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Fluorescent CF® Dye tyramides are used for tyramide signal amplification (TSA) for increasing immunofluorescence sensitivity in multicolor immunocytochemistry (ICC), immunohistochemistry (IHC), or in situ hybridization (ISH).
CF® Dye tyramide conjugates are used for tyramide signal amplification (TSA), a method for high-density labeling of a target protein or nucleic acid in situ.
Also learn about TyraMax™ Amplification Dyes and Kits, Biotium’s next generation tyramide dyes that offer brighter signal compared to the original CF® Dye Tyramides, and have advantages in brightness, photostability, and working solution stability compared to other TSA dyes. We also offer Ready-to-Use Tyramide Amplification Buffer, Tyramide Amplification Buffer Plus (an improved formulation for enhanced TSA sensitivity), and CF® Dye Tyramide Amplification Kits.
Biotium’s next-generation CF® Dyes were designed to be highly water-soluble with advantages in brightness and photostability compared to other commercially available fluorescent dyes. Our CF® Dye Tyramide conjugates are available in 22 colors. Learn more about CF® Dyes.
For researchers considering near-infrared detection, we recommend CF®740 tyramide over CF®750 and CF®754. This is because CF®750 is unstable in oxidizing amplification buffer and should be added to the buffer immediately before performing the staining reaction. The poor stability in oxidizing amplification buffer makes the dye challenging to use for automated staining platforms (ie. BOND RX) that require longer periods with the dye in buffer. CF®754 tyramide is stable in oxidizing amplification buffer, but the dye has a broad absorption peak that can cause channel spillover. CF®740 tyramide was developed to be stable in oxidizing amplification buffer when compared to Alexa Fluor® 750 and CF®750. In addition, CF®740 tyramide has a narrower absorption peak that minimizes spillover and therefore a superior option to CF®754 tyramide. Near-IR tyramide background staining may be tissue dependent and not suitable for all targets. In general, we would recommend using near-IR staining for more abundant targets in your panel.

CF®740 free acid or Alexa Fluor® 750 free acid was diluted to 2 uM in 1X Tyramide Amplification Buffer with 0.0015% hydrogen peroxide and incubated at room temperature, protected from light. Dye concentration was measured by absorbance at the time points shown. CF®740 remained stable in the presence of hydrogen peroxide over 24 hours (≤90% of the starting dye concentration), while Alexa Fluor® 750 degraded to less than 10% of the starting concentration after 24 hours.
TSA is a highly sensitive method for differential gene or protein analysis or detection of low-abundance targets, in fluorescent ICC, IHC, and FISH applications. An antibody- or streptavidin-HRP conjugate catalyzes the deposition of fluorescent dye/biotin tyramides on tyrosine residues on and adjacent to a target protein or nucleic acid sequence in situ. This results in high-density labeling of the target and significantly improves the detection sensitivity up to 100-fold compared to conventional methods. TSA is particularly advantageous for fluorescence detection in human tissue, where conventional ICC or FISH often fails to provide adequate signal over autofluorescence background. In applications where increased sensitivity is not required, TSA enables the use of significantly lower antibody or probe concentrations for the same level of detection sensitivity thereby reducing issues of non-specific binding or cross-reactivity. Furthermore, since binding of the tyramide label is covalent, a large number of targets can be detected in the same sample using multiple rounds of sequential TSA, in which the availability of antibodies from different host species is not a limitation. TSA also can be easily integrated with conventional immunostaining. Learn more about Tyramide Signal Amplification.
