Two same-day protocols (rest/exercise [Protocol 1] and exercise/rest [Protocol 2]) with sestamibi (hexakis 2-methoxy-2-isobutyl-isonitrile) were performed within 2 to 14 days of each other after randomization. The initial study in each protocol was done using a dose of 185-296 MBq of 99mTc- sestamibi. The second study in each protocol used a dose of 555-925 MBq. SPECT imaging was started 30 to 60 min after injection using a 180° anterior arc. Segmental analysis was interpreted as normal, scar or ischemia (20 segments/patient). Among the protocols, there was concordance in 93% of the segments (593/640 segments). In the 11 patients with coronary artery disease and no prior myocardial infarction who had ischemic abnormality, count densities from abnormal and normal zones were compared between the two protocols. Protocol 1 showed greater count differences between abnormal and normal zones on exercise images with better normalization of abnormality on rest images than Protocol 2 (p < 0.05). Technetium-99m-sestamibi provides high quality images using either of the two same-day protocols. However, the rest/exercise protocol provides better image contrast and ability to detect reversibility of perfusion defects, and is the preferred same-day protocol.