The clinical diagnosis of primary hyperparathyroidism is based largely on serum laboratory test results, as patients often are asymptomatic. Surgery, often with bilateral exploration of the neck, has been considered the definitive treatment for symptomatic disease. However, given that approximately 90% of cases are due to a single parathyroid adenoma, a better treatment may be the selective surgical excision of the hyperfunctioning parathyroid gland after its preoperative identification and localization at radiologic imaging. Scintigraphy and ultrasonography are the imaging modalities most often used for preoperative localization. Various scintigraphic protocols may be used in the clinical setting: Single-phase dual-isotope subtraction imaging, dual-phase single-isotope imaging, or a combination of the two may be used to obtain planar or tomographic views. Single photon emission computed tomography (SPECT) with the use of technetium-99m ((99m)Tc) sestamibi as the radiotracer, especially when combined with x-ray-based computed tomography (CT), is particularly helpful for preoperative localization: The three-dimensional functional information from SPECT is fused with the anatomic information obtained from CT. In addition, knowledge of the anatomy and embryologic development of the parathyroid glands and the pathophysiology of primary hyperparathyroidism aid in the identification and localization of hyperfunctioning glands.