| Summary: | Challenges posed by the retention of radiopharmaceuticals in unintended organs affect the quality of patient procedures when undergoing diagnostics and therapeutics. The aim of this study was to formulate a suitable tracer encapsulated in liposomes using different techniques and compounds to enhance the stability, uptake, clearance, and cytotoxic effect of the radiopharmaceutical. Cationic liposomes were prepared by a thin-film method using dipalmitoyl phosphatidylcholine (DPPC) and cholesterol. Whole-body gamma camera images were acquired of intravenously injected New Zealand rabbits. Additionally, liposomes were assessed using stability, toxicity, zeta potential, and particle size tests. In the control cases, Technetium-99m (<sup>99m</sup>Tc)-sestamibi exhibited the lowest heart uptake the blood pool and delayed images compared to both <sup>99m</sup>Tc-liposomal agents. Liver and spleen uptake in the control samples with <sup>99m</sup>Tc-sestamibi increased in 1-h-delayed images, unlike with <sup>99m</sup>Tc-liposomal agents, which were decreased in delayed images. The mean maximum count in the bladder for <sup>99m</sup>Tc-sestamibi loaded liposomes 1 h post-injection was 2354.6 (±2.6%) compared to 178.4 (±0.54%) for <sup>99m</sup>Tc-sestamibi without liposomes. Liposomal encapsulation reduced the cytotoxic effect of the sestamibi. <sup>99m</sup>Tc-MIBI-cationic liposomes exhibited excellent early uptake and clearance compared to <sup>99m</sup>Tc-MIBI without liposomes. Adding cholesterol during liposome formation enhanced the stability and specificity of the targeted organs.
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