iFluor® 460 Styramide
Ordering information
Price | |
Catalog Number | |
Unit Size | |
Quantity |
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 | 975.27 |
Solvent | DMSO |
Spectral properties
Correction Factor (260 nm) | 0.98 |
Correction Factor (280 nm) | 0.46 |
Extinction coefficient (cm -1 M -1) | 800001 |
Excitation (nm) | 468 |
Emission (nm) | 493 |
Quantum yield | ~0.81 |
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 975.27 | Correction Factor (260 nm) 0.98 | Correction Factor (280 nm) 0.46 | Extinction coefficient (cm -1 M -1) 800001 | Excitation (nm) 468 | Emission (nm) 493 | Quantum yield ~0.81 |
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. In combination with our superior iFluor® dyes that have higher florescence intensity, increased photostability and enhanced water solubility, the iFluor® dye-labeled Styramide™ conjugates can generate fluorescence signal with significantly higher precision and sensitivity (more than 100 times) than standard ICC/IF/IHC. Compared to tyramide reagents, the Styramide™ conjugates have ability to label the target at higher efficiency and thus generate significantly higher fluorescence signal. Styramide™ conjugates also allow significantly less consumption of primary antibody compared to standard directly conjugate method or tyramide amplification with the same level of sensitivity. iFluor® 460 Styramide is a new unique PSA reagent for multicolor application with our existing PSA and TSA reagents. It is optimized with the instruments equipped with the 460 nm laser or other equivalent light sources such as LEDs.
Platform
Fluorescence microscope
Excitation | FITC filter set |
Emission | FITC 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™ 460 Styramide stock solution (100X)
Add 100 µL of DMSO into the vial of iFluor™ 460 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™ 460 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® 460 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 | 102.536 µL | 512.679 µL | 1.025 mL | 5.127 mL | 10.254 mL |
5 mM | 20.507 µL | 102.536 µL | 205.071 µL | 1.025 mL | 2.051 mL |
10 mM | 10.254 µL | 51.268 µL | 102.536 µL | 512.679 µL | 1.025 mL |
Molarity calculator
Enter any two values (mass, volume, concentration) to calculate the third.
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Spectrum
Open in Advanced Spectrum Viewer


Spectral properties
Correction Factor (260 nm) | 0.98 |
Correction Factor (280 nm) | 0.46 |
Extinction coefficient (cm -1 M -1) | 800001 |
Excitation (nm) | 468 |
Emission (nm) | 493 |
Quantum yield | ~0.81 |
Product Family
Images

Figure 1. Fluorescence IHC of formaldehyde-fixed, paraffin-embedded human lung adenocarcinoma positive tissue using PSA ™ amplified methods. Human lung adenocarcinoma positive tissue sections were stained with Mouse anti-EpCAM or Control Mouse IgG antibody and then incubated with polyHRP-labeled Goat anti-Mouse IgG secondary antibody followed by iFluor® 460 Styramide™ (Cat#44902).

Figure 2. 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® 460 styramide (Cat No. 44902). Images were captured on a fluorescence microscope equipped with a FITC 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
An Advanced Method for the Immunohistochemical Detection of Nitric Oxide Synthase (NOS) in the Female Genital Tract.
Authors: Ückert, Stefan and Richter, Karin and Fischer, Klaus-Dieter and Albrecht, Knut and Kuczyk, Markus A
Journal: Analytical biochemistry (2021): 114264
Authors: Ückert, Stefan and Richter, Karin and Fischer, Klaus-Dieter and Albrecht, Knut and Kuczyk, Markus A
Journal: Analytical biochemistry (2021): 114264
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
Neutrophils use selective autophagy receptor Sqstm1/p62 to target Staphylococcus aureus for degradation in vivo in zebrafish.
Authors: Gibson, Josie F and Prajsnar, Tomasz K and Hill, Christopher J and Tooke, Amy K and Serba, Justyna J and Tonge, Rebecca D and Foster, Simon J and Grierson, Andrew J and Ingham, Philip W and Renshaw, Stephen A and Johnston, Simon A
Journal: Autophagy (2021): 1448-1457
Authors: Gibson, Josie F and Prajsnar, Tomasz K and Hill, Christopher J and Tooke, Amy K and Serba, Justyna J and Tonge, Rebecca D and Foster, Simon J and Grierson, Andrew J and Ingham, Philip W and Renshaw, Stephen A and Johnston, Simon A
Journal: Autophagy (2021): 1448-1457
Multiplex Immunohistochemistry Analysis of Melanoma Tumor-Infiltrating Lymphocytes.
Authors: Nguyen, Thu and Kocovski, Nikolce and Macdonald, Sean and Yeang, Han Xian Aw and Wang, Minyu and Neeson, Paul J
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 557-572
Authors: Nguyen, Thu and Kocovski, Nikolce and Macdonald, Sean and Yeang, Han Xian Aw and Wang, Minyu and Neeson, Paul J
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 557-572
The autophagic response to Staphylococcus aureus provides an intracellular niche in neutrophils.
Authors: Prajsnar, Tomasz K and Serba, Justyna J and Dekker, Bernice M and Gibson, Josie F and Masud, Samrah and Fleming, Angeleen and Johnston, Simon A and Renshaw, Stephen A and Meijer, Annemarie H
Journal: Autophagy (2021): 888-902
Authors: Prajsnar, Tomasz K and Serba, Justyna J and Dekker, Bernice M and Gibson, Josie F and Masud, Samrah and Fleming, Angeleen and Johnston, Simon A and Renshaw, Stephen A and Meijer, Annemarie H
Journal: Autophagy (2021): 888-902
Monitoring of Active Notch Signaling in Mouse Bladder Urothelium.
Authors: Karakaidos, Panagiotis and Rampias, Theodoros
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 121-134
Authors: Karakaidos, Panagiotis and Rampias, Theodoros
Journal: Methods in molecular biology (Clifton, N.J.) (2021): 121-134
Tagging the vanA gene in wastewater microbial communities for cell sorting and taxonomy of vanA carrying cells.
Authors: Gallego, Sara and Barkay, Tamar and Fahrenfeld, N L
Journal: The Science of the total environment (2020): 138865
Authors: Gallego, Sara and Barkay, Tamar and Fahrenfeld, N L
Journal: The Science of the total environment (2020): 138865
Estimating the ploidy of Gracilariopsis lemaneiformis at both the cellular and genomic level1.
Authors: Chen, Haihong and Feng, Xiaoqing and Jiang, Minjie and Xiao, Baoheng and Zhang, Jingyu and Zhang, Wei and Hu, Yiyi and Sui, Zhenghong
Journal: Journal of phycology (2020): 1339-1348
Authors: Chen, Haihong and Feng, Xiaoqing and Jiang, Minjie and Xiao, Baoheng and Zhang, Jingyu and Zhang, Wei and Hu, Yiyi and Sui, Zhenghong
Journal: Journal of phycology (2020): 1339-1348
Tyramide Signal-Amplified Immunofluorescence of MYCN and MYC in Human Tissue Specimens and Cell Line Cultures.
Authors: Schafer, Johanna M and Pietenpol, Jennifer A
Journal: Bio-protocol (2020): e3677
Authors: Schafer, Johanna M and Pietenpol, Jennifer A
Journal: Bio-protocol (2020): e3677