The aims of this study were: (a) to define the accuracy of a preoperative parathyroid imaging protocol based on the combination of technetium-99m pertechnetate/technetium-99m methoxyisobutylisonitrile (99mTcO4/ 99mTc-MIBI) scan and neck ultrasound (US) in selecting patients with primary hyperparathyroidism (pHPT) eligible for a limited neck exploration, and (b) to investigate the potential role of the intraoperative gamma probe (IGP) in radio-guided minimally invasive surgery. 99mTcO4/99mTc-MIBI subtraction scan was performed by means of potassium perchlorate administration with the aim of effecting rapid 99mTcO4 wash-out from the thyroid. Minimally invasive surgery using an IGP was commenced some minutes following the injection of a low, 70 MBq, 99mTc-MIBI dose. Intraoperative PTH (i-PTH) was measured. On the basis of preoperative imaging, 21 pHPT consecutive patients were selected for a limited neck dissection. In 18 of them, a single parathyroid adenoma was found at surgery and IGP allowed performance of parathyroidectomy through a small, 2-2.5 cm, skin incision with a relatively short surgical duration (mean 38 min). i-PTH rapidly normalised in all cases. In two patients, a parathyroid carcinoma was diagnosed at surgery; consequently, a wide neck exploration associated with a near-total thyroidectomy was performed. No loco-regional metastatic lesions were found and i-PTH rapidly normalised after carcinoma excision. In one patient, i-PTH remained elevated after removal of the enlarged parathyroid gland which was localised by 99mTcO4/99mTc-MIBI scan and US. A bilateral exploration was needed to remove a contralateral enlarged parathyroid gland. Combined, 99mTcO4/99mTc-MIBI scan and US imaging correctly localised a single parathyroid gland in 20/21 patients (95.2%); thus, this protocol appears to be accurate enough for the preoperative selection of pHPT patients eligible for limited neck surgery. Moreover, in these selected patients the IGP seems to be helpful in performing radio-guided minimally invasive surgery.