Comparative studies of thallium-201 (201Tl) and 99Tcm-tetrofosmin for the detection of ischaemic heart disease (IHD) have previously been reported. These 201Tl and 99Tcm-tetrofosmin studies were usually performed separately with different exercise loads at an interval of several days. Here, we used a dual-isotope technique with exercise myocardial SPET (single photon emission tomography) in 17 patients with IHD and 10 patients with normal coronary arteries. The triple-energy window (TEW) method was applied for cross-talk correction. SPET imaging was performed at 10 and 70 min (S1 and S2) after the injection of 99Tcm-tetrofosmin (222 MBq) and 201Tl (74 MBq) at peak exercise to determine the optimal imaging time of 99Tcm-tetrofosmin. The S2 value was obtained 35 min after the subject drank a glass of milk to accelerate hepatobiliary clearance. Twenty-five minutes after S2, 37 MBq of 201Tl were reinjected at rest and SPET imaging (S3) was performed. Immediately after S3, 666 MBq of 99Tcm-tetrofosmin were reinjected and SPET imaging (S4) was performed 50 min later. Representative short and vertical long axis tomograms were divided into 17 segments. Each segment was assessed using a 4-point scoring system. The defect score was defined as the sum of each segmental score. The defect scores for imaging at exercise were 14.3 +/- 11.4 for 201Tl at S1, 11.4 +/- 8 for 99Tcm-tetrofosmin at S1 and 9.7 +/-9.8 for 99Tcm-tetrofosmin at S2 (P < 0.01), respectively. The washout rate of 99Tcm-tetrofosmin for the first hour was 15.5 +/- 7.3% and 11.8 +/- 7.7% (P < 0.01), respectively, for the normal and ischaemic segments. The image quality of 201Tl at S1 was almost equivalent to that of 99Tcm-tetrofosmin at S1/S2. The overall sensitivity and specificity for the detection of ischaemia was 94% and 82% for 201Tl at S1, 89% and 86% for 99Tcm-tetrofosmin at S1, and 88% and 95% for 99Tcm-tetrofosmin at S2, respectively. The overall accuracy was 86%, 88% and 92%, respectively. The myocardial viability score was 7.4 +/- 7.1 for 201Tl at S3 and 5.8 +/- 7.0 for 99Tcm-tetrofosmin at S4 (P < 0.01). The results indicate that the diagnostic accuracy of 99Tcm-tetrofosmin for myocardial ischaemia is almost equivalent to that of 201Tl as assessed by dual SPET imaging, and that the optimal imaging time for 99Tcm-tetrofosmin is within 10-35 min (S1) after exercise.