Overview

  • Product name
  • Description
    Rabbit polyclonal to Axin 2
  • Host species
    Rabbit
  • Specificity
    Milk blocking not recommended
  • Tested applications
    Suitable for: IHC-P, IHC-Fr, ICC/IF, WB, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Chicken
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 800 to the C-terminus of Mouse Axin 2.

    Read Abcam's proprietary immunogen policy (Peptide available as ab32196.)

  • Positive control
    • This antibody gave a positive signal in: SW480, HCT 116, LS147T whole cell lysates; mouse colon tissue lysates and AXIN2 partial recombinant tagged protein

Properties

Applications

Our Abpromise guarantee covers the use of ab32197 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 a concentration of 0.1 - 1 µg/ml.
IHC-Fr Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.
WB Use a concentration of 1 µg/ml. Can be blocked with Mouse Axin 2 peptide (ab32196). Detects a band of approximately 94 kDa (predicted molecular weight: 94 kDa) in endogenous lysates. Ab32197 gave a positive signal against the tagged partial recombinant protein which has an expected molecular weight of 37 kDa.
Flow Cyt 1/50.

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

 

Target

  • Function
    Inhibitor of the Wnt signaling pathway. Down-regulates beta-catenin. Probably facilitate the phosphorylation of beta-catenin and APC by GSK3B.
  • Tissue specificity
    Expressed in brain and lymphoblast.
  • Involvement in disease
    Defects in AXIN2 are involved in colorectal cancer (CRC) [MIM:114500]. They appear to be specifically associated with defective mismatch repair.
    Defects in AXIN2 are the cause of oligodontia-colorectal cancer syndrome (ODCRCS) [MIM:608615]. Affected individuals manifest severe tooth agenesis and colorectal cancer or precancerous lesions of variable types.
  • Sequence similarities
    Contains 1 DIX domain.
    Contains 1 RGS domain.
  • Domain
    The tankyrase-binding motif (also named TBD) is required for interaction with tankyrase TNKS and TNKS2.
  • Post-translational
    modifications
    Probably phosphorylated by GSK3B and dephosphorylated by PP2A.
    ADP-ribosylated by tankyrase TNKS and TNKS2. Poly-ADP-ribosylated protein is recognized by RNF146, followed by ubiquitination and subsequent activation of the Wnt signaling pathway.
    Ubiquitinated by RNF146 when poly-ADP-ribosylated, leading to its degradation and subsequent activation of the Wnt signaling pathway. Deubiquitinated by USP34, deubiquitinated downstream of beta-catenin stabilization step: deubiquitination is important Wnt signaling to positively regulate beta-catenin (CTNBB1)-mediated transcription.
  • Cellular localization
    Cytoplasm.
  • Information by UniProt
  • Database links
  • Alternative names
    • Axil antibody
    • Axin like protein antibody
    • Axin-2 antibody
    • Axin-like protein antibody
    • Axin2 antibody
    • AXIN2_HUMAN antibody
    • Axis inhibition protein 2 antibody
    • Conductin antibody
    • DKFZp781B0869 antibody
    • MGC10366 antibody
    • MGC126582 antibody
    see all

Images

  • Anti-Axin 2 antibody (ab32197) at 1 µg/ml + AXIN2 partial recombinant tagged protein at 0.1 µg

    Secondary
    IRDye 680 Conjugated Goat Anti-Rabbit IgG at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size: 94 kDa



    ab32197 gave a positive signal against the tagged partial recombinant protein which has an expected molecular weight of 37 kDa.

  • ab32197 staining Axin2 in mouse colon sections. Secondary antibody conjugated to Alexa-fluor ® 488 (please see Abreview for further details)

    See Abreview

  • IHC image of ab32197 stained area of normal tissue in a human colorectal cancer FFPE section. Section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30 seconds at 125°C. Section was incubated with ab32197 at a dilution of 1:500 for 1h at room temperature and detected using an HRP conjugated polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
  • All lanes : Anti-Axin 2 antibody (ab32197) at 1 µg/ml

