TripleStain IHC Kit: M&R&G on human tissue (DAB, AP/Red & HRP/Green) (ab183290)

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Overview

  • Product name

    TripleStain IHC Kit: M&R&G on human tissue (DAB, AP/Red & HRP/Green)

  • Assay type

    Polymer Multiplex

  • Product overview

    Abcam's TripleStain IHC Kit (ab183290) is designed to be used with user-supplied one goat, one mouse and one rabbit primary antibodies to detect three distinct antigens on a single human tissue or cell samples. This kit has been tested on paraffin embedded tissur; however it may also be used on frozen or freshly prepared monolayer cell smears.


    Triple staining uses traditional and non-traditional methods in immunohistostaining to reveal three distinct antigens and their co-expression on a single tissue. Abcam's TripleStain IHC Kit (ab183290) supplies polymer enzyme conjugates: Goat HRP Polymer, Rabbit AP Polymer and Mouse HRP Polymer with three chromogens, DAB (brown); Permanent Red (red); and Emerald (green). Abcam's TripleStain IHC Kit (ab183290) is a non-biotin system, avoiding non-specific binding caused by endogenous biotin. This kit has been optimized to have no cross reaction when detecting three different primary antibodies using our unique blocking system. Simplified steps allow the user to complete triple staining within 3 hours (without antigen retrieval) or 4 hours (with antigen retrieval). The well tested protocol allows the user to permanently mount slides with a coverslip.

  • Tested applications

    Suitable for: IHC-P, IHC-Frmore details

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 24 ml 72 ml 240 ml
    DAB Chromogen (20x) 1 x 1.5ml 1 x 2ml 1 x 6ml
    DAB Substrate 1 x 15ml 2 x 18ml 1 x 120ml
    Emerald Chromogen 1 x 15ml 2 x 18ml 1 x 120ml
    Goat HRP Polymer 1 x 6ml 1 x 18ml 1 x 60ml
    Mouse HRP Polymer 1 x 12ml 2 x 18ml 1 x 120ml
    Non-aqueous Mounting Medium 1 x 12ml 2 x 18ml 2 x 30ml
    Permanent Red Activator (5x) 1 x 3ml 1 x 7.2ml 2 x 12ml
    Permanent Red Chromogen (100x) 1 x 150µl 1 x 360µl 1 x 1.2ml
    Permanent Red Substrate 1 x 15ml 2 x 18ml 1 x 120ml
    Plastic pipette 1 unit 1 unit 1 unit
    Rabbit AP Polymer 1 x 6ml 1 x 18ml 1 x 60ml
    Triple Stain Blocker 1 x 12ml 2 x 18ml 1 x 120ml

  • Research areas

Applications

Our Abpromise guarantee covers the use of ab183290 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-P Use at an assay dependent concentration.
IHC-Fr Use at an assay dependent concentration.

Protocols

References

ab183290 has not yet been referenced specifically in any publications.

Publishing research using ab183290? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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Question

I’ve noticed that, in the protocol from this kit (TripleStain IHC Kit: M&R&G on human tissue (DAB, AP/Red & HRP/Green) (ab183290)), blocking with normal serum is not listed as a step: Tissue Preparation (Up to 70 minutes) 1.Incubate slides in peroxidase blocking reagent (3% H2O2 solution) and alkaline phosphatase blocking reagent (ie levamisole) for 10 minutes. No blocking is needed between step 1 and 2???? 2. Rinse the slides using 2 changes of distilled water. 3. Heat induced Epitope Retrieval (HIER) may be required for primary antibody as suggested by manufacturer. 4. Wash three times with PBS-T or TBS-T for 2 minutes per wash. As far as I know, blocking is always recommended in IHC to prevent unwanted binding of the primary antibodies. I wonder what blocking I should use in this case? Would it be okay to use Carbo-Free Blocking Solution (From Vector laboratory), or would you please advise on whatever that is appropriate?

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Answer

Pre-Blocking step (before primary antibodies are applied) is optional with all our detection kits.  This means that it’s not necessary to use any blocking solution when they use any of our detection kits. 
Carbo-Free Blocking Solution (from Vector Laboratories) is ideal for applications using lectins in which glycoproteins contamination could generate background staining or false positive results.  If needed, I would suggest using donkey serum (5%-10%) as a pre-block with ab183290. Please never use goat serum as pre-block with ab1783290.

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