Dual isotope Single Photon Emission Computed Tomography permits in vivo measurements of two separate signals simultaneously, but the quantitative accuracy of these measurements is degraded due to the presence of cross-talk. An iterative triple energy window method was developed to correct for cross-talk in In-111 and Tc-99m dual isotope SPECT. The correction was evaluated in comparison to other TEW-based methods in phantoms and in vivo. In phantoms containing different (Tc-99m)/(In-111) activity fractions, scattered In-111 photons falsely detected as Tc-99m resulted in an over-estimation of
Tc-99m activity by up to 80%. Application of the iterative TEW resulted in quantification of total Tc-99m activity within 5% of the truth across the largest range of activity fractions. In vivo, modified and iterative TEW both quantified total heart Tc-99m activity within 5% in a rat, but the iterative TEW was superior to the modified TEW in removing cross-talk in regions of mostly In-111.