iFluor® 405 Styramide
Ordering information
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Additional ordering information
Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Physical properties
Molecular weight | 1174.49 |
Solvent | DMSO |
Spectral properties
Correction Factor (260 nm) | 0.48 |
Correction Factor (280 nm) | 0.77 |
Extinction coefficient (cm -1 M -1) | 370001 |
Excitation (nm) | 403 |
Emission (nm) | 427 |
Quantum yield | 0.911 |
Storage, safety and handling
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
Storage | Freeze (< -15 °C); Minimize light exposure |
UNSPSC | 12171501 |
Overview | ![]() ![]() |
Molecular weight 1174.49 | Correction Factor (260 nm) 0.48 | Correction Factor (280 nm) 0.77 | Extinction coefficient (cm -1 M -1) 370001 | Excitation (nm) 403 | Emission (nm) 427 | Quantum yield 0.911 |
Power Styramide™ Signal Amplification (PSA™) system is one of the most sensitive methods that can detect extremely low-abundance targets in cells and tissues with improved fluorescence signal 10-50 times higher than the widely used tyramide (TSA) reagents. iFluor® dye-labeled Styramide™ conjugates can generate fluorescence signal with significantly higher precision and sensitivity (more than 100 times) than standard ICC/IF/IHC. PSA utilizes the catalytic activity of horseradish peroxidase (HRP) for covalent deposition of fluorophores in situ. PSA radicals have much higher reactivity than tyramide radicals, making the PSA system much faster, more robust and sensitive than the traditional TSA reagents. iFluor® 405 Styramide is a new unique blue fluorescent PSA reagent for multicolor application with our existing PSA and TSA reagents. It has the maximum fluorescence ~421 nm. iFluor® 405 Styramide is a superior replacement for Alexa Fluor 405 tyramide or other spectrally similar fluorescent tyramide conjugates or TSA reagents. AAT Bioquest offers the largest collection of TSA regents. We are the exclusive source of the superior PSA reagents for multicolor applications.
Platform
Fluorescence microscope
Excitation | DAPI/Violet filter set |
Emission | DAPI/Violet filter set |
Recommended plate | Black wall/clear bottom |
Example protocol
AT A GLANCE
Protocol Summary
- Fix/permeabilize/block cells or tissue
- Add primary antibody in blocking buffer
- Add HRP-conjugated secondary antibody
- Prepare Styramide™ working solution and apply in cells or tissue for 5-10 minutes at room temperature
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.
Note Make single use aliquots, and store unused 100X stock solution at 2-8 oC in dark place.
Note Prepare the 100X H2O2 solution fresh on the day of use.
1. iFluor™ 405 Styramide stock solution (100X)
Add 100 µL of DMSO into the vial of iFluor™ 405 Styramide conjugate to make 100X Styramide stock solution.Note Make single use aliquots, and store unused 100X stock solution at 2-8 oC in dark place.
2. H2O2 stock solution
Add 10 µL of 3% hydrogen peroxide (Not provided) to 90 µL of ddH2O.Note Prepare the 100X H2O2 solution fresh on the day of use.
PREPARATION OF WORKING SOLUTION
1. iFluor™ 405 Styramide working solution (1X)
Every 1 mL of Reaction Buffer requires 10 µL of Styramide stock solution and 10 µL of H2O2 stock solution.Note The Styramide provided is enough for 100 tests based on 100 µL of Styramide working solution needed per coverslip or per well in a 96-well microplate.
Note The Styramide working solution must be used within 2 hours after preparation and avoid direct exposure to light.
2. Secondary antibody-HRP working solution
Make appropriate concentration of secondary antibody-HRP working solution as per the manufacturer's recommendations.SAMPLE EXPERIMENTAL PROTOCOL
This protocol is applicable for both cells and tissues staining.
Protocol can be found at
https://www.aatbio.com/resources/guides/paraffin-embedded-tissue-immunohistochemistry-protocol.html
Cell fixation and permeabilization
- Fix the cells or tissue with 3.7% formaldehyde or paraformaldehyde, in PBS at room temperature for 20 minutes.
- Rinse the cells or tissue with PBS twice.
- Permeabilize the cells with 0.1% Triton X-100 solution for 1-5 minutes at room temperature.
- Rinse the cells or tissue with PBS twice.
Tissue fixation, deparaffinization and rehydration
Deparaffinize and dehydrate the tissue according to the standard IHC protocols. Perform antigen retrieval with preferred specific solution/protocol as needed.Protocol can be found at
https://www.aatbio.com/resources/guides/paraffin-embedded-tissue-immunohistochemistry-protocol.html
Peroxidase labeling
- Optional: Quench endogenous peroxidase activity by incubating cell or tissue sample in peroxidase quenching solution (such as 3% hydrogen peroxide) for 10 minutes. Rinse with PBS twice at room temperature.
- Optional: If using HRP-conjugated streptavidin, it is advisable to block endogenous biotins by biotin blocking buffer.
