Fluorescein isothiocyanate (FITC) is a derivative of
fluorescein
Fluorescein is an organic compound and dye based on the xanthene tricyclic structural motif, formally belonging to triarylmethine dyes family. It is available as a dark orange/red powder slightly soluble in water and alcohol. It is widely used ...
used in wide-ranging applications including
flow cytometry. First described in 1942, FITC is the original fluorescein molecule
functionalized with an
isothiocyanate
In organic chemistry, isothiocyanate is the functional group , formed by substituting the oxygen in the isocyanate group with a sulfur. Many natural isothiocyanates from plants are produced by enzymatic conversion of metabolites called glucosi ...
reactive group (−N=C=S), replacing a
hydrogen
Hydrogen is the chemical element with the symbol H and atomic number 1. Hydrogen is the lightest element. At standard conditions hydrogen is a gas of diatomic molecules having the formula . It is colorless, odorless, tasteless, non-toxic ...
atom on the bottom ring of the structure. It is typically available as a mixture of
isomers, fluorescein 5-isothiocyanate (5-FITC) and fluorescein 6-isothiocyanate (6-FITC). FITC is reactive towards nucleophiles including
amine
In chemistry, amines (, ) are compounds and functional groups that contain a basic nitrogen atom with a lone pair. Amines are formally derivatives of ammonia (), wherein one or more hydrogen
Hydrogen is the chemical element wi ...
and
sulfhydryl groups on
protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, res ...
s. It was synthesized by Robert Seiwald and Joseph Burckhalter in 1958.
A
succinimidyl-ester functional group attached to the fluorescein core, creating "NHS-fluorescein", forms another common amine reactive derivative that has much greater specificity toward primary amines in the presence of other nucleophiles.
FITC has
excitation
Excitation, excite, exciting, or excitement may refer to:
* Excitation (magnetic), provided with an electrical generator or alternator
* Excite Ballpark, located in San Jose, California
* Excite (web portal), web portal owned by IAC
* Electron exc ...
and
emission spectrum peak wavelengths of approximately 495 nm and 519 nm,
giving it a green color. Like most
fluorochrome
A fluorophore (or fluorochrome, similarly to a chromophore) is a fluorescent chemical compound that can re-emit light upon light excitation. Fluorophores typically contain several combined aromatic groups, or planar or cyclic molecules with sev ...
s, it is prone to
photobleaching
In optics, photobleaching (sometimes termed fading) is the photochemical alteration of a dye or a fluorophore molecule such that it is permanently unable to fluoresce. This is caused by cleaving of covalent bonds or non-specific reactions between ...
. Due to the problem of photobleaching, derivatives of fluorescein such as
Alexa 488 and
DyLight 488 have been tailored for various chemical and biological applications where greater
photostability
Photochemistry is the branch of chemistry concerned with the chemical effects of light. Generally, this term is used to describe a chemical reaction caused by absorption of ultraviolet (wavelength from 100 to 400 nm), visible light (400–7 ...
, higher fluorescence intensity, or different attachment groups are needed. In addition, some experiments make use of FITC's propensity for photobleaching in order to measure proteins' lateral mobility in membranes, through the technique of
fluorescence recovery after photobleaching
Fluorescence recovery after photobleaching (FRAP) is a method for determining the kinetics of diffusion through tissue or cells. It is capable of quantifying the two dimensional lateral diffusion of a molecularly thin film containing fluorescently ...
.
References
{{DEFAULTSORT:Fluorescein Isothiocyanate
Benzoic acids
Fluorone dyes
Isothiocyanates
Staining dyes