Abstract: Aim To investigate the possibility of polymethacrylate nanoparticles (NP) for antisense oligodeoxynucleotides delivery system. Methods The nanoparticles were prepared by evaporating ethenol solution containing Eudragit RL100 or RS100, and then mixtured with oligonucleotides. The morphology and size were investigated by a transmission electron microscope and Mastersizer particle characterization systems, and the cytotoxicity was evaluated by Trypan Blue staining and hemolysis test. The flow cytometer was used to determine the uptake of fluorescence-labelled oligodeoxynucleotides. Results The morphology of nanoparticles showed spherical and orderly, the average diameter was about 127 nm,and almost the antisense oligodeoxynucleotides (ODN) were loaded when NP∶ODN was 6.6. The uptake of ODN was significantly increased when loaded by nanoparticles, which well depended on the nanoparticles concentration. Meanwhile, slightly cytotoxicity was observed when high dose of nanoparticles was used. Conclusion The polymethacrylate nanoparticles appeared to be a promising vehicle for gene delivery.