Carbon-nanotube reinforcement of DNA-silica nanocomposites yields programmable and cell-instructive biocoatings.

Biomedical applications require substrata that allow for the grafting, colonization and control of eukaryotic cells. Currently available materials are often limited by insufficient possibilities for the integration of biological functions and means for tuning the mechanical properties. We report on tailorable nanocomposite materials in which silica nanoparticles are interwoven with carbon nanotubes by DNA polymerization. The modular, well controllable and scalable synthesis yields materials whose composition can be gradually adjusted to produce synergistic, non-linear mechanical stiffness and viscosity properties. The materials were exploited as substrata that outperform conventional culture surfaces in the ability to control cellular adhesion, proliferation and transmigration through the hydrogel matrix. The composite materials also enable the construction of layered cell architectures, the expansion of embryonic stem cells by simplified cultivation methods and the on-demand release of uniformly sized stem cell spheroids.

Biopebble Containers: DNA-Directed Surface Assembly of Mesoporous Silica Nanoparticles for Cell Studies.

The development of methods for colloidal self-assembly on solid surfaces is important for many applications in biomedical sciences. Toward this goal, described is a versatile class of mesoporous silica nanoparticles (MSN) that contain on their surface various types of DNA molecules to enable their self-assembly into micropatterned surface architectures useful for cell studies. Monodisperse dye-doped MSN are synthesized by biphase stratification and functionalized with an aptamer oligonucleotide that serves as gatekeeper for the triggered release of encapsulated molecular cargo, such as fluorescent dye rhodamine B or the anticancer drug doxorubicin. One or two additional types of oligonucleotides are installed on the MSN surface to enable DNA-directed immobilization on solid substrates bearing patterns of complementary capture oligonucleotides. It is demonstrated that this strategy can be used for efficient self-assembly of microstructured surface architectures, which not only promote the adhesion and guidance of cells but also are capable of affecting the fate of adhered cells through triggered release of their cargo. It is believed that this approach is useful for diverse applications in tissue engineering and nanobio sciences.

Multiscale Origami Structures as Interface for Cells.

A DNA-based platform was developed to address fundamental aspects of early stages of cell signaling in living cells. By site-directed sorting of differently encoded, protein-decorated DNA origami structures on DNA microarrays, we combine the advantages of the bottom-up self-assembly of protein-DNA nanostructures and top-down micropatterning of solid surfaces to create multiscale origami structures as interface for cells (MOSAIC). In a proof-of-principle, we use this technology to analyze the activation of epidermal growth factor (EGF) receptors in living MCF7 cells using DNA origami structures decorated on their surface with distinctive nanoscale arrangements of EGF ligand entities. MOSAIC holds the potential to present to adhered cells well-defined arrangements of ligands with full control over their number, stoichiometry, and precise nanoscale orientation. It therefore promises novel applications in the life sciences, which cannot be tackled by conventional technologies.

Patterning of polymeric cell culture substrates.

The purpose of this chapter is to provide a summary of polymer patterning technologies for biological applications and detailed instructions for resist-free deep ultraviolet (UV) patterning of poly(styrene). Photochemical modifications of this polymer yield unstable peroxides together with stable oxidized chemical groups. The altered physicochemical properties of the polymer surface influence protein adsorption and cell adhesion. HepG2 (human hepatoma cell line), fibroblasts (L929, murine fibroblast line), and other cell lines exhibit strong adhesion on areas of UV-irradiated polymer. Masked irradiations open a simple, fast (cell patterns are obtained within a few hours), and economical route to obtain chemically patterned cell culture substrates. The described protocol is advantageous compared to silane-based patterning techniques on glass or thiol-based patterning on gold because of the elimination of any chemical treatment and the small size of achieved structures. The protocol is compatible with common clean room technologies; however, even without access to a clean room, structured substrates can be produced. The described technique can be a useful tool for a variety of cell cultures used to study biological processes like intercellular communication and organogenesis and for applications like biosensing or tissue engineering.

Selective immobilization of Sonic hedgehog on benzylguanine terminated patterned self-assembled monolayers.

Patterned two-component, self-assembled monolayers on gold were produced by UV lithography. An oligo(ethylene glycol) terminated disulfide served as inert matrix reducing unspecific protein adsorption and cell adhesion. The second component of the self-assembled monolayer (SAM) presented a benzylguanine moiety for the immobilization of Sonic hedgehog (Shh) fused to a mutant O(6)-alkylguanine-DNA alkyltransferase (SNAP-tag™). The enzymatic activity of the SNAP-tag allows selective and covalent immobilization of the linked Shh. Time-of-flight secondary ion mass spectrometry verified the correct lateral distribution of the benzylguanine head groups in the patterned SAM. The quantification of unspecific and specific protein binding to mixed SAMs showed increased adsorption of albumin with increasing benzylguanine/(ethylene glycol) ratios. However, the immobilization of SNAP-tagged Shh was not blocked by pre-adsorbed albumin. Furthermore, the obtained micro-patterned substrates permitted direct immobilization of SNAP-tagged Shh even in the presence of many competing proteins from conditioned media of transfected HEK293 cells. Therefore, the presented system is suited for the controlled immobilization of fusion proteins from complex mixtures avoiding purification steps.