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Stayright™ Purple HRP Staining Kit

Immunohistochemical detection of EpCAM in FFPE lung adenocarcinoma tissue. The tissue sections were incubated with poly-HRP conjugated Goat anti-Rabbit IgG and then developed with Stayright™ Purple (Left) or DAB (Right), respectively. Cells were also counterstained with hematoxylin. Stayright™ Purple generates an intense stain with high sensitivity and clear resolution similar as DAB.
Immunohistochemical detection of EpCAM in FFPE lung adenocarcinoma tissue. The tissue sections were incubated with poly-HRP conjugated Goat anti-Rabbit IgG and then developed with Stayright™ Purple (Left) or DAB (Right), respectively. Cells were also counterstained with hematoxylin. Stayright™ Purple generates an intense stain with high sensitivity and clear resolution similar as DAB.
Immunohistochemical detection of EpCAM in FFPE lung adenocarcinoma tissue. The tissue sections were incubated with poly-HRP conjugated Goat anti-Rabbit IgG and then developed with Stayright™ Purple (Left) or DAB (Right), respectively. Cells were also counterstained with hematoxylin. Stayright™ Purple generates an intense stain with high sensitivity and clear resolution similar as DAB.
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Telephone1-800-990-8053
Fax1-800-609-2943
Emailsales@aatbio.com
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Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12352200

OverviewpdfSDSpdfProtocol


3,3'-Diaminobenzidine (DAB) has been applied for decades as the most commonly used IHC chromogen because it is inexpensive and sensitive for routine applications. However, DAB has been shown to be mutagenic and hazardous to laboratory workers and the environment. In order to address this issue, AAT provides Stayright™ Purple as a significantly safer IHC chromogen than DAB. Furthermore, Stayright™ Purple provides a rapid and simple method to develop clean and intense purple stain in the presence of HRP with high sensitivity as DAB. The Stayright™ Purple HRP substrate also shows non-mutagenic effects with minimal cytotoxicity. ReadiUse™ Stayright™ Purple Peroxidase (HRP) Substrate is suitable for use in peroxidase (HRP)-based immunohistochemistry (IHC) and in situ hybridization (ISH) staining methods. Upon HRP-induced oxidation, Stayright™ Purple forms a purple insoluble precipitating product at the target site of your assay. The purple end product is insoluble in organic solvents and organic mounting media, thus the distinct purple stain can maintain through regular dehydration and coverslipping steps. For enhanced convenience, you try our ReadiUse™ Stayright™ Purple Peroxidase (HRP) Substrate (#45900 and 45901). It is a stable pre-mixed solution containing hydrogen peroxide so all mixing steps are eliminated and is ready to use.

Platform


Light microscope

Instrument specification(s)White light

Components


Example protocol


AT A GLANCE

Protocol Summary

  1. Apply working Stayright™ Purple solution to tissue section.
  2. Incubate tissue section for 5-15 minutes.
  3. Rinse tissue for 5-10 minutes, counterstain.
  4. Add mounting medium to cover the section.

PREPARATION OF WORKING SOLUTION

Add 10 μL of 100X Stayright™ Purple (Component A) and 1 μL of stabilized 3% Hydrogen peroxide (Component C) in every 1 mL of Stayright™ Purple HRP buffer (Component B).

Note  Unused pre-mixed working solution can be stored for few weeks in 2-4 oC for future application. However, we recommend mixing all these components as needed. 

SAMPLE EXPERIMENTAL PROTOCOL

  1. Cover section with the working Stayright™ Purple solution. Incubate at room temperature for 5-15 minutes.

  2. Immerse slides in dH2O to stop the color development and monitor the staining intensity. If the staining intensity is not bright enough, longer incubation is needed. One can re-apply the Stayright™ Purple solution to continue the development.

  3. Wash with dH2O for 5-10 minutes.

  4. Use a desired counterstain if needed.

  5. Dehydrate with ethanol and permanently mount in organic permanent mounting medium.

Note   For guidelines on sample preparation, please visit   https://www.aatbio.com/resources/guides/paraffin-embedded-tissue-immunohistochemistry-protocol.html

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Citations


View all 25 citations: Citation Explorer
3,3'-diaminobenzidine (DAB)-H2O2-HRP voltammetric enzyme-linked immunoassay for the detection of carcionembryonic antigen
Authors: Zhang, S., Yang, J., Lin, J.
Journal: Bioelectrochemistry (2008): 47-52
HistoGreen: a new alternative to 3,3'-diaminobenzidine-tetrahydrochloride-dihydrate (DAB) as a peroxidase substrate in immunohistochemistry?
Authors: Thomas, M. A., Lemmer, B.
Journal: Brain Res Brain Res Protoc (2005): 107-18
Stability and solubility of 3,3'-diaminobenzidine (DAB)
Authors: Kiernan, J. A.
Journal: Biotech Histochem (2003): 135
Safe diaminobenzidine (DAB) disposal
Authors: Horn, H.
Journal: Biotech Histochem (2002): 229
Non-fluorescent chromosome painting using the peroxidase/diaminobenzidine (DAB) reaction
Authors: K, undefined and a, R., Suzuki, M., Minamihisamatsu, M., Furukawa, A., Odaka, T., Hayata, I.
Journal: Int J Radiat Biol (1998): 529-33
Modified cerium-based and Gomori-based cerium methods for light microscopic phosphatase histochemistry: the cerium-perhydroxide-diaminobenzidine-nickel (Ce-H2O2-DAB-Ni and Ce/Ce-H2O2-DAB-Ni) two-step procedures
Authors: Halbhuber, K. J., Feuerstein, H., Moller, U., Klinger, M.
Journal: Acta Histochem (1992): 87-103
Employment of merocyanine 540 fluorescence to form diaminobenzidine (DAB) oxidation product: a photoconversion method for the visualization of erythrocyte membrane fluidity for light and electron microscopy
Authors: Oehring, H., Halbhuber, K. J.
Journal: Acta Histochem (1991): 127-34
The cerium perhydroxide-diaminobenzidine (Ce-H2O2-DAB) procedure. New methods for light microscopic phosphatase histochemistry and immunohistochemistry
Authors: Halbhuber, K. J., Gossrau, R., Moller, U., Hulstaert, C. E., Zimmermann, N., Feuerstein, H.
Journal: Histochemistry (1988): 289-97
Copper-H2O2 oxidation strikingly improves silver intensification of the nickel-diaminobenzidine (Ni-DAB) end-product of the peroxidase reaction
Authors: Gallyas, F., Merchenthaler, I.
Journal: J Histochem Cytochem (1988): 807-10
The use of gold-substituted silver-intensified diaminobenzidine (DAB) and non-intensified DAB for simultaneous electron microscopic immunoperoxidase labeling of tyrosine hydroxylase and glutamic acid decarboxylase immunoreactivity in the rat medial preopti
Authors: Gorcs, T. J., Leranth, C., MacLusky, N. J.
Journal: J Histochem Cytochem (1986): 1439-47