Interaction of partially denatured insulin with a DSPC floating lipid bilayer.

The carefully controlled permeability of cellular membranes to biological molecules is key to life. In degenerative diseases associated with protein misfolding and aggregation, protein molecules or their aggregates are believed to permeate these barriers and threaten membrane integrity. We used neutron reflectivity to study the interaction of insulin, a model amyloidogenic protein, with a DSPC floating lipid bilayer. Structural changes consistent with protein partitioning to the membrane interior and adsorption to a gel phase model lipid bilayer were observed under conditions where the native fold of the protein is significantly destabilised. We propose that the perturbation of the membrane by misfolded proteins involves long term occupation of the membrane by these proteins, rather than transient perforation events.

Quantitative evaluation of evaporation rate during spin-coating of polymer blend films: Control of film structure through defined-atmosphere solvent-casting.

Thin films of polymer mixtures made by spin-coating can phase separate in two ways: by forming lateral domains, or by separating into distinct layers. The latter situation (self-stratification or vertical phase separation) could be advantageous in a number of practical applications, such as polymer optoelectronics. We demonstrate that, by controlling the evaporation rate during the spin-coating process, we can obtain either self-stratification or lateral phase separation in the same system, and we relate this to a previously hypothesised mechanism for phase separation during spin-coating in thin films, according to which a transient wetting layer breaks up due to a Marangoni-type instability driven by a concentration gradient of solvent within the drying film. Our results show that rapid evaporation leads to a laterally phase-separated structure, while reducing the evaporation rate suppresses the interfacial instability and leads to a self-stratified final film.

A solution concentration dependent transition from self-stratification to lateral phase separation in spin-cast PS:d-PMMA thin films.

Thin films with a rich variety of different nano-scale morphologies have been produced by spin casting solutions of various concentrations of PS:d-PMMA blends from toluene solutions. During the spin casting process specular reflectivity and off-specular scattering data were recorded and ex situ optical and atomic force microscopy, neutron reflectivity and ellipsometry have all been used to characterise the film morphologies. We show that it is possible to selectively control the film morphology by altering the solution concentration used. Low polymer concentration solutions favour the formation of flat in-plane phase-separated bi-layers, with a d-PMMA-rich layer underneath a PS-rich layer. At intermediate concentrations the films formed consist of an in-plane phase-separated bi-layer with an undulating interface and also have some secondary phase-separated pockets rich in d-PMMA in the PS-rich layer and vice versa. Using high concentration solutions results in laterally phase-separated regions with sharp interfaces. As with the intermediate concentrations, secondary phase separation was also observed, especially at the top surface.

The efficiency of encapsulation within surface rehydrated polymersomes.

The key to the use of polymersomes as effective molecular delivery systems is in the ability to design processing routes that can efficiently encapsulate the molecular payload. We have evaluated various surface rehydration mechanisms for encapsulation, in each case characterizing the morphologies formed using DLS and confocal microscopy as well as determining the encapsulation efficiency for the hydrophilic dye Rhodamine B. In contrast to bulk methods, where the encapsulation efficiencies are low, we find that higher efficiencies can be obtained by the rehydration of thin films. We relate these results to the non-equilibrium mechanisms that underlie vesicle formation and discuss how an understanding of these mechanisms can help optimize encapsulation efficiencies. Our conclusion is that, even considering the good encapsulation efficiency, surface methods are still unsuitable for the massive scale-up needed when applied to commercial "mass market" molecular delivery scenarios. However, targeting more specialized applications for high value ingredients (like pharmaceuticals) might be more feasible.

Early-stage roughening of the polymer-polymer interface approaching the glass transition temperature by real-time neutron reflection.

The early-stage roughening of the interface between thin deuterated poly(methyl methacrylate) (d-PMMA) layers on thick polystyrene (PS) films was studied as a function of the temperature using real-time specular neutron reflectivity. By measuring the growth of the interface roughness as a precursor of the dewetting, the characteristic time constant of the early stages of the process was studied as a function of the temperature approaching the glass transition temperature (T(g)) of the two polymers from above and compared with the prediction of the growth of the interface by the spinodal process. Both solid and liquid regimes were probed, in which the viscosity of the thin film or the substrate dominates respectively. The characteristic growth time of the process also depends on the upper film thickness to a power of 5 or 6 in the solid or liquid regimes, respectively, as predicted by the theory of spinodal dewetting.

Measurement of adhesion energies and Young's modulus in thin polymer films using a novel axi-symmetric peel test geometry.

We present a novel method of probing adhesion energies of solids, particularly polymers. This method uses the axi-symmetric deformation of a thin spincast polymer membrane brought into contact with a flat substrate to probe the work of adhesion. The use of a thin membrane minimizes uncertainty in the radius of contact, while the use of spincast films provides very smooth surfaces by means of a very simple method. The experimental profile of the deformed membrane shows good agreement with the expected logarithmic profile. The experimental setup enables the measurement of Young's modulus and the solid-solid work of adhesion for thin films. The value obtained for Young's modulus of polystyrene (PS) was found to be in agreement with other conventional measurement techniques. In addition, measurement of the work of adhesion at the PS/silicon oxide interface was possible. The apparatus is well suited to studying the dependence of Young's modulus, work of adhesion and fracture energy on membrane thickness, temperature, pulling rate, and ageing of the interface, and can readily be modified to study biologically relevant samples.

Mapping the fluorescence decay lifetime of a conjugated polymer in a phase-separated blend using a scanning near-field optical microscope.

