Abstract: We report the real-time monitoring of protein-protein interactions without labeling either of the two interacting proteins, posing minimum effects on the binding properties of the proteins. In particular, the methods provide protein/aptamer complexes to probe the interactions in a competitive assay where the binding of an aptamer to its target protein is altered by a second protein that interacts with the target protein. Two signal transduction strategies, fluorescence resonance energy transfer (FRET) and fluorescence anisotropy, are described.