**1. Introduction**

Fluorescence spectroscopy has been revolutionizing the field of life sciences and clinical routines such as diagnostics and biosensing, due to its impressive sensitivity and the biocompatibility of many fluorescent organic compounds, which allows one to probe biological processes in vivo in noninvasive bioimaging procedures. The improvement of instrumentation has granted optical-based sensing routines a new level of sensitivity, accuracy, and reliability. Very subtle changes in fluorescence intensity, or even extremely low levels of light that might result from an interaction between the fluorescent probe or sensor with the environment under study, are easily detectable nowadays thanks to modern instrumentation such as photomultiplier tubes, in which electronic impulses are created by just a single photon. In the time-resolved fluorescence approach, in which the fluorescence lifetime of the fluorophore is monitored, with current femtosecond pulsed lasers, even more sensitive and reliable measurements are possible, since the intensity decay is not affected by a number of possible undesired factors that interferes in the steady-state intensity. These advantages make the fluorescence spectroscopy a powerful technique, paving the way to the most wished rapid and low-cost sensing in a wide range of biological and environmental applications and point-of-care diagnostics for real-time

monitoring of physiological conditions. Sensing methods of remarkable sensitivity, reliability, and selectivity based on fluorescence spectroscopy dominates the field of sensing and biosensing. DNA sequencing and fragment analysis, fluorescence staining for bioimaging and fluorescence immunoassays are all based on fluorescence techniques.

The countless possibilities of combinations of biorecognition element, support matrix, and the transducing method in biosensors constituted of nanomaterials make it possible to design versatile and selective biosensors. In this review, particular attention is centered on luminescent biosensors based on the Förster resonance energy transfer, or FRET, biosensing transducing method, which encompasses a huge variety of biosensors, due to their unique sensitivity, selectivity, and fast response. For this reason, FRET-based sensors have enabled, for example, intracellular monitoring of ROS kinetics and oxygen sensing, which is vital for elucidating how tumor cells respond to treatment, in order to develop better therapeutic strategies. Before FRET sensors were introduced, these practices were hampered by difficulties and unreliability in real-time monitoring intracellular ROS. In fluorescence bioimaging, thanks to its high spatial and temporal resolution, it is becoming possible to probe in real time biological elements and processes, such as enzymatic reactions, protein/protein, protein/nucleic acid, protein/substrate, and biomembrane interactions.

In the context of environmental monitoring, with the possibility of miniaturization of biosensors based on nanomaterials, it is becoming possible to perform fast and accurate field analysis and real-time surveillance of analytes relevant in the assessment of water quality. A variety of FRET-based biosensors for water pollutants, such as heavy metal ions, pesticides, antibiotics, and halogenated compounds, is reported, some of them capable of detecting concentrations in the pico and nanomolar scales. Such sensitivity levels are far from reach with conventional analytical methods.
