Protocol for brain and ventricle volume comparison after clearing and light-sheet microscopy in mice

Analysis of brain and ventricular volumes provides insights into the physiological and pathological progression of brain development and aging, but accessible and cost-effective approaches are lacking. Here, we present a workflow to quantify relative brain and ventricular volumes in unsectioned cleared mouse brains following fluorescent detection of brain tissue and ventricle lining. We describe steps for brain harvesting, clearing, imaging by light-sheet microscopy, image resampling, and ventricle annotation, and comparing brains from adult, neonate, germ-free, and hydrocephalic mouse models.