Invitrogen™ Alexa Fluor™ 488 Tyramide SuperBoost™ Kit, goat anti-rabbit IgG

Product Code.	Quantity	unitSize
15661892	Sufficient for 150 Slides	Pack of 150
15631902	50 Slides	Pack of 50

Product Code. 15661892 Supplier Invitrogen™ Supplier No. B40922

Description

Features poly-HRP-conjugated goat anti-rabbit IgG secondary antibody. Alexa Fluor™ 488 Tyramide SuperBoost™ Kit combine brightness of AlexaFluor dyes with high signal amplification of a poly-HRP-mediated tyramide labeling reaction to produce s

SuperBoost™ tyramide signal amplification is most sensitive method for detection of low abundant targets in multiplexable fluorescent immunocytochemistry (ICC), immunohistochemistry (IHC ), and in situ hybridization (ISH).

Features:

• High sensitivity for detection of low-level or hard-to-detect targets by fluorescent imaging
• Simple protocol and detection using standard filters
• Suitable for high-resolution multiplex images, co-label with DAPI, secondary antibodies, and other SuperBoost kits
• Requires 10-100 times less primary antibody then standard ICC/IHC/ISH experiments

Benefits:

• Enhancement of signal using Alexa Fluor tyramides: Utilize Alexa Fluor tyramides, which react with HRP to ultimately deposit bright and photostable Alexa Fluor dye on surrounding proteins and other similar molecules
• Poly-HRP enhancement: Several HRP enzymes are conjugated with short polymers, enhancing signal by several fold over regular HRP systems
• Reaction stop solution: Include an HRP stop solution to halt the HRP reaction
• Reduction of background: Include blockers for elimination or reduction of endogenous peroxidase and fluorescent background signals

Shelf Life:

• Stable for 6 months after receiving

Specifications

• Content And Storage: 1 kit sufficient for 150 microscope slides (18 x 18 mm), containing: Blocking buffer (1X), 22.5 mL
  Poly-HRP-conjugated goat anti-rabbit secondary antibody (1X), 22.5 mL
  Alexa Fluor tyramide reagent
  Hydrogen pero
• Quantity: Sufficient for 150 Slides
• Shipping Condition: Approved for shipment at Room Temperature or on Wet Ice
• Conjugate: Alexa Fluor 488
• Product Line: SuperBoost
• Product Type: Tyramide SuperBoost Kit

Frequently Asked Questions (FAQs)

With a SuperBoost tyramide kit, I got excessive and non-specific labeling. What can I do to limit background and acquire a more localized labeling?

To limit background, we recommend performing a pre-blocking step with 3% H2O2 for 60 mins to inactivate endogenous peroxidases. To limit the localization of labeling, we recommend optimizing the final concentration of the primary and secondary antibodies and the dye-tyramide. You may also limit the incubation time of the dye-tyramide on the sample.

Is it possible to perform dual TSA labeling with SuperBoost tyramide kits?

Yes. This involves the sequential application of the antibodies and the tyramides with a HRP-quenching step between antibodies using H2O2.

How are SuperBoost tyramide kits different from the original TSA labeling kits?

The SuperBoost tyramide kits utilize poly-HRP labeled antibodies. This provides a greater number of horseradish peroxidase (HRP) molecules per antibody. The original kits used antibodies and streptavidin that were directly conjugated with HRP and thus, limited the number per antibody or streptavidin.

I used a neuron-specific antibody to label my neurons. I can't get enough signal from my fluorescent dye conjugated primary antibody. What can I do to improve it?

Here are our recommendations:

Use one of our extensive selection of secondary antibodies conjugated to bright, photostable Alexa Fluor dyes. The degree of labeling for each conjugate is 2-8 fluorophores per IgG molecule, with potentially three secondary antibody-binding sites per primary antibody, providing signal amplification of approximately 10-20 fluorophores per primary antibody.
Alternatively, primary antibody labeling can be detected with a biotinylated secondary antibody in conjunction with either a fluorescent streptavidin or a streptavidin bridge followed by a biotinylated reporter such as Qdot biotin. Although processing times increase with additional incubation and endogenous biotin-blocking steps, detection sensitivity also improves as a result of the labeled streptavidin.
For low-abundance targets, signal amplification may be necessary for optimal signal-to-noise ratios. Tyramide signal amplification (TSA) is an enzyme-mediated detection method that utilizes the catalytic activity of horseradish peroxidase (HRP) to generate reactive fluorophore-labeled tyramide radicals. These short-lived tyramide radicals covalently couple to nearby residues, producing an amplified fluorescent signal localized at the HRP-target interaction site.
For improved detection sensitivity with rapidly bleaching dyes, our SlowFade Diamond or ProLong Diamond antifade reagents have been shown to increase photostability and reduce initial fluorescence quenching in fixed cells, fixed tissues, and cell-free preparations.
Please review this web page for further optimization tips (https://www.thermofisher.com/us/en/home/references/newsletters-and-journals/bioprobes-journal-of-cell-biology-applications/bioprobes-issues-2011/bioprobes-66-october-2011/guide-to-immunocytochemistry.html).

I have a very low-abundance antigen. How can I amplify my signal?

A common method for amplifying antibody detection is biotin-streptavidin detection, where a biotinylated secondary antibody is combined with subsequent labeling with a dye-conjugated streptavidin. This will amplify the signal by approximately 2-8 times, but endogenous biotin must be blocked beforehand. Another option is to use tyramide-signal amplification, where a horseradish peroxidase conjugate is used with a dye-labeled tyramide. This will amplify the signal by approximately 10-20 times, but endogenous peroxidase will need to be blocked. A final option may be to use a Qdot nanoparticle antibody or streptavidin conjugate, which can yield a signal as much as 40 times higher than a standard organic dye conjugate, depending on the Qdot color.

For Research Use Only. Not for use in diagnostic procedures.