We aimed to evaluate the diagnostic and preoperative localization capacity of 99mTc methoxyisobutylnitrile (MIBI) parathyroid scintigraphy and ultrasonography (USG) in enlarged parathyroid glands in the primary hyperparathyroidism (pHPT) as well as the relationship between the success rate of these techniques and biochemical values. In this study, we retrospectively evaluated 39 patients with clinical and biological evidence of pHPT who referred to the university hospital for MIBI parathyroid scintigraphy. Patients were examined with USG and double-phase MIBI parathyroid scintigraphy for the detection of enlarged parathyroid glands. Preoperative serum intact parathyroid hormone (iPTH) levels, calcium (Ca), phosphate and alkaline phosphatase measurements were obtained. A total of 45 parathyroid lesions in 39 patients were reviewed. Thirty-four patients had a single adenoma and 5 patients with multi-gland disease had 11 abnormal parathyroid glands including three adenomas, whereas the remaining 8 glands showed hyperplasia. The overall sensitivities of MIBI parathyroid scintigraphy, USG and combined techniques were 85.3%, 72.5% and 90.4%, respectively; the positive predictive values (PPV) were 89.7%, 85.2% and 92.6%, respectively. The most successful approach for detection of enlarged parathyroid glands in hyperparathyroidism is the concurrent application of USG and MIBI parathyroid scintigraphy modalities. The concomitancy of thyroid diseases decreases the sensitivity of both MIBI parathyroid scintigraphy and USG in enlarged parathyroid glands.