Patient and organ motion is a potentially limiting factor in gamma camera single-photon emission tomography (SPET) imaging, as highlighted in stress thallium myocardial SPET, where the heart may exhibit a systematic axial motion (cardiac creep) following stress. Multi-rotation SPET has previously been described as a means of obtaining better raw data for motion detection and correction. This study describes the validation of a computerised motion detection algorithm applied to multi-rotation SPET, and reports measured motions in thallium myocardial stress SPET studies from a single-headed gamma camera. Forty-two patients underwent pharmacological stress (dipyridamole) with leg raising, with injection of 75 MBq thallium-201 and imaging after a 10-min delay to detect or evaluate coronary artery disease. Multi-rotation gamma camera SPET was performed with a single-headed gamma camera, with five sequential rapid (4.5 min) continuous SPET mode rotations over 180 degrees. A one-dimensional cross-correlation alignment technique was applied to the projection images to perform motion detection and correction in the axial direction prior to combining the five data sets for tomographic reconstruction. Validation of the cross-correlation alignment analysis was carried out by performing imaging with measured whole-body axial motions in nine subjects, and by reproducibility measurements on multi-rotation data sets. The effect of the applied motion correction was evaluated by calculating mean differences between image pairs before and after shifting, and the general reliability of the automatic motion detection was checked to within one pixel by visual assessment of 160 image pairs. Validation measurements of the cross-correlation technique gave a mean absolute error of 1.5+/-0.4 mm (0.24+/-0.06pixels) with a maximum error of 3.7 mm (0.6 pixels). In 40 subjects undergoing pharmacological stress 201Tl myocardial SPET imaging, the mean cardiac axial creep movement was calculated as 3.1+/-0.7 mm (0. 49+/-0.11 pixels), with 13 out of 40 (32%) having a calculated motion of 1 pixel (6.3 mm) or more. The automatic image shift was visually judged to be within 1 pixel in all 160 image pair analyses, and the mean pixel value difference between image pairs was reduced following image shifting. It is concluded that multi-rotation 180 degrees SPET imaging provides raw data which allow objective and accurate motion detection of cardiac motion in thallium stress myocardial imaging, whilst the one-dimensional cross-correlation technique demonstrates adequate accuracy and reliability to be applied as an automatic motion screening technique on these data.