Background There currently is no consensus on the optimal localization procedure and imaging protocol for parathyroid adenoma. Parathyroid four-dimensional (4D) CT has emerged as a promising method for preoperative localization. Purpose To evaluate the diagnostic performance of parathyroid 4D CT and technetium 99m-sestamibi (hereafter, referred to as sestamibi) SPECT/CT in preoperative localization in patients with primary hyperparathyroidism. Materials and Methods This was a single-institution retrospective study of patients with primary hyperparathyroidism who underwent a combined imaging protocol of sestamibi SPECT/CT and 4D CT (noncontrast, contrast agent-enhanced, arterial, and delayed venous phases) acquired in a single setting from February 2013 to May 2016, with subsequent parathyroidectomy within 6 months. Reference standard for correct localization was on the basis of location denoted on operative reports, with pathologic confirmation of parathyroid adenoma or hyperplasia. By using a four-quadrant analysis, sensitivity, specificity, and area under the curve (AUC) for localization of the hyperfunctioning parathyroid gland or glands at sestamibi SPECT/CT and 4D CT were compared, per modality and in combination. Results Four hundred patients (319 women, 81 men; mean age, 61 years ± 14 [standard deviation]) were evaluated. Similar diagnostic performance was found in both combined 4D CT with sestamibi SPECT/CT and 4D CT alone (area under the curve [AUC], 0.88 [95% CI: 0.86, 0.90] and 0.87 [95% CI: 0.85, 0.90], respectively; P = .82). Both modalities outperformed sestamibi SPECT/CT (AUC, 0.78; 95% CI: 0.76, 0.81; P < .001). Four-dimensional CT showed higher sensitivity than did sestamibi SPECT/CT (sensitivity, 79.3% [414 of 522] vs 58.0% [303 of 522], respectively; P < .001). In a subset analysis, 4D CT had higher sensitivity than sestamibi SPECT/CT in patients with single-gland disease (sensitivity, 92.5% [297 of 321] vs 75.1% [241 of 321], respectively; P < .001) and with multigland disease (sensitivity, 58.2% [117 of 201] vs 30.8% [62 of 201], respectively; P < .001). Conclusion Four-dimensional CT provided superior preoperative localization compared with sestamibi SPECT/CT in patients with single and multigland disease. The combination of the two modalities did not improve diagnostic performance compared with four-dimensional CT alone. © RSNA, 2019 Online supplemental material is available for this article. See also the editorial by Sinha and Oates in this issue.