Fluorophore molecules could be either utilized alone, or serve as a fluorescent motif of a functional system. Based on molecular complexity and synthetic methods, fluorophore molecules could be generally classified into four categories: proteins and peptides, small organic compounds, synthetic oligomers and polymers, and multi-component systems.
Fluorescent proteins GFP, YFP, and RFP (green, yellow, and red, respectively) can be attached to other specific proteins to form a fusion protein, synthesized in cells after transfection of a suitable plasmid carrier.
Non-protein organic fluorophores belong to following major chemical families:
* Xanthene derivatives: fluorescein, rhodamine, Oregon green, eosin, and Texas red
* Cyanine derivatives: cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine, and merocyanine
* Squaraine derivatives and ring-substituted squaraines, including Seta and Square dyes
* Squaraine rotaxane derivatives: See Tau dyes
* Naphthalene derivatives (dansyl and prodan derivatives)
* Coumarin derivatives
* Oxadiazole derivatives: pyridyloxazole, nitrobenzoxadiazole, and benzoxadiazole
* Anthracene derivatives: anthraquinones, including DRAQ5, DRAQ7, and CyTRAK Orange
* Pyrene derivatives: cascade blue, etc.
* Oxazine derivatives: Nile red, Nile blue, cresyl violet, oxazine 170, etc.
* Acridine derivatives: proflavin, acridine orange, acridine yellow, etc.
* Arylmethine derivatives: auramine, crystal violet, malachite green
* Tetrapyrrole derivatives: porphin, phthalocyanine, bilirubin
* Dipyrromethene derivatives: BODIPY, aza-BODIPY
These fluorophores fluoresce due to delocalized electrons which can jump a band and stabilize the energy absorbed. For example, benzene, one of the simplest aromatic hydrocarbons, is excited at 254 nm and emits at 300 nm. This discriminates fluorophores from quantum dots, which are fluorescent semiconductor nanoparticles.
They can be attached to proteins to specific functional groups, such as amino groups (active ester, carboxylate, isothiocyanate, hydrazine), carboxyl groups (carbodiimide), thiol (maleimide, acetyl bromide), and organic azide (via click chemistry or non-specifically (glutaraldehyde)).
Additionally, various functional groups can be present to alter their properties, such as solubility, or confer special properties, such as boronic acid which binds to sugars or multiple carboxyl groups to bind to certain cations. When the dye contains an electron-donating and an electron-accepting group at opposite ends of the aromatic system, this dye will probably be sensitive to the environment's polarity (solvatochromic), hence called environment-sensitive. Often dyes are used inside cells, which are impermeable to charged molecules; as a result of this, the carboxyl groups are converted into an ester, which is removed by esterases inside the cells, e.g., fura-2AM and fluorescein-diacetate.
The following dye families are trademark groups, and do not necessarily share structural similarities.
* CF dye (Biotium)
* DRAQ and CyTRAK probes (BioStatus)
* BODIPY (Invitrogen)
* EverFluor (Setareh Biotech)
* Alexa Fluor (Invitrogen)
* Bella Fluor (Setareh Biotech)
* DyLight Fluor (Thermo Scientific, Pierce)
* Atto and Tracy (Sigma Aldrich)
* FluoProbes (Interchim)
* Abberior Dyes (Abberior)
* DY and MegaStokes Dyes (Dyomics)
* Sulfo Cy dyes (Cyandye)
* HiLyte Fluor (AnaSpec)
* Seta, SeTau and Square Dyes (SETA BioMedicals)
* Quasar and Cal Fluor dyes (Biosearch Technologies)
* SureLight Dyes (APC, RPEPerCP, Phycobilisomes) (Columbia Biosciences)
* APC, APCXL, RPE, BPE (Phyco-Biotech, Greensea, Prozyme, Flogen)
* Vio Dyes (Miltenyi Biotec)