In humans, damage caused by injuries to the heart result in the permanent loss of myocardial tissue. Lost myocardium is instead replaced with non-functional scar tissue that causes heart failure and death. This lack of healing results in the high morbidity and mortality seen in heart disease in the US and around the world. The goal of my work was to develop a novel material that could improve wound healing in the hearts of adult mammals, with the goal of scarless healing that restores lost myocardium. To do this, I looked to natural examples of cardiac regeneration. Some animals exhibit cardiac regeneration after injury, and the components of the extracellular matrix of these species have been shown to stimulate regeneration in non-regenerative species. The first portion of my work focused on identifying a larger and more scalable source of regenerative extracellular matrix than those already characterized. I showed that goldfish regenerate resection injuries in the hearts by 70 days post injury in a scarless manner. To do so, they induce higher levels of cardiomyocyte proliferation, which peaks at about 14 days post injury. Additionally, they increase the expression of extracellular matrix components known to stimulate cardiac regeneration in other species, with this peak coinciding with peak cardiomyocyte proliferation. The second portion of my work focused on evaluating the ability of goldfish cardiac extracellular matrix to stimulate a pro-regenerative phenotype in mammalian cells relevant to the cardiac wound response. I found that injured goldfish cardiac extracellular matrix acts in an oxygen concentration dependent manner on cardiac fibroblasts, macrophages, and neonatal cardiomyocytes. The cell behaviors and phenotypes I saw resembled some of the responses seen in regenerative species, including fibroblast inactivation as well as different macrophage polarization and cardiomyocyte proliferation under hypoxia. In sum, my work identified and validated goldfish cardiac extracellular matrix as a potential source of a regenerative biomaterial, though more work is needed to evaluate its effects in vivo.