Secondary upper limb lymphoedema is usually caused by lymphatic system dysfunction. Diagnosis is primarily based on clinical features. However, there are no distinct diagnostic criteria for lymphoedema. Although conventional lymphoscintigraphy is a useful technique to diagnose the severity of lymphoedema, the resultant data are two-dimensional. In this study, we examined the pathology of lymphoedema using single photon emission computed tomography-computed tomography lymphoscintigraphy (SPECT-CT LSG), a new technique that provides 3-dimensional information on lymph flow. We observed lymph flow pathways in the subcutaneous and muscle layers of the upper limbs. A significant positive correlation was found between the dermal back flow (DBF) type and the visualization of lymph nodes around the clavicle (p = 0.000266), the type of lymph flow pathways and the visualization of lymph nodes around the clavicle (p = 0.00963), and the DBF type and the lymph flow pathway (p = 0.00766). As the severity of lymphoedema increased, the DBF appeared more distally in the upper limb and the flow into the lymph nodes around the clavicle decreased, whereas the lymph flow pathways in the muscle layer became dominant. These findings demonstrate the features of lymphoedema pathology and the functional anatomy and physiology of the lymphatic system without the need for cadaver dissection.