Fluorescence resonance energy transfer (FRET) is widely used method in bio-imaging, especially for measuring distance in real time. FRET between two conventional fluorophores such as organic dyes and fluorescence proteins is generally used. However, observation duration is not sufficiently long (<10 s) to be utilized for bio-application. Unlike fluorophores, FRET using inorganic nanoparticles such as quantum dots (QDs) shows better efficiency in aspect of the photostability. Herein, we used QD and Au film to observe intensity changes in real time according to the distance between QD and Au film. To construct this system, surface chemistry for phase transfer (organic to aqueous) of QD was required for using QDs in biological applications as FRET probes. Then, to precisely control the distance between QD and Au, various ligands and biomolecules were used. As a result, QDs showed different intensities with various distances between QD and Au film due to FRET. From the correlation between the intensity and the distance, QD-Au complex with various distance could be used as the nano-ruler for real time bio-imaging.