A range of antitumor agents for cancer treatment is available; however, they show low specificity, which often limit their use. Recently, we have reported the preparation of folate-coated long-circulating and pH-sensitive liposomes (SpHL-folate-PTX) loaded with paclitaxel (PTX), an effective drug for the treatment of solid tumors, including breast cancer. The purpose of this study was to prepare and characterize SpHL-PTX and SpHL-folate-PTX radiolabeled with technetium–99 m ( ^99m Tc). Biodistribution studies and scintigraphic images were performed after intravenous administration of ^99m Tc-PTX, ^99m Tc-SpHL-PTX and ^99m Tc-SpHL-folate-PTX into healthy and tumor-bearing mice. High radiochemical purity ( > 98%) and in vitro stability ( > 90%) were achieved for both liposome formulations. The pharmacokinetic properties of ^99m Tc-SpHL-DTPA-PTX and ^99m Tc-SpHL-folate-DTPA-PTX decreased in a monophasic manner showing half-life of 400.1 and 541.8 min, respectively. Scintigraphic images and biodistribution studies showed a significant uptake in liver, spleen and kidneys, demonstrating these routes as way for excretion. At 8 h post-injection, the liposomal tumor uptake was higher than ^99m Tc-PTX. Interesting, 4 h after administration, the liposome folate coated showed higher tumor-to-muscle ratio than ^99m Tc-SpHL-DTPA-PTX and ^99m Tc-PTX. In conclusion, the liposomal systems, showed high tumor uptake by scintigraphic images, especially the ^99m Tc-SpHL-folate-DTPA-PTX that showed a sustained and higher tumor-to-muscle ratio than non-functionalized liposome, which indicate its feasibility as a PTX delivery system to folate positive tumors.