Actively helping customers, employees and the global community during the coronavirus SARS-CoV-2 outbreak.  Learn more >>

XFD488 tyramide reagent *Same Structure to Alexa Fluor™ 488 tyramide*

Ordering information
Price ()
Catalog Number11070
Unit Size
Find Distributor
Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Physical properties
Molecular weight856.02
SolventDMSO
Spectral properties
Correction Factor (260 nm)0.3
Correction Factor (280 nm)0.11
Extinction coefficient (cm -1 M -1)73000
Excitation (nm)499
Emission (nm)520
Quantum yield0.921
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageFreeze (< -15 °C); Minimize light exposure
UNSPSC12171501
Related products
XFD488 azide *Same Structure to Alexa Fluor™ 488 azide*
XFD488 alkyne *Same Structure to Alexa Fluor™ 488 alkyne*
XFD488 NHS Ester *Same Structure to Alexa Fluor™ 488 NHS Ester*
XFD488 C5 Maleimide *Same Structure to Alexa Fluor™ 488 C5 Maleimide*
XFD350 NHS Ester *Same Structure to Alexa Fluor™ 350 NHS Ester*
XFD532 NHS Ester *Same Structure to Alexa Fluor™ 532 NHS Ester*
XFD594 NHS Ester *Same Structure to Alexa Fluor™ 594 NHS Ester*
XFD350 C5 Maleimide *Same Structure to Alexa Fluor™ 350 C5 Maleimide*
XFD532 C5 Maleimide *Same Structure to Alexa Fluor™ 532 C5 Maleimide*
XFD594 C5 Maleimide *Same Structure to Alexa Fluor™ 594 C5 Maleimide*
XFD488 Hydroxylamine *Same Structure to Alexa Fluor™ 488 Hydroxylamine*
XFD350 goat anti-mouse IgG (H+L) *Cross Adsorbed, XFD350 Same Structure to Alexa Fluor™ 350*
XFD488 goat anti-mouse IgG (H+L) *Cross Adsorbed, XFD488 Same Structure to Alexa Fluor™ 488*
XFD594 goat anti-mouse IgG (H+L) *Cross Adsorbed, XFD594 Same Structure to Alexa Fluor™ 594*
XFD350 goat anti-rabbit IgG (H+L) *Cross Adsorbed, XFD350 Same Structure to Alexa Fluor™ 350*
XFD488 goat anti-rabbit IgG (H+L) *Cross Adsorbed, XFD488 Same Structure to Alexa Fluor™ 488*
XFD594 goat anti-rabbit IgG (H+L) *Cross Adsorbed, XFD594 Same Structure to Alexa Fluor™ 594*
XFD488 amine *Same Structure to Alexa Fluor™ 488 amine*
XFD594 amine *Same Structure to Alexa Fluor™ 594 amine*
XFD350-streptavidin conjugate *XFD350 Same Structure to Alexa Fluor™ 350*
XFD488-streptavidin conjugate *XFD488 Same Structure to Alexa Fluor™ 488*
XFD594-streptavidin conjugate *XFD594 Same Structure to Alexa Fluor™ 594*
XFD350 Phalloidin *XFD350 Same Structure to Alexa Fluor™ 350*
XFD488 Phalloidin *XFD488 Same Structure to Alexa Fluor™ 488*
XFD594 Phalloidin *XFD594 Same Structure to Alexa Fluor™ 594*
XFD532 acid *Same Structure to Alexa Fluor™ 532 acid*
XFD488 acid *Same Structure to Alexa Fluor™ 488 acid*
XFD488 NHS Ester-UltraPure Grade *XFD488 Same Structure to Alexa Fluor™ 488*
XFD555 NHS Ester *Same Structure to Alexa Fluor™ 555 NHS Ester*
XFD647 NHS Ester *Same Structure to Alexa Fluor™ 647 NHS Ester*
XFD680 NHS Ester *Same Structure to Alexa Fluor™ 680 NHS Ester*
XFD700 NHS Ester *Same Structure to Alexa Fluor™ 700 NHS Ester*
XFD750 NHS Ester *Same Structure to Alexa Fluor™ 750 NHS Ester*
XFD647 maleimide *C2, XFD647 Same Structure to Alexa Fluor™ 647*
XFD546 NHS Ester *Same Structure to Alexa Fluor™ 546 NHS Ester*
XFD568 NHS Ester *Same Structure to Alexa Fluor™ 568 NHS Ester*
XFD350 acid *Same Structure to Alexa Fluor™ 350 acid*
XFD546 acid *Same Structure to Alexa Fluor™ 546 acid*
XFD568 acid *Same Structure to Alexa Fluor™ 568 acid*
XFD488 tetrazine *Same Structure to Alexa Fluor™ 488 tetrazine*
XFD488 aldehyde *Same Structure to Alexa Fluor™ 488 aldehyde*
XFD™488-dUTP *1 mM in Tris Buffer (pH 7.5)*
XFD514 acid
XFD514 NHS Ester *Same Structure to Alexa Fluor™ 514 NHS Ester*
XFD514 tyramide
XFD532 tyramide
XFD647 Azide
XFD647 Alkyne
XFD488 TCO
Show More (49)

