Drop-On-Drop Multimaterial 3D Bioprinting Realized by Peroxidase-Mediated Cross-Linking

Abstract

A cytocompatible inkjet bioprinting approach that enables the use of a variety of bioinks to produce hydrogels with a wide range of characteristics is developed. Stabilization of bioinks is caused by horseradish peroxidase (HRP)-catalyzed cross-linking consuming hydrogen peroxide (H₂ O₂ ). 3D cell-laden hydrogels are fabricated by the sequential dropping of a bioink containing polymer(s) cross-linkable through the enzymatic reaction and H₂ O₂ onto droplets of another bioink containing the polymer, HRP, and cells. The ≈95% viability of enclosed mouse fibroblasts and subsequent elongation of the cells in a bioprinted hydrogel consisting of gelatin and hyaluronic acid derivatives suggest the high cytocompatibility of the developed printing approach. The existence of numerous polymers, including derivatives of polysaccharides, proteins, and synthetic polymers, cross-linkable through the HRP-catalyzed reaction, means the current approach shows great promise for biofabrication of functional and structurally complex tissues.

Keywords: 3D printer; bioprinting; horseradish peroxidase; hydrogels; ink jet; inkjetting.