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Anti-mouse IgG VeriBlot for IP secondary antibody (HRP) (ab131368)

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Overview

  • Product nameAnti-mouse IgG VeriBlot for IP secondary antibody (HRP)See all IgG secondary antibodies ...
  • Target speciesMouse
  • SpecificityVeriBlot for IP secondary antibodies are anti-mouse immunoblotting reagents that enable the trouble-free detection of immunoblotted target protein bands, without interference from denatured IgG. This allows to detect the (co-)immunoprecipitated protein without masking by the IgG heavy (50 kDa) and light chains (25 kDa). In general this interference tends to originate from secondary antibodies which recognize primary antibodies released with the antigen during the immunoprecipitation procedure or endogenous IgGs from the lysate itself. VeriBlot for IP secondary antibodies only recognize native (non-reduced) antibodies and therefore the detection of heavy and light chains is highly minimized, if the immunoprecipitate is fully reduced.
  • Tested applicationsWB more details

Properties

  • FormLiquid
  • Storage instructionsStore at -20°C.
  • Storage bufferConstituents: 1% Proprietary component, 99% Water
  • Concentration information loading...
  • Clonality Monoclonal
  • IsotypeIgG
  • General notesNumber of blots:
    At least 20 blots (based on a 1 µl/ml dilution in 5 ml milk).

    Important protocol notes:

    1. The anti-mouse VeriBlot for IP secondary antibody (HRP) detects mouse IgG antibodies (subtypes: IgG1, IgG2a, IgG2b, IgG3).

    2. The anti-mouse VeriBlot for IP secondary antibody (HRP) preferentially detects the non-reduced form of mouse IgG (IgG1, IgG2a, IgG2b, IgG3) over the reduced, SDS-denatured forms.

    3. IP sample should be completely reduced/denatured before loaded onto a western blot.

    4. Milk should be used as the blocking protein for the immunoblot.

    Western blot and IP resources:
    a) Western blot a beginner's guide

    • b) IP protocol
    c) IP troubleshooting tips
  • Research Areas

Applications

Our Abpromise guarantee covers the use of ab131368 in the following tested applications.

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

Application Notes
WB WB: Use at an assay dependent concentration. The dilution will depend on the sensitivity of the HRP substrate. The recommended dilution is 1 µl in 1 ml milk. Based on the recommended dilution in 5 ml milk researchers can perform 20 western blots. This product is recommended for the western blot detection of IP samples.

Anti-mouse IgG VeriBlot for IP secondary antibody (HRP) images

  • Western blot - Anti-mouse IgG VeriBlot for IP secondary antibody (HRP) (ab131368)
    Western blot - Anti-mouse IgG VeriBlot for IP secondary antibody (HRP) (ab131368)

    IP sample preparation: Caspase 7 was immunoprecipitated from 0.5 ml of 1x107 Jurkat cells/ml with 5 ug mouse anti-human Caspase 7.

    WB conditions: Precipitate from 1x106 cells was subjected to electrophoresis, transferred to an PVDF membrane, and immunoblotted with an anti-Caspase 7 antibody.

    Detection:
    Lane 1: Detection with anti-mouse IgG VeriBlot for IP secondary antibody (HRP) (ab131368)
    Lane 2: Detection with an HRP-conjugated anti-mouse IgG H&L secondary antibody
    Lane 3: Lane 1 was re-immunoblotted using an HRP-conjugated anti-mouse IgG H&L secondary antibody. The heavy and light-chains can now be seen, confirming that although the immunoprecipitating heavy and light-chains are present, ab131368 detects only native antibody and not denatured heavy and light-chains.
    Please note the detection of the heavy and light-chains of the immunoprecipitating antibody in Lane 2 but not in Lane 1.

  • Western blot - monoclonal Secondary Antibody to (HRP) (ab131368)
    Western blot - monoclonal Secondary Antibody to (HRP) (ab131368)
    Developed using ECL technique under reducing conditions; exposure time 3 mins; blocking and antibody incubation steps done in 5% milk/TBST
    Lanes:
    1: Marker
    2: Human heart homogenate Tissue Lysate – 20 µg
    3: HeLa Cell Lysate – 20 µg
    4: Mouse heart homogenate Tissue Lysate - 20 µg
    5: NIH3T3 Cell Lysate – 20 µg
    6: Rat heart homogenate Tissue Lysate – 20 µg
    7: H9C2 cell lysate – 20 µg

    All Lanes:
    Anti-Sodium ATPase antibody – Plasma Membrane Marker
    Anti-GAPDH antibody – Cytosolic Marker
    Anti-Histone H3 (di methyl k9) antibody – Nuclear Marker

    Secondary: Anti-Mouse IgG VeriBlot for IP secondary antibody (ab131368) at 1/1000 dilution

    Predicted Sodium Potassium ATPase band size: 112 kDa
    Observed Sodium Potassium ATPase band size: 100 kDa
    Predicted GAPDH band size: 37 kDa
    Observed GAPDH band size: 37 kDa
    Predicted Histone H3 (di methyl k9) band size: 17 kDa
    Observed Histone H3 (di methyl k9 band size: 17 kDa

Protocols

References for Anti-mouse IgG VeriBlot for IP secondary antibody (HRP) (ab131368)

ab131368 has not yet been referenced specifically in any publications.

