In the recent progress of gene and cell therapy, novel drug delivery system (DDS) has been required for efficient delivery of small molecules/drugs and also the safety for clinical usage. We have already developed the unique transfection technique by preparing magnetic vector and using permanent magnet. This technique can improve the transfection efficiency. In this study, we directly associated plasmid DNA with magnetic nanoparticles, which can potentially enhance their transfection efficiency by magnetic force. Magnetic nanoparticle, such as magnetite, its average size of 18.7 nm, can be navigated by magnetic force and is basically consisted with oxidized Fe that is commonly used as the supplement drug for anemia. The magnetite particles coated with protamine sulfate, which gives a cationic surface charge onto the magnetite particle, significantly enhanced the transfection efficiency in vitro cell culture system. The magnetite particles coated with protamine sulfate also easily associated with cell surface, leading to high magnetic seeding percentage. From these results, it was found that the size and surface chemistry of magnetic particles would be tailored to meet specific demands on physical and biological characteristics accordingly. Overall, magnetic nanoparticles with different surface modification enhance the association with plasmid DNA and cell surface as well as HVJ-E, which potentially help to improve the drug delivery system.