression in tissue engineering and drug discovery applications. This includes the capability to modify material surface properties within well-defined liquid environments in cell culture systems. One successful approach to mimic extra cellular matrix is with porous electrospun polymer fiber scaffolds, while microfluidic networks have been shown to efficiently generate spatially and temporally defined liquid microenvironments. Here, a method to integrate electrospun fibers with microfluidic networks was developed in order to form complex cell microenvironments with the capability to vary relevant parameters. Spatially defined regions of electrospun fibers of both aligned and random orientation were patterned on glass substrates that were irreversibly bonded to microfluidic networks produced in poly-dimethyl-siloxane. Concentration gradients obtained in the fiber containing channels were characterized experimentally and compared with values obtained by computational fluid dynamic simulations. Velocity and shear stress profiles, as well as vortex formation, were calculated to evaluate the influence of fiber pads on fluidic properties. The suitability of the system to support cell attachment and growth was demonstrated with a fibroblast cell line. The potential of the platform was further verified by a functional investigation of neural stem cell alignment in response to orientation of electrospun fibers versus a microfluidic generated chemoattractant gradient of stromal cell-derived factor 1 alpha. The described method is a competitive strategy to create complex microenvironments invitro that allow detailed studies on the interplay of topography, substrate surface properties, and soluble microenvironment on cellular fate processes.