The case reported here describes a transthoracic approach for removal of a splenic lymphoma from a 5-year-old mule. Nine months prior to presentation at Iowa State University College of Veterinary Medicine (ISUCVM), abdominal ultrasound was performed as part of a work-up to investigate the cause of lethargy, anorexia and anaemia. Ultrasound examination identified a haemoabdomen and multiple splenic masses. Transabdominal biopsies were collected for histopathological evaluation and lymphoma diagnosed. Based on a number of clinical and laboratory examinations there was a lack of support for multicentric involvement and all results suggested solitary splenic involvement. Surgical removal of the spleen was accomplished via a transthoracic approach without complications. Histopathology of the splenic masses revealed a B cell lymphoma. Bone marrow was collected during surgery and submitted for cytology and was found to contain a normal lymphoid population with no evidence of neoplastic cells. The mule was discharged from the hospital 11 days after surgery.