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The definitive goal of this research is to develop protein‐based scaffolds for use in soft tissue regeneration, particularly in the field of dermal healing. The premise of this investigation was to characterize the mechanical properties of gelatin cross‐linked with microbial transglutaminase (mTGase) and to investigate the cytocompatibility of mTGase cross‐linked gelatin. Dynamic rheological analysis revealed a significant increase in the storage modulus and thermal stability of gelatin after cross‐linking with mTGase. Static, unconfined compression tests showed an increase in Young's modulus of gelatin gels after mTGase cross‐linking. A comparable increase in gel strength was observed with 0.03% mTGase and 0.25% glutaraldehyde cross‐linked gelatin gels. In vitro studies using 3T3 fibroblasts indicated cytotoxicity at a concentration of 0.05% mTGase after 72 h. However, no significant inhibition of cell proliferation …