In this study, NFkB decoy oligodeoxynucleotides (NDON) having water-soluble and anionic properties were used as the model gene-drug.Several types of NDON loaded PLGA nanospheres were prepared by a new technique based on Emulsion Solvent Diffusion (ESD) method in the aqueous PVA solution.It was found that the PLGA nanospheres could carry NDON both inside themselves and on their outer layers.Furthermore, applying the delayed type hypersensitivity (DTH) model with mice, the inhibition efficacy of the NDON loaded PLGA nanospheres on the delayed type allergic reaction was evaluated.The NDON loaded PLGA nanospheres showed the same bioavailability as that of conventional NDON ointment, but with less than one-tenth of NDON concentration.This result demonstrated that the PLGA nanospheres could improve the skin permeability, drug delivery efficiency, and gene expression efficiency of NDON.