We have studied a blend of the polymers poly(9,9-dioctylfluorene) (F8) dispersed in an inert matrix of polystyrene (PS) using time-resolved scanning near-field optical microscopy (SNOM). On spin-casting, phase separation occurs between the two polymers resulting in a thin film characterized by an F8-rich phase and a PS-rich phase. By spatially mapping the intensity of photoluminescence from the film, we find that there is a low concentration of F8 trapped within the PS-rich phase. We find that the fluorescence emission lifetime (measured at 440 nm) of F8 trapped within the PS-rich phase is significantly longer than that from the F8-rich phase (290 ps compared to 235 ps). Furthermore, spectral measurements indicate that the F8 emission from the PS-rich phase is characterized by a reduced fraction of emission from fluorenone defect states. Taken together, our measurements suggest that in the PS-rich phase interchain exciton diffusion between F8 molecules is suppressed significantly by the effect of dilution.

Approaching criticality in polymer-polymer systems.

The interfacial width of polyolefins blends has been probed as a function of distance away from the critical point by using neutron reflectivity. For strongly immiscible polymer pairs, the width of the interface increases slowly when the degree of immiscibility is decreased and the interfacial width varies with the interaction parameter chi of the polymers. Closer to the critical point the dependence on the degree of miscibility becomes stronger and the way in which the interfacial width diverges, as criticality is approached, is related to both the chain length and chi. The self-consistent field theory numerical calculations, with the additional contribution due to capillary waves, provides a good description of the width of the interface between two polymer bulk phases in particular at intermediate values of the degree of immiscibility.

Early stages of polymer interdiffusion.

We have examined the wavelength dependence of the diffusion coefficient in a multilayer deuterated polystyrene-hydrogenated polystyrene system. The measurements were performed using neutron reflectivity. The decay of the Bragg peak was also followed in great detail. The measured wavelength diffusion coefficient D(k) was also not found spatially dependent on the distance of the diffusional couples to the substrate. On the basis of our results, it was possible to reconstruct the broadening interface of polymer bilayer sample geometry which had characteristics of a diffuse tail and a sharp core.

Effect of physical ageing in thin glassy polymer films.

We have studied the effect of physical ageing in thin supported glassy polystyrene films by using ellipsometry to detect overshooting in the expansivity-temperature curve upon heating of aged samples. Films with thickness 10-200 nm have been aged at 70(degrees) C and 80(degrees) C (below the bulk glass transition temperature). We observe clear relaxation peaks in the expansivity-temperature curve for films thicker than 18 nm but not for the 10 nm film. The intensity of the relaxation peak is inversely proportional to the film thickness, while the temperatures characteristic to the relaxation peak are almost independent of the film thickness. These observations are successfully interpreted by the idea that the surface layer of the order of 10 nm has liquid-like thermal properties.

Surface denaturation and amyloid fibril formation of insulin at model lipid-water interfaces.

We consider the effects that different lipid surfaces have upon the denaturation and subsequent formation of amyloid fibrils of bovine insulin. The adsorption and unfolding kinetics of insulin being adsorbed onto the different lipid surfaces under denaturing conditions are studied using FTIR ATR spectroscopy and are compared to the bulk solution behavior of the protein. Atomic force microscopy studies are also performed to compare the fibrils growing on the different surfaces. This study shows that both the adsorption and unfolding kinetics of insulin can be described by a sum of exponential processes and that different surfaces behave differently, with respect both to one another and to the bulk protein solution. The proteins adsorbed onto the surfaces are observed to have faster unfolding kinetics than those in the bulk, and the fibril-like structures formed at the surfaces are shown to be different in a number of ways from those found in bulk solution. The beta-sheet content and growth kinetics of the adsorbed proteins also differ from those of the bulk system. An attempt is made to describe the observed behavior in terms of simple physical arguments involving adsorption, unfolding, and aggregation of the proteins.

Swelling-induced morphology in ultrathin supported films of poly(d,l-lactide).

In this study we describe a surface morphology that arises when ultrathin supported films of poly(d,l-lactide) are immersed in water. The films are initially flat with a rms roughness of approximately 2 nm. After immersion the surfaces of the films are covered with craters. The craters have a narrow distribution of sizes and are typically micrometers in diameter. They have depths in the 10-100 nm range. In situ atomic force microscopy shows that the craters occur as a result of a blistering process, which occurs when the films delaminate from the silicon substrate. The films buckle away from the substrate to give a nonzero initial diameter and then the blisters proceed to grow until they reach a maximum size. At any point during the growth process, the blisters can be made to collapse by removing the films from water. This phenomenon is explained in terms of a laterally confined swelling film, which has a buckling instability and releases excess strain energy by wrinkling. An expression for the initial buckling wavelength is extracted using the expressions for a buckling plate. Information about the mechanical properties of the films and the surface interaction between the film and substrate can also be obtained by considering the kinetics of blister growth.

The timescale of spinodal dewetting at a polymer/polymer interface.

We investigate the dynamics of spinodal dewetting in liquid-liquid polymer systems. Dewetting of poly(methyl-methacrylate) (PMMA) thin films on polystyrene (PS) "substrates" is followed in situ using neutron reflectivity. By following the development of roughness at the PS/PMMA interface and the PMMA surface we extract characteristic growth times for the dewetting process. These characteristic growth times are measured as a function of the molecular weight of the two polymers. By also carrying out experiments in the regime where the dynamics are independent of the PS molecular weight, we are able to use dewetting to probe the scaling of the PMMA thin film viscosity with temperature and molecular weight. We find that this scaling reflects bulk behaviour. However, absolute values are low compared to bulk viscosities, which we suggest may be due in part to slippage at the polymer/polymer interface.