This document presents a protocol for inducing cardiac cryoinjury in neonatal mice to study cardiac regeneration. The protocol aims to improve reproducibility over other neonatal mouse heart injury models that have produced variable results. Key steps include using reusable cryoprobes to induce a controlled injury of scalable size without blood loss or open chest manipulation. The cryoinjury model induces scar formation and decreases cardiomyocyte proliferation, mimicking features of pediatric heart disease. It allows direct comparison to regenerating models and testing of interventions to stimulate regeneration. While technical challenges exist, following the structured protocol approach can help researchers successfully learn and apply this technique.