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Cell Supports & Scaffolds (invited presentation)

M. Soichet, R. Wylie
University of Toronto, Canada, CA

Keywords: hydrogels, tissue engineering


Three-dimensional biomolecule patterned hydrogels provide cellular microenvironments reaching complexities found in native tissues. We are particularly interested the fabrication of complex materials to spatially control stem cell fate towards the creation of tissue analogues. To this end, we have designed a 3D protein patterning system where multiple differentiation factors were simultaneously immobilized within distinct volumes utilizing two-photon chemistry and orthogonal physical binding pairs. Using this system, two differentiation factors for retinal stem progenitor cells were immobilized: 1) ciliary neurotrophic factor (CNTF); and 2) N-terminal sonic hedgehog (SHH). Agarose hydrogels were modified with bromohydroxy-coumarin protected thiols, which upon two-photon excitation are deprotected in defined volumes yielding reactive thiols. Maleimide modified binding proteins, such as maleimide-streptavidin, react with exposed thiols, yielding 3D patterns of covalently immobilized streptavidin in agarose hydrogels. Growth factors are then introduced as fusion proteins with binding domains, such as biotin-CNTF, for complexation and thus 3D immobilization. Similarly, a series of proteins have been immobilized using other high affinity physical binding pairs. By combining multiple binding systems with two-photon patterning, we were able to simultaneously 3D immobilize proteins for the creation biomimetic hydrogels.
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