donor protein emit light with the same wavelengths. Yet researchers can detect differences in the polarisation between the light which excites the fluorophores and the light which is emitted, in a technique called FRET anisotropy imaging; the level of quantified anisotropy (difference in polarisation between the excitation and emission beams) then becomes an indicative guide to how many FRET events have happened. Others Various compounds beside fluorescent proteins. Applications The applications of fluorescence resonance energy transfer (FRET) have expanded tremendously in the last 25 years, and the technique has become a staple in many biological and biophysical fields. FRET can be used as a spectroscopic ruler to measure distance and detect molecular interactions in a number of systems and has applications in biology and biochemistry. Proteins FRET is often used to detect and track interactions between proteins. Additionally, FRET can be used to measure distances between domains in a single protein by tagging different regions of the protein with fluorophores and measuring emission to determine distance. This provides information about protein conformation, including secondary structures and protein folding. This extends to tracking functional changes in protein structure, such as conformational changes associated with myosin activity. Applied in vivo, FRET has been used to detect the location and interactions of cellular structures including integrins and membrane proteins. Membranes FRET can be used to observe membrane fluidity, movement and dispersal of membrane proteins, membrane lipid-protein and protein-protein interactions, and successful mixing of different membranes. FRET is also used to study formation and properties of membrane domains and lipid rafts in cell membranes and to determine surface density in membranes. Chemosensory FRET-based probes can detect the presence of various molecules: the probe's structure is affected by small molecule binding or activity, which can turn the FRET system on or off. This is often used to detect anions, cations, small uncharged molecules, and some larger biomacromolecules as well. Similarly, FRET systems have been designed to detect changes in the cellular environment due to such factors as pH, hypoxia, or mitochondrial membrane potential. Signaling pathways Another use for FRET is in the study of metabolic or signaling pathways. For example, FRET and BRET have been used in various experiments to characterize G-protein coupled receptor activation and consequent signaling mechanisms. Other examples include the use of FRET to analyze such diverse processes as bacterial chemotaxis and caspase activity in apoptosis. Proteins and nucleotides folding kinetics Proteins,