Objective To evaluate the capability of ultrasound for preoperative localization in primary hyperparathyroidism. Study Design Prospective study. Setting Multi-institutional Midwest Head and Neck Cancer Consortium. Subjects and Methods Two hundred twenty patients who underwent preoperative localization and had parathyroid surgery were evaluated. The findings of preoperative localization studies were correlated with surgical findings. Results Preoperative ultrasonography, sestamibi scintigraphy, or both were obtained in 77%, 93%, and 69% of the patients, respectively. Preoperative ultrasonography and sestamibi scintigraphy localized an abnormality in 71% and 79% of patients, respectively. At the time of surgery, the localization by ultrasound was accurate in 82%. The accuracy of localization was similar for sestamibi scintigraphy (85%). In patients with inaccurate ultrasound localization, the sestamibi scintigraphy correctly identified the site of disease in only 45%. In patients with a nonlocalizing ultrasound, sestamibi scintigraphy was able to localize disease in only 47%, with 2 being in the mediastinum. Conclusions Ultrasonography is an acceptable initial localization study for patients with primary hyperparathyroidism. In patients with nonlocalizing ultrasound, sestamibi scintigraphy should be obtained, but can be expected to detect an abnormality in less than 50% of patients.