Background: This study examines the usefulness of thallium-201 single photon emission computed tomography (201Tl SPECT) in differentiating pulmonary malignancies from benign lesions by using dual phase 201Tl scintigraphy. Methods: One hundred and six patients with thoracic lesions and confirmed diagnoses were assessed by 201Tl chest SPECT examinations; of these 106 enrolled thoracic lesions, 59 were malignancies and 47 were benign lesions. Dual phase 201Tl SPECT was performed with an early image acquired at 10–20 min and a delayed image at 2–3 h after intravenous injection of 2–3 mCi of 201Tl. The results of 201Tl SPECT images were classified as either malignant or benign lesions by visual interpretation: the lesion was interpreted as positive for malignancy if the uptake of 201Tl in the lung lesion in the delayed phase was increased or persistent as compared with that in the early phase image; otherwise, it was considered as a benign lesion. Simultaneously, the traditional method of retention index (RI) was also calculated to help in the differential diagnosis of pulmonary lesions. Then, both methods of dual phase 201Tl SPECT, visual reading and traditional RI, were compared to differentiate pulmonary malignancies from benign lesions. Results: Analyzing the image results, we found that dual phase 201Tl SPECT could differentiate pulmonary malignancies from benign lesions with a sensitivity of 83%, a specificity of 91% and an accuracy of 87%. Moreover, by using the traditional RI method of 201Tl SPECT, it could differentiate pulmonary malignancy from benign lesions with a sensitivity of 79.3%, a specificity of 80.8% and an accuracy of 80%. Conclusions: We conclude that using dual phase 201Tl SPECT with visual interpretation is a simpler and potentially more effective method for differentiating pulmonary malignancies from benign lesions, with results compatible with the traditional RI method of 201Tl SPECT.