- Block with preferred blocking solution (such as PBS with 1% BSA) for 30 minutes at 4 °C.
- Remove blocking solution and add primary antibody diluted in recommended antibody diluent for 60 minutes at room temperature or overnight at 4 °C.
- Wash with PBS three times for 5 minutes each.
- Apply 100 µL of secondary antibody-HRP working solution to each sample and incubate for 60 minutes at room temperature.
Note Incubation time and concentration can be varied depending on the signal intensity. - Wash with PBS three times for 5 minutes each.
Styramide labeling
- Prepare and apply 100 µL of Styramide working solution to each sample and incubate for 5-10 minutes at room temperature.
Note If you observe non-specific signal, you can shorten the incubation time with Styramide. You should optimize the incubation period using positive and negative control samples at various incubation time points. Or you can use lower concentration of Styramide in the working solution. - Rinse with PBS three times.
Counterstain and fluorescence imaging
- Counterstain the cell or tissue samples as needed. AAT provides a series of nucleus counterstain reagents as listed in Table 1. Follow the instruction provided with the reagents.
- Mount the coverslip using a mounting medium with anti-fading properties.
- Use the appropriate filter set to visualize the signal from the Styramide labeling.
Cat# | Product Name | Ex/Em (nm) |
17548 | Nuclear Blue™ DCS1 | 350/461 |
17550 | Nuclear Green™ DCS1 | 503/526 |
17551 | Nuclear Orange™ DCS1 | 528/576 |
17552 | Nuclear Red™ DCS1 | 642/660 |
Calculators
Common stock solution preparation
Table 1. Volume of DMSO needed to reconstitute specific mass of iFluor® 405 Styramide to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 85.143 µL | 425.717 µL | 851.433 µL | 4.257 mL | 8.514 mL |
5 mM | 17.029 µL | 85.143 µL | 170.287 µL | 851.433 µL | 1.703 mL |
10 mM | 8.514 µL | 42.572 µL | 85.143 µL | 425.717 µL | 851.433 µL |
Molarity calculator
Enter any two values (mass, volume, concentration) to calculate the third.
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
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Spectrum
Open in Advanced Spectrum Viewer


Spectral properties
Correction Factor (260 nm) | 0.48 |
Correction Factor (280 nm) | 0.77 |
Extinction coefficient (cm -1 M -1) | 370001 |
Excitation (nm) | 403 |
Emission (nm) | 427 |
Quantum yield | 0.911 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield | Correction Factor (260 nm) | Correction Factor (280 nm) |
iFluor® 405 amine | 403 | 427 | 370001 | 0.911 | 0.48 | 0.77 |
iFluor® 405 hydrazide | 403 | 427 | 370001 | 0.911 | 0.48 | 0.77 |
iFluor® 405 maleimide | 403 | 427 | 370001 | 0.911 | 0.48 | 0.77 |
iFluor® 350 Styramide *Superior Replacement for Alexa Fluor 350 tyramide* | 345 | 450 | 200001 | 0.951 | 0.83 | 0.23 |
iFluor® 488 Styramide *Superior Replacement for Alexa Fluor 488 tyramide and Opal 520* | 491 | 516 | 750001 | 0.91 | 0.21 | 0.11 |
iFluor® 546 Styramide *Superior Replacement for Alexa Fluor 546 tyramide* | 541 | 557 | 1000001 | 0.671 | 0.25 | 0.15 |
iFluor® 555 Styramide *Superior Replacement for Alexa Fluor 555 tyramide and Opal 570* | 557 | 570 | 1000001 | 0.641 | 0.23 | 0.14 |
iFluor® 568 Styramide *Superior Replacement for Alexa Fluor 568 tyramide* | 568 | 587 | 1000001 | 0.571 | 0.34 | 0.15 |
iFluor® 594 Styramide *Superior Replacement for Alexa Fluor 594 tyramide* | 588 | 604 | 1800001 | 0.531 | 0.05 | 0.04 |
Show More (16) |
Images

Figure 1. Fixed and permeabilized HeLa cells were incubated with rabbit anti-tubulin antibody, then labeled with HRP-labeled Goat anti-Rabbit IgG (Cat No. 16793), and detected using iFluor® 405 styramide (Cat No. 44908). Images were captured on a fluorescence microscope equipped with a DAPI filter set.
References
View all 50 references: Citation Explorer
Immunofluorescent Staining of Adult Murine Paraffin-Embedded Skeletal Tissue.
Authors: Felsenthal, Neta and Zelzer, Elazar
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 337-344
Authors: Felsenthal, Neta and Zelzer, Elazar
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 337-344
Highly Sensitive and Multiplexed In Situ RNA Profiling with Cleavable Fluorescent Tyramide.
