Tide Fluor™ dyes by AAT Bioquest are a series of highly water-soluble fluorescent reagents designed to label oligonucleotides and peptides for molecular imaging and diagnostic applications. Conjugates produced with Tide Fluor™ dyes exhibit exceptionally bright fluorescence and better photostability outperforming most conventional and proprietary dye-labeled conjugates, including fluorescein, cyanine, and most Alexa Fluor® dye-labeled conjugates. For optimum results in FRET biosensing, Tide Fluor™ dyes should be used collectively with complementary non-fluorescent Tide Quencher™ dyes
. The efficient FRET relationship between Tide Fluor™-Tide Quencher™ pairs facilitates the detection of molecular interactions within live cells at the nanometer range.
Tide Fluor™ Dyes for Labeling Oligonucleotides and Peptides
Tide Fluor™ dyes are available in 10 distinct emission colors with absorption spectra that span the UV to infrared spectrum and match the primary wavelengths of common excitation sources. The significantly stronger fluorescence and higher photostability exhibited by Tide Fluor™ dyes and their conjugates minimize photobleaching effects caused by high light intensities and long-term illumination giving researchers plenty of time to observe and capture images. Tide Fluor™ dyes are made available in a variety of conjugation chemistries, including amine-reactive (e.g., succinimidyl ester), thiol-reactive (.e.g., maleimide), and click chemistry formats (e.g., alkyne and azide) for labeling N-terminal, C-terminal, and cysteine residues. For in-synthesis labeling (e.g., solid-phase synthesis), Tide Fluor™ 1 CPG (Cat No. 2240
) and Tide Fluor™ 3 phosphoramidite dyes (Cat No. 2274
) are available in either 100 mg or 100 µmoles unit sizes, respectively. In addition, Tide Fluor™ 3-labeled FMOC amino acids are available for directly labeling aspartic acid, glutamic acid, and lysine residues during FMOC solid-phase synthesis.
Benefits of Tide Fluor™ dyes and their conjugates include:
- Efficient absorption of excitation light from common light sources
- pH-insensitive fluorescence between pH 3 to 11
- Brighter fluorescence and better photostability than conventional dyes and Alexa Fluor® dyes
- Exceptional FRET efficiency with Tide Quencher™ acceptor dyes
Table 1. Tide Fluor™ Dyes Spectral Properties
|Tide Fluor™ 1||345||442||20000||0.95||0.246||0.187|
|Tide Fluor™ 2WS||491||516||75000||0.9||0.211||0.091|
|Tide Fluor™ 2||500||527||75000||0.9||0.288||0.201|
|Tide Fluor™ 3WS||555||565||150000||0.105||0.079||0.079|
|Tide Fluor™ 3||555||584||85000||0.85||0.331||0.201|
|Tide Fluor™ 4||590||618||90000||0.91||0.489||0.436|
|Tide Fluor™ 5WS||649||664||250000||0.25||0.023||0.027|
|Tide Fluor™ 6WS||676||695||220000||0.18||0.111||0.009|
|Tide Fluor™ 7WS||749||775||275000||0.12||0.009||0.049|
|Tide Fluor™ 8WS||775||807||250000||0.08||0.103||0.109|
- ε = extinction coefficient (cm-1M-1) at their maximum absorption wavelength.
- Φ = fluorescence quantum yield in aqueous buffer (pH 7.2)
- CF at 260 nm is the correction factor used for eliminating the dye contribution to the absorbance at 260 nm (for oligos and nucleic acid labeling)
- CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptide and protein labeling)
Tide Fluor™ Dye Alternatives to Common Fluorophores
Tide Fluor™ dyes share nearly identical fluorescence spectra with common fluorophores, making upgrading to Tide Fluor™ dyes effortless. Traditional fluorophores and their conjugates can easily be replaced with spectrally similar Tide Fluor™ dyes without affecting instrument configuration (e.g., lasers and optical filters). For example, spectra of Tide Fluor™ 2WS and Tide Fluor™ 3WS dyes match those of fluorescein and Cy3 dyes and can be detected with standard fluorescein or TRITC/Cy3 filter sets, respectively. Each of the Tide Fluor™ dyes and their equivalents is listed below in Table 2.