OverviewpdfSDSpdfProtocol


Molecular weight
856.02
Correction Factor (260 nm)
0.3
Correction Factor (280 nm)
0.11
Extinction coefficient (cm -1 M -1)
73000
Excitation (nm)
499
Emission (nm)
520
Quantum yield
0.921
XFD488 is manufactured by AAT Bioquest, and it has the same chemical structure of Alexa Fluor® 488 (Alexa Fluor® is the trademark of ThermoFisher). For many immunohistochemical (IHC) applications, the traditional enzymatic amplification procedures are sufficient for achieving adequate antigen detection. However, several factors limit the sensitivity and utility of these procedures. Tyramide signal amplification (TSA) has proven to be a particularly versatile and powerful enzyme amplification technique with improved assay sensitivity. TSA is based on the ability of HRP, in the presence of low concentrations of hydrogen peroxide, to convert labeled tyramine-containing substrate into an oxidized, highly reactive free radical that can covalently bind to tyrosine residues at or near the HRP. To achieve maximal IHC detection, tyramine is prelabeled with a fluorophore. The signal amplification conferred by the turnover of multiple tyramide substrates per peroxidase label translates ultrasensitive detection of low-abundance targets and the use of smaller amounts of antibodies and hybridization probes. In immunohistochemical applications, sensitivity enhancements derived from TSA method allow primary antibody dilutions to be increased to reduce nonspecific background signals, and can overcome weak immunolabeling caused by suboptimal fixation procedures or low levels of target expression. XFD488 tyramide contains the bright XFD488 dye that can be readily detected with the standard FITC filter set.

Platform


Fluorescence microscope

ExcitationFITC filter set
EmissionFITC filter set
Recommended plateBlack wall/clear bottom
Instrument specification(s)FITC filter set

Example protocol


AT A GLANCE

Protocol Summary
  1. Fix/permeabilize/block cells or tissue
  2. Add primary antibody in blocking buffer
  3. Add HRP-conjugated secondary antibody
  4. Prepare tyramide 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

Tyramide stock solution (200X)

Add 100 µL DMSO to vial and mix well. Note: Unused Tyramide stock solution can be stored at 2-8 °C.

PREPARATION OF WORKING SOLUTION

Tyramide working solution (1X)

Add 100 µL of Tyramide stock solution into 20 mL of buffer of your choice containing 0.003% H2O2. Note: Tris Buffer, pH=7.4 can be used for similar performance. Note: Tyramide working solution should be used immediately and made fresh on the day of use. Note: 20 mL solution is good for 200 tests.

SAMPLE EXPERIMENTAL PROTOCOL

This protocol is applicable for both cells and tissues staining.

