Acute kidney injury (AKI), characterized by a sudden and sustained decline in renal function, is linked to significant morbidity and mortality. The regeneration of the kidney following AKI is a complex process in which the activation of stem and progenitor cells plays a crucial role. Numerous studies have demonstrated that endogenous Sox9+ cells contribute to this regeneration. Traditionally, the status of kidney regeneration after AKI has been evaluated through histopathological examination and renal function indices, which are limited in providing real-time and dynamic insights. To address these limitations, we propose a novel approach using two-photon live imaging to track lineage-labeled endogenous Sox9+ cells in AKI mouse models, allowing long-term monitoring and visualization of the kidney regeneration process.