Molecular excitation of photosensitizing agents provides reactive excited states, which can initiate chemical reactions, but it can also lead to molecular relaxation via radiative photophysical processes, providing the basis for fluorescence diagnostics. The best-known example of the former is Photodynamic Therapy (PDT), which is now approved for the treatment of a number of neoplastic and non-neoplastic pathologies. Although the concept of the use of photodynamic agents in diagnostics is as old as their use in therapy, the focused development of this aspect has been relatively recent. Typically, photodynamic agents have high triplet yields and relatively long triplet lifetimes (microsecond range), which allows them to interact and destroy molecular targets near them either directly or indirectly by producing other toxic molecular species. Associated with a high triplet yield is the fortunate attribute of most PDT agents in having low but finite fluorescence quantum yields. Fluorescence from these molecules may be used not only for diagnostics of disease de novo but also for guided surgery, PDT dosimetry and therapeutic monitoring. Other uses of fluorescence in PDT (not necessarily from the PDT agents) include the development of technologies that allow tracking of cells during treatment in vivo, studies of sub-cellular localization of molecules for mechanistic studies and photosensitizer tracking for specific targeting. An overview of studies on these aspects from different laboratories will be presented.