Fluorescent sensors classes. Class I: Ligand-binding sensors. This first class accounts for probes in which the binding of the ligand results in changes of the physicochemical properties of the sensor such as: (A) FPs that intrinsically respond to a parameter of interest, e.g., pH probes; (B) FPs, in which the binding domain is inserted in the sequence of an FP; (C) intramolecular reaction, as in chemiluminescence, where binding of the ligand causes the oxidation of the cofactor and photon emission. Class II: FRET-based sensors. Includes probes where ligand binding changes the FRET efficiency between the two chromophores. They are subdivided into (A) intermolecular FRET sensors; (B) intramolecular FRET sensors; and (C) BRET, the donor chromophore is a chemiluminescent protein and the acceptor is an FP. Class III: Translocation- or accumulation-based biosensors. (A) Translocation probe: the GFP-PKC example. The GFP-tagged protein kinase C accumulates at the PM upon increases in DG levels at the PM. (B) Accumulation probe: the TMRM example. This dye accumulates within mitochondria depending on their membrane potential. For other examples such as PM potential dyes that reorient in the plane of the membrane after changes or nonfluorescent probes, see Targeting organic sensors to subcellular compartments (PM section).