In this work, we demonstrate near-infrared-to-visible upconverting fluorescent nanoparticles as a promising platform for lanthanide based or luminescence resonance energy transfer (LRET)-based biodetection of DNA release in live cells. Highly monodispersed, stable aqueous suspension of nanoparticles, surface-functionalized with amino groups for binding to DNA, were prepared and characterized. These amino-functionalized nanoparticles were able to electrostatically bind to DNA and protect it from DNaseI degradation as shown by gel electrophoresis. Attachment of DNA to the nanoparticles was also confirmed by LRET, which was observed to occur between the donor nanoparticle and acceptor POPO-3 dye intercalating the DNA. The intrinsic fluorescence property of upconverting fluorescent nanoparticles makes them useful for tracking their cellular localization without the use of any additional fluorescent tag. We were able to track the movement of these DNA-loaded nanoparticles into the cell cytoplasm where they successfully released their genetic cargo. Successful transfection of the loaded DNA material in vitro and in vivo was confirmed by expression of its encoded green fluorescent protein (GFP) in Hela cells and induction of specific antibody in mice, respectively.