RESUMO
Three-dimensional printing at the micro-/nanoscale represents a new challenge in research and development to achieve direct printing down to nanometre-sized objects. Here, FluidFM, a combination of microfluidics with atomic force microscopy, offers attractive options to fabricate hierarchical polymer structures at different scales. However, little is known about the effect of the substrate on the printed structures and the integration of (bio)functional groups into the polymer inks. In this study, we printed micro-/nanostructures on surfaces with different wetting properties, and integrated molecules with different functional groups (rhodamine as a fluorescent label and biotin as a binding tag for proteins) into the base polymer ink. The substrate wetting properties strongly affected the printing results, in that the lateral feature sizes increased with increasing substrate hydrophilicity. Overall, ink modification only caused minor changes in the stiffness of the printed structures. This shows the generality of the approach, as significant changes in the mechanical properties on chemical functionalization could be confounders in bioapplications. The retained functionality of the obtained structures after UV curing was demonstrated by selective binding of streptavidin to the printed structures. The ability to incorporate binding tags to achieve specific interactions between relevant proteins and the fabricated micro-/nanostructures, without compromising the mechanical properties, paves a way for numerous bio and sensing applications. Additional flexibility is obtained by tuning the substrate properties for feature size control, and the option to obtain functionalized printed structures without post-processing procedures will contribute to the development of 3D printing for biological applications, using FluidFM and similar dispensing techniques.
RESUMO
The curvature of lipid membranes plays a key role in many relevant biological processes such as membrane trafficking, vesicular budding and host-virus interactions. In vitro studies on the membrane curvature of simplified biomimetic models in the nanometer range are challenging, due to their complicated nanofabrication processes. In this work, we propose a simple and low-cost platform for curvature sensitive protein screening, prepared through scanning probe lithography (SPL) methods, where lipid bilayer patches of different compositions can be multiplexed onto substrate areas with tailored local curvature. The curvature is imposed by anchoring nanoparticles of the desired size to the substrate prior to lithography. As a proof of principle, we demonstrate that a positive curvature membrane sensitive protein derived from the BAR domain of Nadrin2 binds selectively to lipid patches patterned on substrate areas coated with 100 nm nanoparticles. The platform opens up a path for screening curvature-dependent protein-membrane interaction studies by providing a flexible and easy to prepare substrate with control over lipid composition and membrane curvature.
