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Background Mediastinal ectopic parathyroid adenoma is a frequent cause of persistent or recurrent hyperparathyroidism, traditionally treated by open surgery. Thoracoscopic access is associated with reduced morbidity in mediastinal surgery. The aim of this study was to evaluate the feasibility and effectiveness of robot-assisted dissection for mediastinal ectopic parathyroid glands. Methods Two patients with recurrent secondary hyperparathyroidism and three with complicated primary hyperparathyroidism were operated on between July 2004 and August 2008 for ectopic mediastinal parathyroid glands. Fusion of single-photon emission computed tomography and computed tomography led to an exact identification of the culprit glands. Surgery was performed thoracoscopically with the da Vinci® robotic system using a three-trocar approach. Results All procedures were completed successfully with the robotic system. No perioperative morbidity or mortality was noted. Median operating time was 58 (range 42–125) min. Intraoperative parathyroid hormone reduction indicated complete resection. Median hospital stay was 3 (range 2–4) days. Conclusion Robot-assisted dissection is a promising approach for resection of ectopic parathyroid glands in remote narrow anatomical locations such as the mediastinum.