Fluo-3 is a

fluorescence Fluorescence is the emission of light by a substance that has absorbed light or other electromagnetic radiation. It is a form of luminescence. In most cases, the emitted light has a longer wavelength, and therefore a lower photon energy, tha ...

indicator of intracellular

calcium Calcium is a chemical element with the symbol Ca and atomic number 20. As an alkaline earth metal, calcium is a reactive metal that forms a dark oxide-nitride layer when exposed to air. Its physical and chemical properties are most similar t ...

(Ca²⁺), developed by Roger Y. Tsien and colleagues. It is used to measure Ca²⁺ inside living cells in flow cytometry, and

confocal laser scanning microscopy Confocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser confocal scanning microscopy (LCSM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a sp ...

using visible light excitation (compatible with

argon laser An ion laser is a gas laser that uses an ionized gas as its lasing medium. Like other gas lasers, ion lasers feature a sealed cavity containing the laser medium and mirrors forming a Fabry–Pérot resonator. Unlike helium–neon lasers, the ...

sources operating at 488 nm). Fluo-3 and derivatives (Fluo-4, Fluo-5 etc) have also been widely used with

two-photon excitation microscopy Two-photon excitation microscopy (TPEF or 2PEF) is a fluorescence imaging technique that allows imaging of living tissue up to about one millimeter in thickness, with 0.64 μm lateral and 3.35 μm axial spatial resolution. Unlike traditional fl ...

. Fluo-3 is an essentially nonfluorescent compound, but upon binding of Ca²⁺ its fluorescence increases sharply with an emission maximum at 525 nm suitable for conventionally used detectors designed for

fluorescein isothiocyanate Fluorescein isothiocyanate (FITC) is a derivative of fluorescein used in wide-ranging applications including flow cytometry. First described in 1942, FITC is the original fluorescein molecule functionalized with an isothiocyanate reactive group ( ...

(FITC) measurements. This large change in fluorescence coupled with a good yield of photons provides very high contrast which allowed the detection of microscopic Ca²⁺ release events inside cells called "

Calcium sparks A calcium spark is the microscopic release of calcium ( Ca2+) from a store known as the sarcoplasmic reticulum (SR), located within muscle cells. This release occurs through an ion channel within the membrane of the SR, known as a ryanodine rece ...

". Whereas the salts of fluo-3 are unable to penetrate cells, loading can be achieved using its acetoxymethyl (AM)

ester In chemistry, an ester is a compound derived from an oxoacid (organic or inorganic) in which at least one hydroxyl group () is replaced by an alkoxy group (), as in the substitution reaction of a carboxylic acid and an alcohol. Glycerides a ...

derivative. Once inside the cell, unspecific

esterase An esterase is a hydrolase enzyme that splits esters into an acid and an alcohol in a chemical reaction with water called hydrolysis. A wide range of different esterases exist that differ in their substrate specificity, their protein structure ...

s cleave the ester effectively trapping fluo-3. As calcium is a key second messenger within cells, the specific properties of fluo-3 enable researchers to investigate the time-resolved dynamics of intracellular signal transduction in a diverse range of cells.Lambert, DG. ''Calcium Signaling Protocols''. Humana Press, 2006

References

{{Reflist Cell imaging Fluorone dyes Acetic acids Anilines Phenol ethers Chloroarenes