Cell fixation and permeabilization
  1. Fix the cells or tissue with 3.7% formaldehyde or paraformaldehyde, in PBS at room temperature for 20 minutes.
  2. Rinse the cells or tissue with PBS twice.
  3. Permeabilize the cells with 0.1% Triton X-100 solution for 1-5 minutes at room temperature.
  4. 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-tissueimmunohistochemistry-protocol.html

Peroxidase labeling
  1. 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.
  2. Optional: If using HRP-conjugated streptavidin, it is advisable to block endogenous biotins by biotin blocking buffer.
  3. Block with preferred blocking solution (such as PBS with 1% BSA) for 30 minutes at 4 °C.
  4. Remove blocking solution and add primary antibody diluted in recommended antibody diluent for 60 minutes at room temperature or overnight at 4 °C.
  5. Wash with PBS three times for 5 minutes each.
  6. 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.
  7. Wash with PBS three times for 5 minutes each.
Tyramide labeling
  1. Prepare and apply 100 µL of Tyramide 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 Tyramide. You should optimize the incubation period using positive and negative control samples at various incubation time points. Or you can use lower concentration of Tyramide in the working solution.
  2. Rinse with PBS three times.
Counterstain and fluorescence imaging
  1. 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.
  2. Mount the coverslip using a mounting medium with anti-fading properties.
  3. Use the appropriate filter set to visualize the signal from the Tyramide labeling.

Table 1. Products recommended for nucleus counterstain

Cat#Product NameEx/Em (nm)
17548Nuclear Blue™ DCS1350/461
17550Nuclear Green™ DCS1503/526
17551Nuclear Orange™ DCS1528/576
17552Nuclear Red™ DCS1642/660

Calculators


Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of XFD488 tyramide reagent *Same Structure to Alexa Fluor™ 488 tyramide* to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

0.1 mg0.5 mg1 mg5 mg10 mg
1 mM116.82 µL584.099 µL1.168 mL5.841 mL11.682 mL
5 mM23.364 µL116.82 µL233.639 µL1.168 mL2.336 mL
10 mM11.682 µL58.41 µL116.82 µL584.099 µL1.168 mL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)Molecular weightVolume (Calculate)Concentration (Calculate)Moles
/=x=

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Correction Factor (260 nm)0.3
Correction Factor (280 nm)0.11
Extinction coefficient (cm -1 M -1)73000
Excitation (nm)499
Emission (nm)520
Quantum yield0.921

Product family


NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)Quantum yieldCorrection Factor (260 nm)Correction Factor (280 nm)
XFD546 tyramide reagent *Same Structure to Alexa Fluor™ 546 tyramide*5615721120000.7910.210.12
XFD594 tyramide reagent *Same Structure to Alexa Fluor™ 594 tyramide*590618920000.6610.430.56
XFD350 tyramide reagent *Same Structure to Alexa Fluor™ 350 tyramide*34344119000-0.250.19
XFD568 tyramide reagent *Same Structure to Alexa Fluor™ 568 tyramide*579603880000.6910.450.46

Citations


View all 17 citations: Citation Explorer
Stage-specific requirement for METTL3-dependent m6A modification during dental pulp stem cell differentiation
Authors: Luo, Haiyun and Liu, Wenjing and Zhou, Yachuan and Zhang, Yanli and Wu, Junrong and Wang, Ruolan and Shao, Longquan
Journal: Journal of Translational Medicine (2022): 1--15
The midbody and midbody remnant are assembly sites for RNA and active translation
Authors: Skop, Ahna and Park, Sungjin and Dahn, Randall and Kurt, Elif and Presle, Adrien and VandenHeuvel, Kathryn and Moravec, Cara and Jambhekar, Ashwini and Olukoga, Olushola and Shepherd, Jason and others,
Journal: (2022)
Cloning, pattern of gonadal soma-derived factor mRNA in the orange-spotted grouper, Epinephelus coioides
Authors: Huang, Jingjun and Wei, Qianhao and Zhao, Mi and Zhou, Libin and Shi, Herong and Zhang, Yong and Chen, Huapu
Journal: Aquaculture Reports (2021): 100754
An ultrasensitive electrochemical immunosensor for procalcitonin detection based on the gold nanoparticles-enhanced tyramide signal amplification strategy
Authors: Liu, P., Li, C., Zhang, R., Tang, Q., Wei, J., Lu, Y., Shen, P.
Journal: Biosens Bioelectron (2019): 543-550
A amperometric immunosensor for sensitive detection of circulating tumor cells using a tyramide signal amplification-based signal enhancement system
Authors: Zhou, X., Li, Y., Wu, H., Huang, W., Ju, H., Ding, S.
Journal: Biosens Bioelectron (2019): 88-94
Gold nanoparticle labeling with tyramide signal amplification for highly sensitive detection of alpha fetoprotein in human serum by ICP-MS
Authors: Li, X., Chen, B., He, M., Xiao, G., Hu, B.
Journal: Talanta (2018): 40-46
Selective Proteomic Proximity Labeling Assay Using Tyramide (SPPLAT): A Quantitative Method for the Proteomic Analysis of Localized Membrane-Bound Protein Clusters
Authors: Rees, J. S., Li, X. W., Perrett, S., Lilley, K. S., Jackson, A. P.
Journal: Curr Protoc Protein Sci (2017): 19 27 1-19 27 18
High Resolution Fluorescent In Situ Hybridization in Drosophila Embryos and Tissues Using Tyramide Signal Amplification
Authors: J, undefined and ura, A., Hu, J., Wilk, R., Krause, H. M.
Journal: J Vis Exp (2017): se name="11070.enl" path="C:\Website\Referenc
Droplet-Free Digital Enzyme-Linked Immunosorbent Assay Based on a Tyramide Signal Amplification System
Authors: Akama, K., Shirai, K., Suzuki, S.
Journal: Anal Chem (2016): 7123-9
Selective Proteomic Proximity Labeling Assay Using Tyramide (SPPLAT): A Quantitative Method for the Proteomic Analysis of Localized Membrane-Bound Protein Clusters
Authors: Rees, J. S., Li, X. W., Perrett, S., Lilley, K. S., Jackson, A. P.
Journal: Curr Protoc Protein Sci (2015): 19 27 1-18

