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Thermo Scientific™ Clean-Blot™ IP Detection Kit (HRP)

Catalog No. PI21232
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PI21232 25 blots
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Catalog No. PI21232 Supplier Thermo Scientific™ Supplier No. 21232
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Description

Avoid interference from denatured IP antibody fragments in Western blots of immunoprecipitated proteins with these HRP and AP conjugates.

Thermo Scientific™ Clean-Blot IP Detection Reagents are HRP and AP conjugates that are optimized for post-immunoprecipitation Western blot detection of primary antibodies without interference from denatured IP antibody fragments.

Clean-Blot IP Reagents allow trouble-free Western blot detection of target proteins following IP assays (immunoprecipitation). They function by specifically binding to functional primary antibodies (whole IgG) without also binding to fragments of the IP antibodies, which usually accompany the immunoprecipitated protein through electrophoresis and membrane transfer. The Clean-Blot IP Reagents and HRP Kit eliminate detection-interference from both heavy-chain (approx. 50kDa) and light-chain (25kDa) IgG-fragments of antibodies used for the initial immunoprecipitation assay. Clean-Blot IP products include detection reagents with alkaline phosphatase conjugate or horseradish peroxidase conjugate and the complete Clean-Blot IP Detection Kit (HRP) for chemiluminescent substrate development.

Highlights:

  • Universal – bind and detect most species, IgG subclasses and isotypes of primary antibodies that are commonly used for Western blotting

  • Compatible – effective with IP assays performed using Protein A, Protein G, or anti-IgG agarose beads and any blocking buffer

  • Cost-effective – eliminates the cost and extra work associated with covalently immobilizing IP antibodies as a means of overcoming Western blot interference

  • Flexible – available in alkaline phosphate (AP) and horseradish peroxidase (HRP) reagents for detection with chemiluminescent, fluorescent or colorimetric substrates

  • Easy to use – no need to change the Western blotting protocol; simply replace conventional secondary HRP conjugate with the Clean-Blot IP Detection Reagent

  • Unobstructed detection – clear Western blot results for immmunoprecipitation assays without significant interference from denatured IgG bands

Specifications

Description Clean-Blot IP Detection Kit (HRP)
Product Type IP Detection Reagent (HRP)
Detection Method Chemiluminescence
Form Liquid
For Use With (Application) Western Blot
Product Line Clean-Blot
Quantity 25 blots
Conjugate HRP
Reactivity All species
Sufficient For 25 Mini Blots, 2000 sq. cm Membrane
Content And Storage Sufficient For: 25 mini blots or 2000 cm2 membrane
• Clean-Blot Detection Reagent (HRP), 2.5 mL
• StartingBlock T20 (TBS) Blocking Buffer, 1 L
• Pierce ECL Detection Reagent 1, Peroxide Solution, 125 mL
• Pierce ECL Detection Reagent 2, Luminol Enhancer Solution, 125 mL

Store at 4°C.
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Frequently Asked Questions (FAQs)

Can I strip and reprobe my blot after using the Clean-Blot IP Detection reagent?

Yes. When using a chemiluminescent substrate, membranes can be stripped and reprobed similar to probing with secondary antibodies.
What substrates are compatible with the Clean-Blot IP Detection reagent?

For best results, use the the Clean-Blot IP Detection with SuperSignal Substrates or Thermo Scientific ECL Western Blotting Substrate. When using other HRP or AP substrates, empirically determine the optimal concentration of the Clean-Blot IP Detection Reagent.
What blocking buffers are compatible with the Clean-Blot IP Detection reagent?

The Clean-Blot IP Detection is compatible with bovine serum albumin, SuperBlock and StartingBlock Blocking Buffers and 5% nonfat milk. Verify compatibility with other blocking buffers by dot-blot analysis.
Does the Clean-Blot IP Detection Reagent bind to all antibody classes and subclasses?

The reagent binds to IgG from a wide range of species: bovine IgG2, goat IgG2, human IgG1, IgG2, IgG4, mouse IgG2a, IgG2b, IgG3, rat IgG2c, sheep IgG2, pig, dog and cat. It will not bind to rat IgG2a and IgG2b or mouse IgG1. If in doubt whether this detection reagent will bind to a specific antibody, perform a dot-blot analysis before the experiment.
How does the Clean-Blot IP Detection Kit work?

This assay uses a HRP conjugate or AP conjugate that binds to native IgG from various host species, allowing clear, specific Western blot detection from IP experiments without interference from denatured IgG. It is used simply as a substitute for secondary antibodies.

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