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Computer simulations, a phantom study and a human study were performed to determine whether a slowly rotating single-photon computed emission tomography (SPECT) system could provide accurate arterial input functions for quantification of myocardial perfusion imaging using kinetic models. The errors induced by data inconsistency associated with imaging with slow camera rotation during tracer injection were evaluated with an approach called SPECT/P (dynamic SPECT from positron emission tomography (PET)) and SPECT/D (dynamic SPECT from database of SPECT phantom projections). SPECT/P simulated SPECT-like dynamic projections using reprojections of reconstructed dynamic (94)Tc-methoxyisobutylisonitrile ((94)Tc-MIBI) PET images acquired in three human subjects (1 min infusion). This approach was used to evaluate the accuracy of estimating myocardial wash-in rate parameters K(1) for rotation speeds providing 180° of projection data every 27 or 54 s. Blood input and myocardium tissue time-activity curves (TACs) were estimated using spatiotemporal splines. These were fit to a one-compartment perfusion model to obtain wash-in rate parameters K(1). For the second method (SPECT/D), an anthropomorphic cardiac torso phantom was used to create real SPECT dynamic projection data of a tracer distribution derived from (94)Tc-MIBI PET scans in the blood pool, myocardium, liver and background. This method introduced attenuation, collimation and scatter into the modeling of dynamic SPECT projections. Both approaches were used to evaluate the accuracy of estimating myocardial wash-in parameters for rotation speeds providing 180° of projection data every 27 and 54 s. Dynamic cardiac SPECT was also performed in a human subject at rest using a hybrid SPECT/CT scanner. Dynamic measurements of (99m)Tc-tetrofosmin in the myocardium were obtained using an infusion time of 2 min. Blood input, myocardium tissue and liver TACs were estimated using the same spatiotemporal splines. The spatiotemporal maximum-likelihood expectation-maximization (4D ML-EM) reconstructions gave more accurate reconstructions than did standard frame-by-frame static 3D ML-EM reconstructions. The SPECT/P results showed that 4D ML-EM reconstruction gave higher and more accurate estimates of K(1) than did 3D ML-EM, yielding anywhere from a 44% underestimation to 24% overestimation for the three patients. The SPECT/D results showed that 4D ML-EM reconstruction gave an overestimation of 28% and 3D ML-EM gave an underestimation of 1% for K(1). For the patient study the 4D ML-EM reconstruction provided continuous images as a function of time of the concentration in both ventricular cavities and myocardium during the 2 min infusion. It is demonstrated that a 2 min infusion with a two-headed SPECT system rotating 180° every 54 s can produce measurements of blood pool and myocardial TACs, though the SPECT simulation studies showed that one must sample at least every 30 s to capture a 1 min infusion input function.