Lymphoscintigraphy readily provides confirmation of chylothorax but not detailed localization of the leakage site. A 77-year-old woman developed traumatic chylothorax and underwent lymphoscintigraphy with radiolabeled albumin to identify the site of lymph leakage. Dynamic imaging demonstrated appearance of focal activity presumably in the upper mediastinum, followed by spread to the left hemithorax. Subsequently, SPECT/CT showed that the leakage site was located in the left side of the upper mediastinum, and this location was confirmed during video-assisted thoracoscopic surgery. Dynamic imaging demonstrated the site of first appearance of abnormal activity, and SPECT/CT enabled detailed localization of the abnormal activity with anatomic correlation. The combination of dynamic imaging with SPECT/CT appears to be recommendable for lymphoscintigraphic assessment of chylothorax.