Adding subtraction single-photon emission computed tomography/computed tomography (SPECT/CT) to dual isotope (I-123 and Tc-99m-sestamibi) subtraction parathyroid scintigraphy is not widely implemented. We aimed to assess the added value of dual isotope subtraction SPECT/CT over single isotope SPECT/CT as an adjunct to dual isotope planar pinhole subtraction scintigraphy. Methods: Parathyroid scintigraphies from 106 patients with an estimated total of 415 parathyroid glands who (1) were

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... with primary hyperparathyroidism, (2) underwent dual isotope subtraction scintigraphy in the Department of Nuclear Medicine, Gentofte Hospital, Denmark throughout 2017 and (3) underwent subsequent parathyroidectomy, were included. The original dual isotope planar pinhole subtraction plus dual isotope subtraction SPECT/CT (dual/dual method) exams were retrospectively re-evaluated using only Tc-99m-sestamibi SPECT/CT (dual/single method). Statistics were calculated per parathyroid. Surgical results confirmed by pathology served as reference standard. Results: The dual/dual method had higher sensitivity than the dual/single method (82% (95%CI 74%–88%) vs. 69% (95%CI 60%–77%)) while specificity, positive and negative predictive values (PPV and NPV) were similar (specificity 96% vs. 93%, PPV's 87% vs. 82% and NPV's 89% vs. 93%). Reader confidence was higher when employing the dual/dual method (p = 0.001). Conclusions: The dual/dual method can be considered superior to the dual/single method in the preoperative imaging in primary hyperparathyroidism.