Photoimmobilization of biomolecules within a 3-dimensional hydrogel matrix

It has been recognized that a three-dimensional cell invasive scaffold that provides both topographical and chemical cues is desirable in regenerative tissue engineering to encourage cell attachment, migration, regrowth and ultimately tissue repair. Carbohydrate hydrogels are attractive for such applications because they are generally biocompatible and able to match the mechanical properties of most soft tissues. Although carbohydrate hydrogels have been previously modified with cell adhesive peptides and proteins, complicated hydrogel matrix activation was required prior to biomolecule coupling and, perhaps more importantly, the overall immobilization yield was low at approximately 1%. In this study, we report the photo-immobilization of a model biomolecule, ovalbumin (OVA), to agarose gel. We describe two methods of modification where the photoactive moiety is coupled to either the protein (i.e. OVA) or the matrix (i.e. agarose) prior to immobilization. We found that the photo-immobilization yield depends on the location of the photoactive moiety. Using photoactive OVA, 1.8% of the OVA initially incorporated into the agarose gel is immobilized; using photoactive agarose, 9.3% of the OVA initially mixed with the agarose is immobilized. The latter is a significant improvement over previous yields and may be useful in attaining our goal of immobilizing a biomolecule gradient for guided tissue regeneration.