Authors: Xiao, Lu and Labaer, Joshua and Guo, Jia
Journal: Cells (2021)
Authors: Xiao, Lu and Labaer, Joshua and Guo, Jia
Journal: Cells (2021)
Single-cell RNA sequencing of human liver reveals hepatic stellate cell heterogeneity.
Authors: Payen, Valéry L and Lavergne, Arnaud and Alevra Sarika, Niki and Colonval, Megan and Karim, Latifa and Deckers, Manon and Najimi, Mustapha and Coppieters, Wouter and Charloteaux, Benoît and Sokal, Etienne M and El Taghdouini, Adil
Journal: JHEP reports : innovation in hepatology (2021): 100278
Authors: Payen, Valéry L and Lavergne, Arnaud and Alevra Sarika, Niki and Colonval, Megan and Karim, Latifa and Deckers, Manon and Najimi, Mustapha and Coppieters, Wouter and Charloteaux, Benoît and Sokal, Etienne M and El Taghdouini, Adil
Journal: JHEP reports : innovation in hepatology (2021): 100278
Multiplexed In Situ Protein Profiling with High-Performance Cleavable Fluorescent Tyramide.
Authors: Pham, Thai and Liao, Renjie and Labaer, Joshua and Guo, Jia
Journal: Molecules (Basel, Switzerland) (2021)
Authors: Pham, Thai and Liao, Renjie and Labaer, Joshua and Guo, Jia
Journal: Molecules (Basel, Switzerland) (2021)
Accessibility-dependent topology studies of membrane proteins using a SpyTag/SpyCatcher protein-ligation system.
Authors: Bae, Yoonji and Lee, Sang Kwon and Chae, Young Chan and Park, Chan Young and Kang, Sebyung
Journal: International journal of biological macromolecules (2021): 171-178
Authors: Bae, Yoonji and Lee, Sang Kwon and Chae, Young Chan and Park, Chan Young and Kang, Sebyung
Journal: International journal of biological macromolecules (2021): 171-178
Immunohistochemical Detection of 5-Hydroxymethylcytosine and 5-Carboxylcytosine in Sections of Zebrafish Embryos.
Authors: Jessop, Peter and Gering, Martin
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 193-208
Authors: Jessop, Peter and Gering, Martin
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 193-208
Phenoxy Radical Reactivity of Nucleic Acids: Practical Implications for Biotinylation.
Authors: Wilbanks, Brandon and Garcia, Brian and Byrne, Shane and Dedon, Peter and Maher, L James
Journal: Chembiochem : a European journal of chemical biology (2021): 1400-1404
Authors: Wilbanks, Brandon and Garcia, Brian and Byrne, Shane and Dedon, Peter and Maher, L James
Journal: Chembiochem : a European journal of chemical biology (2021): 1400-1404
Adoptive cell therapy of triple negative breast cancer with redirected cytokine-induced killer cells.
Authors: Sommaggio, Roberta and Cappuzzello, Elisa and Dalla Pietà, Anna and Tosi, Anna and Palmerini, Pierangela and Carpanese, Debora and Nicolè, Lorenzo and Rosato, Antonio
Journal: Oncoimmunology (2020): 1777046
Authors: Sommaggio, Roberta and Cappuzzello, Elisa and Dalla Pietà, Anna and Tosi, Anna and Palmerini, Pierangela and Carpanese, Debora and Nicolè, Lorenzo and Rosato, Antonio
Journal: Oncoimmunology (2020): 1777046
A simple, real-time assay of horseradish peroxidase using biolayer interferometry.
Authors: Kojima, Takaaki and Nakane, Ayako and Zhu, Bo and Alfi, Almasul and Nakano, Hideo
Journal: Bioscience, biotechnology, and biochemistry (2019): 1822-1828
Authors: Kojima, Takaaki and Nakane, Ayako and Zhu, Bo and Alfi, Almasul and Nakano, Hideo
Journal: Bioscience, biotechnology, and biochemistry (2019): 1822-1828
The EMARS Reaction for Proximity Labeling.
Authors: Honke, Koichi and Miyagawa-Yamaguchi, Arisa and Kotani, Norihiro
Journal: Methods in molecular biology (Clifton, N.J.) (2019): 1-12
Authors: Honke, Koichi and Miyagawa-Yamaguchi, Arisa and Kotani, Norihiro
Journal: Methods in molecular biology (Clifton, N.J.) (2019): 1-12
Application notes
A Meta-Analysis of Common Calcium Indicators
A New Protein Crosslinking Method for Labeling and Modifying Antibodies
A Novel Fluorescent Probe for Imaging and Detecting Hydroxyl Radical in Living Cells
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Abbreviation of Common Chemical Compounds Related to Peptides
A New Protein Crosslinking Method for Labeling and Modifying Antibodies
A Novel Fluorescent Probe for Imaging and Detecting Hydroxyl Radical in Living Cells
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Abbreviation of Common Chemical Compounds Related to Peptides
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Are there any calcium indicators that don't require probenecid (PBC)?