Recombinant
RabMAb

Recombinant Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)

Submit a review Q&A (3)References (8)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR3101(2)] to Wnt2/IRP
  • Suitable for: WB
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-Wnt2/IRP antibody [EPR3101(2)]
    See all Wnt2/IRP primary antibodies

  • Description

    Rabbit monoclonal [EPR3101(2)] to Wnt2/IRP

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WBmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide (GST-tag) within Human Wnt2/IRP aa 250-350 (N terminal). The exact sequence is proprietary.
    (Peptide available as ab197186)

  • Positive control

    • Jurkat,MCF-7,T47-D, and SKBR-3 cell lysates Human fetal lung tissue

  • General notes

    Previously labelled as Wnt2. 

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

Applications

Our Abpromise guarantee covers the use of ab109222 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
WB 1/1000. Detects a band of approximately 34 kDa (predicted molecular weight: 40 kDa).Can be blocked with Wnt2/IRP peptide (ab197186).

Target

  • Function

    Ligand for members of the frizzled family of seven transmembrane receptors. Probable developmental protein. May be a signaling molecule which affects the development of discrete regions of tissues. Is likely to signal over only few cell diameters.

  • Tissue specificity

    Expressed in brain in the thalamus, in fetal and adult lung and in placenta.

  • Sequence similarities

    Belongs to the Wnt family.

  • Cellular localization

    Secreted > extracellular space > extracellular matrix.
  • Information by UniProt

  • Database links

    • Form

      The WNT gene family consists of structurally related genes which encode secreted signaling proteins. These proteins have been implicated in oncogenesis and in several developmental processes, including regulation of cell fate and patterning during embryogenesis.

    • Alternative names

      • Int 1 related protein antibody
      • Int-1-like protein 1 antibody
      • Int-1-related protein antibody
      • INT1L 1 antibody
      • INT1L1 antibody
      • IRP antibody
      • IRP protein antibody
      • ONCOGENE INT1 LIKE 1 antibody
      • Protein Wnt-2 antibody
      • Secreted growth factor antibody
      • Wingless type MMTV integration site family member 2 antibody
      • WNT 2 antibody
      • WNT2 antibody
      • WNT2_HUMAN antibody
      see all

    Images

    • Indirect ELISA - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Indirect ELISA - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)

      Representative standard curve using ab109222

    • Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222) at 1/5000 dilution (purified) + Mouse brain tissue lysate at 10 µg

      Secondary
      HRP conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

      Predicted band size: 40 kDa
      Observed band size: 34 kDa
      why is the actual band size different from the predicted?



      Blocking Buffer: 5% NFDM/TBST

      Dilution Buffer: 5% NFDM/TBST

    • Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222) at 1/1000 dilution (purified) + Rat brain tissue lysate at 20 µg

      Secondary
      HRP conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

      Predicted band size: 40 kDa
      Observed band size: 34 kDa why is the actual band size different from the predicted?



      Blocking Buffer: 5% NFDM/TBST

      Dilution Buffer: 5% NFDM/TBST

    • Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      All lanes : Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222) at 1/5000 dilution (purified)

      Lane 1 : Jurkat cell lysate
      Lane 2 : MCF7 cell lysate
      Lane 3 : SKBR-3 cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : HRP conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

      Predicted band size: 40 kDa
      Observed band size: 34 kDa why is the actual band size different from the predicted?



      Blocking Buffer: 5% NFDM/TBST

      Dilution Buffer: 5% NFDM/TBST

    • Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      Western blot - Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222)
      All lanes : Anti-Wnt2/IRP antibody [EPR3101(2)] (ab109222) at 1/1000 dilution (unpurified)

      Lane 1 : Jurkat cell lysates
      Lane 2 : MCF-7 cell lysates
      Lane 3 : T47-D cell lysates
      Lane 4 : SKBR-3 cell lysates

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Standard HRP labelled goat anti-rabbit at 1/2000 dilution

      Predicted band size: 40 kDa
      Observed band size: 34 kDa why is the actual band size different from the predicted?

    References (8)

    Publishing research using ab109222? Please let us know so that we can cite the reference in this datasheet.

    ab109222 has been referenced in 8 publications.

