It is challenging to assess protein-membrane interactions because of the lack of appropriate tools to detect position changes of single proteins in the ∼4 nm range of biological membranes. We developed an assay recently, termed surface-induced fluorescence attenuation (SIFA). It is able to track both vertical and lateral dynamic motion of singly labeled membrane proteins in supported lipid bilayers. Similar to the FRET (fluorescence resonance energy transfer) principle, SIFA takes advantage of the energy transfer from a fluorophore to a light-absorbing surface to determine the distance at 2-8 nm away from the surface. By labeling a protein with a proper fluorophore and using graphene oxide as a two-dimensional quencher, we showed that SIFA is capable of monitoring three-dimensional movements of the fluorophore-labeled protein not only inside but also above the lipid bilayer atop the graphene oxide. Our data show that SIFA is a well-suited method to study the interplay between proteins and membranes.