Overview

  • Product name
    10X Blocking Buffer
  • Tested applications
    Suitable for: ICC, WB, ELISA, In-Cell ELISA, ICC/IFmore details
    Unsuitable for: Flow Cyt
  • General notes

    Protein blocking buffer (pH 7.5) (ab126587) for serum-free blocking of non-specific antibody binding in IHC, ICC, ELISA and western blot. BSA-free

    Preparation: Dilute to 1X in deionized water or PBS (e.g. 1mL 10X blocking buffer + 9mL water/PBS). Diluting in PBS instead of water makes a somewhat more stringent blocking buffer due to the presence of salt in the PBS. Blocking Buffer is pH 7.5. The user should determine empirically whether dilution in water or PBS is most appropriate for the assay at hand. Use diluted block solution promptly. The diluted block solution can be stored at 4C for short periods of time.
    Shelf life for the 10X stock is 1 year.

Properties

Applications

Our Abpromise guarantee covers the use of ab126587 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
ICC Use at an assay dependent concentration.
WB Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
In-Cell ELISA Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
  • Application notes
    Is unsuitable for Flow Cyt.
  • Images

    • Primary :

      All Lanes : top membranes with Anti AMPK pThr172 + Thr183 (ab133448) at 1/1000. Bottom membranes with Anti AMPKa (ab110036) at 1/1000

      Lane 1 = AMPKa1 protein

      Lane 2 = C2C12 cells treated with AICAR

      Lane 3 = untreated C2C12 cells

      Secondary: Goat polyclonal to Rabbit IgG – H&L – Pre-Adsorbed (HRP) and Goat polyclonal to Mouse IgG – H&L – Pre-Adsorbed (HRP) at 1:10,000 developed using the ECL technique.

      Performed under reducing conditions (50mM DTT – Sample heated at 60ºC). Predicted band size for AMPKa: 64 kDa Observed band size for AMPKa: 64 kDa

      Left membrane All blocking steps = 1X blocking reagent ab126587 diluted in PBST

      Right membrane All blocking steps = 5% Milk in PBST Exposure time : 5 minutes



      Primary :

      All Lanes : top membranes with Anti AMPK pThr172 + Thr183 (ab133448) at 1/1000. Bottom membranes with Anti AMPKa (ab110036) at 1/1000

      Lane 1 = AMPKa1 protein

      Lane 2 = C2C12 cells treated with AICAR

      Lane 3 = untreated C2C12 cells

      Secondary: Goat polyclonal to Rabbit IgG – H&L – Pre-Adsorbed (HRP) and Goat polyclonal to Mouse IgG – H&L – Pre-Adsorbed (HRP) at 1:10,000 developed using the ECL technique.

      Performed under reducing conditions (50mM DTT – Sample heated at 60ºC). Predicted band size for AMPKa: 64 kDa Observed band size for AMPKa: 64 kDa

      Left membrane All blocking steps = 1X blocking reagent ab126587 diluted in PBST

      Right membrane All blocking steps = 5% Milk in PBST Exposure time : 5 minutes

    References

    This product has been referenced in:
    • Spence EF  et al. In vivo proximity proteomics of nascent synapses reveals a novel regulator of cytoskeleton-mediated synaptic maturation. Nat Commun 10:386 (2019). Read more (PubMed: 30674877) »
    • Li Y  et al. Autocrine motility factor promotes endometrial cancer progression by targeting GPER-1. Cell Commun Signal 17:22 (2019). Read more (PubMed: 30836961) »
    See all 5 Publications for this product

    Customer reviews and Q&As

    1-4 of 4 Abreviews or Q&A

    Question
    Answer

    The the lab has confirmed this blocking solution is casein based with no BSA, so it may be an option for your ELISA if this is a concern for you.

    Please let me know if you have any further questions!

    Read More

    Question
    Answer

    Thank you for contacting us.



    Yes – in our experience, as with PBS, the extra salt in TBS will make a more stringent block/ prevent non specific binding.





    I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

    Read More

    Answer


    1. ab126420, the components of the blocking buffer are proprietary, and the solution is a standard blocking solution.

    2. ab111542: The blocking solution in the kit is the same as the 10X blocking solution sold separately (ab126587). It is casein based and the % of casein is proprietary.

    Read More

    Answer

    Thank you for contacting us.

    In this case a casein block was used as BSA and serums would cause high background. We do carry a casein based blocking solution, ab126587,which we have used extensively and feel is a superior product in ICC. More information about ab126587 may be found at the following link:

    https://www.abcam.com/10x-blocking-buffer-ab126587.html

    I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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