| Product | Ex/Em | MW (g/mol) | Size | Catalog No. | Dye Features |
|---|---|---|---|---|---|
| CF®350 Tyramide | 347/448 nm | ~614 | 0.5 mg | 92170 | CF®350 Features |
| CF®405S Tyramide | 404/431 nm | ~689 | 0.5 mg | 92197 | CF®405S Features |
| CF®405M Tyramide | 408/452 nm | ~621 | 0.5 mg | 96057 | CF®405M Features |
| CF®405L Tyramide | 395/545 nm | ~1692 | 0.5 mg | 92198 | CF®405L Features |
| CF®430 Tyramide | 426/498 nm | ~707 | 0.5 mg | 96053 | CF®430 Features |
| CF®488A Tyramide | 490/515 nm | ~666 | 0.5 mg | 92171 | CF®488A Features |
| CF®514 Tyramide | 516/548 nm | ~1337 | 0.5 mg | 92199 | CF®514 Features |
| CF®532 Tyramide | 527/558 nm | ~804 | 0.5 mg | 96066 | CF®532 Features |
| CF®543 Tyramide | 541/560 nm | ~1006 | 0.5 mg | 92172 | CF®543 Features |
| CF®550R Tyramide | 551/577 nm | ~806 | 0.5 mg | 96077 | CF®550R Features |
| CF®555 Tyramide | 555/565 nm | ~1120 | 0.5 mg | 96021 | CF®555 Features |
| CF®568 Tyramide | 562/583 nm | ~833 | 0.5 mg | 92173 | CF®568 Features |
| CF®583R Tyramide | 586/609 nm | ~892 | 0.5 mg | 96085 | CF®583R Features |
| CF®594 Tyramide | 593/614 nm | ~848 | 0.5 mg | 92174 | CF®594 Features |
| CF®620R Tyramide | 617/639 nm | ~857 | 0.5 mg | 92194 | CF®620R Features |
| CF®640R Tyramide | 642/662 nm | ~951 | 0.5 mg | 92175 | CF®640R Features |
| CF®647 Tyramide | 650/665 nm | ~1104 | 0.5 mg | 96022 | CF®647 Features |
| CF®660R Tyramide | 663/682 nm | ~1007 | 0.5 mg | 92195 | CF®660R Features |
| CF®680R Tyramide | 680/701 nm | ~1031 | 0.5 mg | 92196 | CF®680R Features |
| CF®710 Tyramide | 712/736 nm | ~977 | 0.5 mg | 96127 | CF®710 Features |
| CF®725 Tyramide | 729/750 nm | ~1005 | 0.5 mg | 96128 | CF®725 Features |
| CF®740 Tyramide | 742/767 nm | ~1005 | 0.5 mg | 96124 | CF®740 Features |
| CF®750 Tyramide* | 755/779 nm | ~3040 | 0.5 mg | 96052 | CF®750 Features |
| CF®754 Tyramide | 748/793 nm | ~1000 | 0.5 mg | 96090 |
* CF®750 Tyramide is not stable in TSA buffer, and should be added to the buffer immediately before performing the staining reaction.
1. Journal of Immunology Research (2020), 2020:2328675. DOI:10.1155/2020/2328675
2. Scientific Reports (2019), 9:1144. DOI:10.1038/s41598-018-38171-5
3. bioRxiv (2019) preprint. DOI:10.1101/651083
4. OncoImmunology (2019), 8(6):e1581528-10. DOI:10.1080/2162402X.2019.1581528
5. Microbiological Research (2018), 217: 69-80. DOI:10.1016/j.micres.2018.08.017
6. Mol Neurodegeneration (2017), 12:68. DOI:10.1186/s13024-017-0202-z
7. Biology Open (2017), 6: 891-896. DOI:10.1242/bio.025809
8. The American Journal of Pathology (2016), 186(10):2650-2664. DOI:10.1016/j.ajpath.2016.06.020
Download a list of CF® Dye references.
1. Journal of Immunology Research (2020), 2020:2328675. DOI:10.1155/2020/2328675
2. Scientific Reports (2019), 9:1144. DOI:10.1038/s41598-018-38171-5
3. bioRxiv (2019) preprint. DOI:10.1101/651083
4. OncoImmunology (2019), 8(6):e1581528-10. DOI:10.1080/2162402X.2019.1581528
5. Microbiological Research (2018), 217: 69-80. DOI:10.1016/j.micres.2018.08.017
6. Mol Neurodegeneration (2017), 12:68. DOI:10.1186/s13024-017-0202-z
7. Biology Open (2017), 6: 891-896. DOI:10.1242/bio.025809
8. The American Journal of Pathology (2016), 186(10):2650-2664. DOI:10.1016/j.ajpath.2016.06.020
Download a list of CF® Dye references.
Bioscience kits
The guaranteed shelf life from date of receipt for bioscience kits is listed on the product information sheet. Some kits have an expiration date printed on the kit box label, this is the guaranteed shelf life date calculated from the day that the product shipped from our facility. Kits often are functional for significantly longer than the guaranteed shelf life. If you have an older kit in storage that you wish to use, we recommend performing a small scale positive control experiment to confirm that the kit still works for your application before processing a large number of samples or precious samples.
Antibodies and other conjugates
The guaranteed shelf life from date of receipt for antibodies and conjugates is listed on the product information sheet. Antibodies and other conjugates often are functional for significantly longer than the guaranteed shelf life. If you have an older conjugate in storage that you wish to use, we recommend performing a small scale positive control experiment to confirm that the product still works for your application before processing a large number of samples or precious samples.
For lyophilized antibodies, we recommend reconstituting the antibody with glycerol and antimicrobial preservative like sodium azide for the longest shelf life (note that sodium azide is not compatible with HRP-conjugates).
Chemicals, dyes, and gel stains
Biotium guarantees the stability of chemicals, dyes, and gel stains for at least a year from the date you receive the product. However, the majority of these products are highly stable for many years, as long as they are stored as recommended. Storage conditions can be found on the product information sheet or product safety and data sheet, material safety data sheet, and on the product label. Fluorescent compounds should be protected from light for long term storage.
If you have a Biotium compound that has been in storage for longer than one year that you wish to use, we recommend performing a small scale positive control experiment to confirm that the compound still works for your application before processing a large number of samples or precious samples.
Expiration date based on date of manufacture (DOM)
If your institution requires you to document expiration date based on date of manufacture for reagents, please contact techsupport@biotium.com for assistance.
