The increasing use of nanoparticles (NPs) in technological applications and in commercial products has escalated environmental health and safety concerns. The detection of NPs in the environment and in biological systems is challenged by limitations associated with commonly used analytical techniques. In this paper we report on the development and characterization of NP binding antibodies, termed NProbes. Phage display methodology was used to discover antibodies that bind NPs dispersed in solution. We present a proof-of-concept for the generation of NProbes and their use for detecting quantum dots and titanium dioxide NPs in vitro and in an ex vivo human skin model. Continued development and refinement of NProbes to detect NPs that vary in composition, shape, size, and surface coating will comprise a powerful tool kit that can be used to advance nanotechnology research particularly in the nanotoxicology and nanotherapeutics fields.