The self-assembly of a bichromophoric naphthalenediimide (NDI) into nanofibers enables efficient energy transfer to perylenediimide (PDI) molecules through host-guest interactions, as visualized using fluorescence microscopy and AFM. Fluorescence imaging and microspectroscopy revealed packing defects, inhomogeneities, and exciton mobility along the NDI nanofibers, showing strong blue polarized emission from NDI, while PDI guest molecules exhibit minimal polarization.