Chemical products with special stability considerations:
Esters
Ester compounds include the following:
Ester dyes are stable in solid form as long as they are protected from light and moisture. Esters are not stable in aqueous solution. Concentrated stock solutions should be prepared in anhydrous DMSO (see Biotium catalog no. 90082). Stock solutions in anhydrous DMSO can be stored desiccated at -20°C for one month or longer. Esters should be diluted in aqueous solution immediately before use. Succinimidyl esters (SE) should be dissolved in a solution that is free of amine-containing compounds like Tris, glycine, or protein, which will react with the SE functional group. AM esters and diacetate compounds should be dissolved in a solution that is free of serum, because serum could contain esterases that would hydrolyze the compound.
A note on CF® Dye succinimidyl ester stability
Succinimidyl esters (SE) are generally susceptible to hydrolysis, which can result in lower labeling efficiency. Many commercially available fluorescent dyes used for life science research are heavily sulfonated dyes which makes them particularly hygroscopic, worsening the hydrolysis problem. In addition, for several commercially available SE reactive dyes, the SE group is derived from an aromatic carboxylic acid, while the SE group in all of Biotium’s CF® Dyes is prepared from an aliphatic carboxylic acid. This structural difference reduces the susceptibility of CF® Dye SE reactive groups to hydrolysis, resulting in relatively stable reactive dyes with consistently higher labeling efficiency compared to other SE derivatives of other fluorescent dyes.
Maleimides, MTS and thiosulfate dyes
Like the succinimidyl ester dyes, these dyes are also susceptible to hydrolysis, although generally to a much lower degree. Thus, for long term storage, anhydrous DMSO is recommended for making stock solutions.
Other reactive dyes
Amines, aminooxy (also known as oxylamine), hydrazide, azide, alkyne, BCN, and tyramide reactive dyes, as well as dye free acids, are generally stable in aqueous solution when stored at -20°C for 6-12 months or longer, as long as no compounds are present that may react with the dye’s functional group. See the product information sheets for specific reactive dyes more information.
Coelenterazines and D-luciferin
Coelenterazines are stable in solid form when stored as recommended; they are not stable in aqueous solution. Concentrated coelenterazine stock solutions (typically 1-100 mg/mL) should be prepared in ethanol or methanol; do not use DMSO or DMF to dissolve coelenterazines, because these solvents will oxidize the compounds. Ethanol or methanol stocks of coelenterazine can be stored at -20°C or below for six months or longer; alcohol stocks may evaporate during storage, so use tightly sealing screw cap vials and wrap the vials with Parafilm for long term storage. Propylene glycol also can be used as a solvent to minimize evaporation. If the solvent evaporates, the coelenterazine will still be present in the vial, so note the volume in the vial prior to storage so that you can adjust the solvent volume to correct for evaporation if needed. Prepare working solutions in aqueous buffers immediately before use. Coelenterazines are stable for up to five hours in aqueous solution.
Aquaphile™ coelenterazines are water soluble formulations of coelenterazines. They are stable in solid form when stored as recommended. Aquaphile™ coelenterazines should be dissolved in aqueous solution immediately before use. They are stable for up to five hours in aqueous solution.
Note that coelenterazines are predominantly yellow solids, but may contain dark red or brown flecks. This does not affect product stability or performance. If your coelenterazine is uniformly brown, then it is oxidized and needs to be replaced.
D-luciferin is stable in solid form and as a concentrated stock solution when stored as recommended; it is not stable at dilute working concentrations in aqueous solution. Prepare concentrated D-luciferin stock solutions (typically 1-100 mg/mL) in water, and store in aliquots at -20°C or below for six months or longer. Prepare working solutions immediately before use.
For dyes or reagents that are supplied lyophilized (as solids), it is hard to compare quantities based on appearance of the dye in the tube, because during the lyophilization process the dye can dry down in different ways, either spread out all over the tube, clumped together, or coating the sides or bottom of the tube. Centrifugation of the tube may not help in collecting the dye solid to the bottom of the tube as this generally works for solutions. However, lyophilized solids are packaged based on highly accurate absorbance measurement of the reagent solution prior to drying, so the vial will contain the correct amount of dye.
Biotium ships all antibodies (primary, secondary and conjugates) at room temperature. We guarantee their quality and performance under these conditions based upon our stability testing. Antibodies were subjected to accelerated stability testing by storing them at various temperatures (4°C, room temperature, or 37°C) for 1 week to mimic simulated shipping conditions and tested in immunostaining experiments. All antibodies showed the expected brightness and specificity, even after storage at sub-optimal temperatures for a week or longer. You can also download our Product Storage Statement here.
In line with our goal to be more environmentally friendly by reducing the use of excess packaging, and lowering shipping costs for our customers, products that have passed our stability testing are shipped at room temperature.
Once you have received the antibody vial, please follow the long-term storage instructions on the product information (PI) sheet.