Publishing research using ab131368 ? Please let us know so that we can cite the reference in this datasheet

Product Wall

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Displaying 1 - 9 of 9 results for Abreviews and Q&A

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Hi. Does the antibody react to the mouse antibody run under a non-reducing SDS-PAGE? I just want to verify the description above. It detects mouse antibody run under native PAGE but not reducing SDS-PAGE. How about in the condition of non-reducing SDS-PAG...

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ab131368 has only been tested in a gel with SDS, but it should detect non-reduced mouse IgG run with or without SDS as it binds to the non-reduced native mouse IgG primary antibody that is used to probe the SDS-PAGE blot

This is what I am looking for. Just one question. Which light chain it is detecting? Kappa or lambda? or both?

We do not know where the epitope this rat monoclonal antibody recognizes is located on mouse IgG, whether light chain or heavy chain, or if it contains amino acids from the light and also the heavy chain. We do know that the antibody will recognize mouse ...

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for example this secondary antibody: http://www.abcam.com/Goat-F-ab-2-polyclonal-Secondary-Antibody-to-Mouse-IgG-Fab-2-HRP-pre-adsorbed-ab98762.html
It is a full-length human protein with a C-terminally located Fc-tag.
We want to detect sialic ac...

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Thank you very much for the time taken giving me furthrt infotmation.

I can confirm that the Goat F(ab')2 polyclonal Secondary Antibody to Mouse IgG - (Fab')2 (HRP), pre-adsorbed could overcome the cross-reactivity with the human Fc. However we r...

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I would like to try the ab218999 for immunoprecipitation, which is the working dilution range for this assay? Thanks,

Thank you for contacting us.

When using ab21899 Anti-beta 2 Microglobulin antibody [B2M-01] (Biotin) in IP we can recommend a working range of 1-10 ug/ml. The exact concentration for your IP experiments should be within this range but could b...

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Thanks for you quick response! See attach is the picture with the label of marker bands.
For Figure 1:
a. Lane 2 and Lane 7 are expected. Lane 2 was the total lysate of 293T transfected with V5-KGA and Lane 7 was the total lysate of Hela as posi...

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There is a protocol note for ab131368 that states:
"4. Milk should be used as the blocking protein for the immunoblot."
I suggest using milk as the blocking buffer in efforts to reduce background on the blots.
Also, please ensure that ...

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Recently, I got the ab60709 and ab131368 from abcam, which are primary antibody for Glutaminase and
anti-mouse secondary antibody for IP, respectively. I have try to validate the WB and IP for ab60709.
Attach is the results I got. Unfortuna...

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Thank you for your enquiry and for including all your data. That was very helpful.
Regarding Figure 1:
a. In which cell lines do you expect Glutaminase to be expressed?,
b. background is quite high in both figures 1 and 3. What blocking buf...

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I recently purchased an anti-Lamin A+C antibody, made aliquots and then stored it at -20C. I used it for the first time the other day for a western blot of nuclear fractions,mitochondrial and ER fractions from mouse heart and liver. The attached file is t...

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Thank you for contacting us. I am sorry that ab8984 is giving poor results in WB.

Have you validated your transfer at the higher molecular weights by Ponceau stain? If the transfer is incomplete, that could account for the lack of the specific b...

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I had performed western blot on rat kidney lysate(whole cell lysate). Lysate was prepared fresh in RIPA buffer with proteolytic inhibitors. 140µg of protein was loaded per well on a 12% SDS-PAGE with 4% stacking. Blocking: 1 hour at 25ºC in 5% ...

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Thank you for contacting us. I am sorry that this antibody is detecting multiple bands in WB on rat samples.
Is the secondary antibody you are using pre-adsorbed against rat? It is possible that the anti-mouse secondary may be detecting the heavy and...

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I just wanted to thank you again for you incredible help

Such an amazing service and responsibility for your products



I feel like I'm in good hands!

Thanks again and have a great day.

Thank you for your feedback! We are always happy to help, and if you have any further questions please let me know.

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"