References


View all 74 references: Citation Explorer
Tyramide Signal Amplification for Immunofluorescent Enhancement
Authors: Faget L, Hnasko TS.
Journal: Methods Mol Biol (2015): 161
Enhanced detection of Porcine reproductive and respiratory syndrome virus in fixed tissues by in situ hybridization following tyramide signal amplification
Authors: Trang NT, Hirai T, Ngan PH, Lan NT, Fuke N, Toyama K, Yamamoto T, Yamaguchi R.
Journal: J Vet Diagn Invest (2015): 326
Rapid and sensitive detection of Escherichia coli O157:H7 in milk and ground beef using magnetic bead-based immunoassay coupled with tyramide signal amplification
Authors: Aydin M, Herzig GP, Jeong KC, Dunigan S, Shah P, Ahn S.
Journal: J Food Prot (2014): 100
Multiplexed immunohistochemistry, imaging, and quantitation: a review, with an assessment of Tyramide signal amplification, multispectral imaging and multiplex analysis
Authors: Stack EC, Wang C, Roman KA, Hoyt CC.
Journal: Methods (2014): 46
KSHV cell attachment sites revealed by ultra sensitive tyramide signal amplification (TSA) localize to membrane microdomains that are up-regulated on mitotic cells
Authors: Garrigues HJ, Rubinchikova YE, Rose TM.
Journal: Virology (2014): 75
Sensitive whole-mount fluorescent in situ hybridization in zebrafish using enhanced tyramide signal amplification
Authors: Lauter G, Soll I, Hauptmann G.
Journal: Methods Mol Biol (2014): 175
Characterization of GABAergic neurons in the mouse lateral septum: a double fluorescence in situ hybridization and immunohistochemical study using tyramide signal amplification
Authors: Zhao C, Eisinger B, Gammie SC.
Journal: PLoS One (2013): e73750
Quantification of alpha-tubulin isotypes by sandwich ELISA with signal amplification through biotinyl-tyramide or immuno-PCR
Authors: Draberova E, Stegurova L, Sulimenko V, Hajkova Z, Draber P.
Journal: J Immunol Methods (2013): 63
Pitfalls using tyramide signal amplification (TSA) in the mouse gastrointestinal tract: endogenous streptavidin-binding sites lead to false positive staining
Authors: Horling L, Neuhuber WL, Raab M.
Journal: J Neurosci Methods (2012): 124
Integrated tyramide and polymerization-assisted signal amplification for a highly-sensitive immunoassay
Authors: Yuan L, Xu L, Liu S.
Journal: Anal Chem (2012): 10737