BACKGROUND The objective of this study was to determine if dual-tracer protocol using thallium-201 (201TI) chloride for rest and technetium-99m (99mTc)-sestamibi for stress images (TI-MIBI) detects ischemia better than single-tracer protocol using 99mTc-sestamibi for both rest and stress (MIBI) myocardial perfusion images. RESULTS A total of 460 patients with suspected or known coronary artery disease (CAD) were included in the study, with 230 (50%) undergoing single-tracer MIBI protocol and 230 (50%) undergoing dual-tracer TI-MIBI protocol. Patient demographics were matched, and the proportion of patients having treadmill stress or dipyridamole stress was similar between the two groups. Stress and rest single photon emission computed tomographic images were recorded in all patients and dual-tracer 201TI and 99mTc-sestamibi (TI-MIBI) perfusion images were acquired separately. The scintigraphic images were analyzed visually, and a semiquantitative method of scoring reversible ischemia was used. Patients' reversibility scores in each group were compiled (ischemia index) and compared to determine the relative ability of each protocol in identifying ischemia. CONCLUSIONS The frequency of ischemia and ischemia index was found to be significantly (P < 0.05) higher in the TI-MIBI group compared with the MIBI group. For detecting myocardial ischemia or viability, the dual-tracer TI-MIBI acquisition technique appears superior to the single-tracer MIBI protocol.