Fluoresceins

Fluorescein derivatives are one of the most commonly used fluorescent labels for biological detection in flow cytometry and immunofluorescence due to their high absorptivity, excellent fluorescence quantum yields and good water solubility. The most widely used fluoresceins include fluorescein isothiocyanate (FITC) for labeling proteins (in particular antibodies) and carboxyfluoresceins (5-FAM and 5(6)-FAM) for labeling peptides and oligonucleotides. We offer a broad selection of fluorescein based-dyes, substrates and conjugates, as well as a series of superior iFluor® fluorescent labeling dyes with improved characteristics optimized for cellular labeling and detection.

Spectral Properties

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iFluor® 488 - superior alternative to FITC & Alexa Fluor® 488

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Despite its bright green fluorescence, fluorescein based-dyes and conjugates have a number of caveats that may restrict its use in certain applications, including rapid photobleaching, pH-sensitive fluorescence and susceptibility to quenching. The iFluor® 488 dye-with identical spectral properties - produces brighter, more photostable conjugates with pH-insensitive fluorescence over a wide molar range. iFluor® 488 dye molecules can be conjugated to biomolecules at high molar ratios with minimal self-quenching (Figure 2 and Figure 3).

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1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm^-1M^-1).

Fluorescein Products

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We offer a broad selection of fluorescein products including reactive fluorescein and fluorescein derivatives for labeling antibodies, nucleic acids and other biomolecules, conjugates, indicators, and substrates for detecting enzymatic activity in cells, homogenates and solution.

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Fluorescein Reactive Dyes

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Reactive fluorescent fluorescein and fluorescein derivatives are widely used to covalently label biomolecules such as antibodies, carbohydrates, nucleic acids, proteins, peptides and oligonucleotides. Resulting in intensely bright and photostable green fluorescence, fluorescein conjugates are often used as substrates for detecting enzymatic activity or to provide contrast in multicolored fluorescence investigation when combined with blue, yellow or red fluorescent probes. Fluorescein and fluorescein derivatives are readily available in a variety of dye-labeling chemistries, these include:

• Succinimidyl esters/NHS esters - for labeling free amines (-NH₂) on antibodies, proteins, peptides, amine-modified oligonucleotides and other biomolecules
• Maleimides - for labeling sulfhydryl groups on antibodies, proteins and peptides
• Azides - for labeling ethylene groups via click chemistry methods
• Alkynes - for labeling azides via click chemistry methods
• Hydrazides - for labeling aldehyde and ketone groups
• Carboxylic acids - for labeling amines after pre-activation with carbodiimides or for Steglich esterification of alcohols
• Amines - for labeling with various electrophilic compounds such as activated esters
• Tetrazines - for labeling trans-cyclooctenes (TCO) compounds via Inverse-Electron-Demands Diels-Alder reaction
• Phosphoramidite - for labeling oligonucleotides

1. ε = molar extinction coefficient at their maximum absorption wavelength (Units = cm^-1M^-1).

Fluorescein-Labeled Enzyme Substrates

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Fluorescein and fluorescein derivatives are frequently used to prepare fluorogenic substrates for detecting enzymatic activity in cells and solutions. When hydrolyzed by the enzyme of interest, fluorescein-modified substrates exhibit an increase in fluorescence intensity that is linearly proportional to enzyme activity. AAT Bioquest provides a large assortment of fluorescein modified substrates for detecting activity from caspases, proteases, peptidases, phosphatases and more.

Fluorescein-Labeled Conjugates

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Fluorescein-labeled conjugates have a wide range of applications in bio-imaging and bio-labelling due to their bright green fluorescence and excellent photostability. Fluorescein derivatives can be labeled to antibodies, streptavidin, phalloidin, annexin v, dextran and other biomolecules to study apoptosis, label organelles or improve assay sensitivity by amplifying weak signals.

Fluorescein-Derived Indicators

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Fluorescein-derived indicators are important tools in biochemical and cellular studies. They have been used in all major types of fluorescence imaging applications including flow cytometry, fluorescence microscopy and fluorescence plate-based assays. AAT Bioquest offers a wide range of fluorescein-derived indicators for measuring cellular pH, calcium concentrations, cell proliferation, cell viability and more.