    • Bläuer M  et al. Wnt/ß-catenin signalling plays diverse functions during the process of fibrotic remodelling in the exocrine pancreas. Pancreatology 19:252-257 (2019). PubMed: 30792046
    • Xie B  et al. Benzyl isothiocyanate suppresses development and metastasis of murine mammary carcinoma by regulating the Wnt/ß-catenin pathway. Mol Med Rep 20:1808-1818 (2019). PubMed: 31257529
    • Zhou D  et al. Sonic hedgehog connects podocyte injury to mesangial activation and glomerulosclerosis. JCI Insight 4:N/A (2019). PubMed: 31647783
    • Zhang Z  et al. Wnt2 contributes to the progression of gastric cancer by promoting cell migration and invasion. Oncol Lett 16:2857-2864 (2018). PubMed: 30127872
    • Ding SM  et al. The role of cancer-associated fibroblast MRC-5 in pancreatic cancer. J Cancer 9:614-628 (2018). WB ; Human . PubMed: 29483967
    • Feng J  et al. Protective effect of tanshinone IIA against cardiac hypertrophy in spontaneously hypertensive rats through inhibiting the Cys-C/Wnt signaling pathway. Oncotarget 8:10161-10170 (2017). PubMed: 28053285
    • Lu YY  et al. 1118-20, an indazole diarylurea compound, inhibits hepatocellular carcinoma HepG2 proliferation and tumour angiogenesis involving Wnt/ß-catenin pathway and receptor tyrosine kinases. J Pharm Pharmacol 67:1393-405 (2015). WB ; Human . PubMed: 26076716
    • Zhang Z  et al. Association of Wnt2 and sFRP4 expression in the third trimester placenta in women with severe preeclampsia. Reprod Sci 20:981-9 (2013). PubMed: 23322712

    Customer reviews and Q&As

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    1-3 of 3 Abreviews or Q&A

    Question

    Thanks for getting back in touch.
    I compared muscle stem cells from two metabolically distinct muscles – I found that these populations stained differently for Wnt2, with one population having more intense nuclear staining that the other. I have attached images for you to see the difference. In the image with the darkest staining (nuclear), you can see that there are also cells which are negative for the stain, which suggests that it not therefore indiscriminately staining all nuclei. There is some light staining of the cytoplasm also for both populations which I don’t know if this is as expected or due to too high Ab conc.

    OK. I use the Vecta stain Elite ABC kit
    - Grow cells as monolayer and fix in 70% ethanol, wash 2x pbs, 1x water, store in water until staining

    - Block in 1% normal horse serum 30 mins

    - Remove NHS and add primary antibody – Wnt2 (AB109222) 1/200 dilution in PBS (range on data sheet 1/100- 1/250)

    - Inc 1 h at room Temp on rocker

    - Wash 3x pbs

    - Add secondary antibody – biotinylated anti-rabbit (1%), inc as before.

    - Wash cells 3x pbs

    - Add 1% A+B reagent as Vecta protocol states for 30 min

    - Was cells 4x pbs

    - Add substrate – peroxidase (VIP substrate kit) ˜5 mins

    - Wash 3x pbs

    - Wash 1x water, store in water.

    - done

    I look forward to your comments.

    Kind regards

    Read More
    Answer

    Thank you very much for providing the images and protocol info.

    I have checked different publications and databases; all show cytoplasmic staining. The links to the few are attached below;

    http://www.proteinatlas.org/ENSG00000105989

    http://cebp.aacrjournals.org/content/20/5/1021.abstract

    http://www.nature.com/cgt/journal/v16/n4/full/cgt200878a.html

    The following Abcam antibody also show cytoplasmic staining

    https://www.abcam.com/WNT2-antibody-ab27794.html#WNT2-Primary-antibodies-ab27794-3.jpg

    I would suggest tweaking a protocol and then checking if the result improve

    - Fix cell in acetone for 10 minutes, or 4% icecold PFA or icecold methanol
    - Use 5 - 10% NHS for blocking (1hour)
    - Try the normal ICC/IF procedure using fluorescent secondary antibody
    - Use PBST (0.1% Tween) instead of PBS
    -Try no primary control also

    The other question I have is
    - Have you stained nucleus by any DNA dye?

    I hope the results will improve after following these suggestions. Please let me know if you have any feedback or would like to discuss this further.

    Read More

    Question

    Nuclear staining observed instead of cytoplasmic
    Porcine cells monolayer, muscle stem cells
    Vecta stain Kit with AP conjugated antibody

    Read More
    Answer

    Thank you for contacting us.

    I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

    I am attaching our questionnaire so that we can gather further information regarding the samples tested and the protocol used. Once we receive the completed questionnaire, we will look at the protocol and see if there are any suggestions we can make that may improve the results. This information will also allow us to investigate this case internally and initiate additional testing where necessary. If the product was purchased in the last six months and is being used according to our Abpromise, we would be happy to replace or refund the antibody.

    I look forward to receiving your reply.

    PS: Please either fill in the attached questionnaire or attach the protocol you have been using.

    Read More

    Question

    What is epitope site of this antibody, even the general region would be helpful? Also do you know if this antibody will react with other WNT family members?

    Read More
    Answer

    Thank you for contacting Abcam. The peptide maps between amino acids ********* of human Wnt2 (UniProt:  P09544). We have not yet tested whether this antibody will react with other WNT family members, but I did Blast the immunogen sequence and it did not pull out any other Wnt family members. If there is anything else I can help you with, please let me kno

    Read More

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