UC Berkeley

3D Bioprinting is a popular method of fabricating scaffolds for tissue engineering. While soft bioinks such as alginate are useful for reproducing the softest tissues of the human body, it remains difficult to 3D print objects out of bioinks that cannot support their own weight. 3D cryoprinting is a promising method of printing objects out of soft bioinks and creating highly porous scaffolds for tissue engineering. However, crosslinking the frozen objects before they thaw and collapse remains a challenge. Here we investigate a process which we name “freezing-modulated-crosslinking” for crosslinking frozen objects produced by 3D cryoprinting. During freezing-modulated-crosslinking, frozen objects are thawed in a crosslinker bath at a controlled melting rate, so that crosslinking occurs layer by layer and the object can maintain its printed shape. First, we examine the process with a mathematical model to determine the important thermal parameters. Second, we validate our results experimentally by printing a variety of multi-layer alginate objects. By systematically examining this crosslinking approach, we expand the options for 3D cryoprinting. 3D cryoprinted alginate scaffolds can be seeded with cells for use in 3D cell culture or for tissue regeneration.