In Mexico, urban rabies has been reduced during the last decade thanks to intensive canine control and vaccination campaigns; however, rabies transmitted by wild animals, especially by bats, has been increasing due to vampire bats feeding on livestock. Vampire bat populations has been controlled by culling with vampiricides, reducing indiscriminately other bat species. Hence, bat vaccination for rabies offers an alternative for culling. Nevertheless, available rabies vaccines are not suitable for their use in wildlife from emerging countries. This project presents an alternative for the use of plasmid vaccines using bio-nanotechnology, to create low-cost and accessible vaccines. To accomplish this goal, chitosan nanoparticles were synthesized by ionic gelation and conjugated by coacervation with a pDNA rabies vaccine to test their attachment efficiency. Also, the conjugate was functionalized with Protoporphyrin IX and Folic acid as biomarkers. The nanoparticles complex was characterized by ultraviolet visible spectroscopy, infrared spectroscopy, transmission electron microscopy, dynamic light scattering, and the Z potential was obtained. In vitro tests were performed on cell viability and transfection. The nanoparticles possessed a low polydispersity, a mean size of 118.5 ± 13.6 nm and a Z potential of 17.3 mV. The attachment efficiency was of 100% independent of pDNA added. In contrast to functionalized nanoparticles which showed a max attachment efficiency of 99.6% dependent of pDNA concentration and the method of functionalization. The conjugate did not influence the viability and they improved the transfection efficiency. Results suggest that these nanoparticles are easy to prepare, inexpensive, and exhibit potential for plasmid delivery as it improves transfection efficiency of pDNA vaccines.