The particle sizes and encapsulation ratio of pDNA/γ-CyD/AF-liposomes should be involved in the enhancing effect of γ-CyD on gene transfer activity of AFliposomes. The particle size of pDNA/γ-CyD/AF-liposomes was decreased in the presence of γ-CyD, although the ζpotential value of pDNA/γ-CyD/AF-liposomes was almost equivalent to that of pDNA/AF-liposomes and pDNA/HP-γ-CyD/AF-liposomes ( Table 2 ), suggesting that γ-CyD inhibits the aggregation of pDNA/AF-liposomes, because the particle size shows more than 200 nm, despite the fact that the liposomes were extruded through a filter membrane having a pore size of 200 nm. Meanwhile, the encapsulation ratio of pDNA was significantly increased by adding γ-CyD to pDNA/AF-liposomes (Table 3) . Thus, these lines of evidence speculate that addition of γ-CyD enhances cellular uptake of pDNA/AF-liposomes. In fact, the CLSM study demonstrated that cellular uptake of pDNA/γ-CyD/AF-liposomes was higher than that of pDNA/γ-CyD/AF-liposomes ( Figure 11 ). Furthermore, we confirmed that transfection efficiency of pDNA/γ-CyD/AF-liposomes was increased, as the encapsulation ratio of pDNA into pDNA/γ-CyD/AF-liposomes was augmented (Table 4 ). In view of the findings, the particle size of pDNA/γ-CyD/AF-liposomes and encapsulation ratio of pDNA into pDNA/γ-CyD/AF-liposomes are crucial role for enhancing transfection efficiency of pDNA/AF-liposomes.