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Nanosized liposomes composed of 1,2-distearoyl-sn-glycerol-3-phosphatidylcholine (DSPC), cholesterol and polyethylene glycol-conjugated phospholipid (PEG), incorporating FITC-dextran (FITC) and in some cases also Rhodamine-conjugated phospholipid (RHO) (as labels) were constructed by the thin film hydration method, followed by extrusion; membranes with pore diameters from 50 to 400 nm were used, while charged vesicles were produced by partially replacing DSPC with 1,2-distearoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DSPG). The uptake of liposomes by hCMED/D3 cells was evaluated by measuring FITC in cells, and their permeability across cell monolayers was evaluated, by measuring the FI of liposome associated-FITC and RHO in the receiving side of a monolayer-transwell system. Results prove that liposome size has a significant effect on their uptake and permeability (for both charged and non-charged vesicles). The effect of liposome charge on cell uptake was slight (but significant), however charge (in the range from −2 to −16 mV) did not significantly affect vesicle permeability; a significant decrease was only demonstrated for the liposome with the highest charge.