Emission spectra of Tide Fluor™ dyes.
Table 2. Tide Fluor™ Dyes Common Equivalents
|EDANS||Tide Fluor™ 1 (TF1)|
|FAM, FITC, Alexa Fluor® 488||Tide Fluor™ 2 (TF2)|
|Alexa Fluor® 488||Tide Fluor™ 2WS (TF2WS)|
|Cy3®, Alexa Fluor® 555||Tide Fluor™ 3 (TF3)|
|Cy3®, Alexa Fluor® 555||Tide Fluor™ 3WS (TF3WS)|
|ROX, Texas Red®, Alexa Fluor® 594||Tide Fluor™ 4 (TF4)|
|Cy5®, Alexa Fluor® 647||Tide Fluor™ 5WS (TF5WS)|
|Cy5.5®, IRDye® 700, Alexa Fluor® 680||Tide Fluor™ 6WS (TF6WS)|
|Cy7®, Alexa Fluor® 750||Tide Fluor™ 7WS (TF7WS)|
|IRDye® 800||Tide Fluor™ 8WS (TF8WS)|
Tide Fluor™-Tide Quencher™ Pairs for Designing FRET Probes
Together Tide Fluor™ and Tide Quencher™ dyes can be used to produce highly sensitive FRET probes for a variety of applications, including enzyme studies, pharmacological drug screening, and qPCR analysis. Since the absorbances of Tide Quencher™ dyes are well-tuned to quench their Tide Fluor™ equivalent, substrates are completely non-fluorescent, and the potential for any background interference resulting from nonsensitized acceptor excitation is eliminated. Separating the two dyes either by proteolysis (i.e., FRET peptides) or nucleic acid hybridization (i.e., FRET nucleotides) restores fluorescence characteristic of the Tide Fluor™ label.
Table 3. Recommended FRET pairs for developing FRET oligonucleotides
|Tide Fluor™ 1||+++||+++||+||-||-||-||-||-|
Tide Fluor™ 2
Tide Fluor™ 3
|Tide Fluor™ 4||-||-||-||+||+++||+||-||-|
Tide Fluor™ 5
- TQ = Tide Quencher™
Tide Fluor™ and Tide Quencher™ M/Z Values
- Succinimidyl esters label biomolecules with primary amines (NH2) in slightly alkaline conditions to yield stable amide bonds.
- Maleimides react with sulfhydryl groups (-SH) at near-neutral conditions to form stable thioether linkages.
- Acids react with NH2 and OH groups.
- Amines conjugate to carboxylates.
- Alkynes react with azide groups via Cu(I)-catalyzed Alknye-Azide (CUAAC) or Cu(I)-free strain-promoted Alkyne-Azide Click Chemistry (SPAAC) reaction.
- Azides react with alkyne groups via Cu(I)-catalyzed Alknye-Azide (CUAAC) or Cu(I)-free strain-promoted Alkyne-Azide Click Chemistry (SPAAC) reaction.
- CPGs are conjugated to peptides and oligos during synthesis.
- DBCOs react with azide groups.
- Phosphoramidites are conjugated to peptides and oligos during synthesis.
Product ordering information
Table 4. Tide Fluor™ dyes for labeling peptides, oligonucleotides and other biomolecules.
|Tide Fluor™ 1 acid [TF1 acid] *Superior replacement for EDANS*||100 mg||2238|
|Tide Fluor™ 1 alkyne [TF1 alkyne]||5 mg||2237|
|Tide Fluor™ 1 amine [TF1 amine] *Superior replacement for EDANS*||5 mg||2239|
|Tide Fluor™ 1 azide [TF1 azide]||5 mg||2236|
|Tide Fluor™ 1 CPG [TF1 CPG] *1000 Å* *Superior replacement for EDANS*||100 mg||2241|
|Tide Fluor™ 1 CPG [TF1 CPG] *500 Å* *Superior replacement for EDANS*||100 mg||2240|
|Tide Fluor™ 1 maleimide [TF1 maleimide]||5 mg||2242|