    Lane 1 : SW480 (Human colon adenocarcinoma cell line) Whole Cell Lysate
    Lane 2 : Colon (Mouse) Tissue Lysate
    Lane 3 : HCT 116 (Human Colorectal Carcinoma) Whole Cell Lysate
    Lane 4 : LS147T (Colon cancer cells) (Human) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) (ab65484) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size: 94 kDa
    Observed band size: 94 kDa
    Additional bands at: 32 kDa, 75 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 16 minutes
  • Flow Cytometry analysis of human permeabilised REH B cells labeling Axin 2 with ab32197 (red). Cells were fixed with paraformaldehyde and permeabilized with saponin, followed by incubation with 2ul of undiluted Anti-Axin 2 antibody (ab32197)  in 100µl of cells in PBS for 15 minutes at 4°C. An undiluted polyclonal goat anti-rabbit Alexa Fluor® 488 was used as the secondary antibody. Control - rabbit IgG (white).

    See Abreview

References

This product has been referenced in:
  • Ding Y  et al. Deubiquitinase inhibitor b-AP15 activates endoplasmic reticulum (ER) stress and inhibits Wnt/Notch1 signaling pathway leading to the reduction of cell survival in hepatocellular carcinoma cells. Eur J Pharmacol 825:10-18 (2018). Read more (PubMed: 29454609) »
  • Yuan L & Tian J LIN28B promotes the progression of colon cancer by increasing B-cell lymphoma 2 expression. Biomed Pharmacother 103:355-361 (2018). WB ; Human . Read more (PubMed: 29669301) »
See all 33 Publications for this product

Customer reviews and Q&As

1-10 of 20 Abreviews or Q&A

Application
Western blot
Loading amount
50 µg
Gel Running Conditions
Reduced Denaturing
Sample
Mouse Tissue lysate - whole (colon cancer colitis induced)
Specification
colon cancer colitis induced
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jul 25 2014

Application
Western blot
Loading amount
50 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (HCT116 colorectal adenocarcinoma)
Specification
HCT116 colorectal adenocarcinoma
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jul 25 2014

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Colon)
Specification
Colon
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10 mM Citrate
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Feb 13 2013

Answer

Thank you for your enquiry which has been forwarded to the scientific support team. I can confirm that there was indeed an error and the datasheet has been amended by our datasheets team. I sincerely apologise for any confusion.

The quality of our antibodies and datasheets is very important to us, as is any feedback and data we receive. It is regrettable that the information we had on this particular datasheet was not of the standard both we and our customers expect from our products.

I would like to reassure you that this antibody (ab32197) is tested and covered by our 6 month guarantee for mouse samples, we have positive mouse data from other sources. In the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund. ab32197 Axin 2 antibody is covered by our guarantee for IHC-P, IHC-Fr, ICC/IF, WB, Flow Cyt and in use, Rat and Human samples.

For more details regarding our guarantee, please see the following page from our website:

https://www.abcam.com/index.html?pageconfig=abpromise

I hope this information will be helpful to you. if you have any further questions or concerns, please do not hesitate to contact me.

Read More
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (mouse E10.5 embryo)
Specification
mouse E10.5 embryo
Fixative
Paraformaldehyde
Permeabilization
Yes - PBST (PBS/0.5%v/v TritonX100 (Sigma)
Blocking step
NOVOCASTRA protein block as blocking agent for 30 minute(s) · Concentration: 100% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jun 08 2012

Answer

Thank you very much for these precisions.

I would recommend to use 5% BSA as blocking agent. This is what is used in our lab to characterise the antibodies. We also use this blocking solution to dilute both primary and secondary antibodies.

If the results do not improve with the abovesuggestion,I would be pleased to arrange a replacement, a refund or credit note in compensation.

Read More

Question
Answer

Thanks for sending along the sequence.

Unfortunately the alignment results show there is no sequence similarity between the peptide immunogen used to produce ab32197 and canine Axin2. So I don't think this antibody will cross react with your samples.

I hope this is helpful. Please let me know if you have any further questions.

Read More

Question

I have answered your questions below under your questions.
Thank you for your help

- Could you specify the species and type of lysates and lysis buffer used?
The samples were from human colorectal cancer xenografts (LS174T).
- At what temp the lysates were treated with sample buffer?
Sample buffer added at room temp, with reducing agent. They are then heated to 70C for 10mins and placed on ice until loaded on gel.
- How much protein was loaded in wells?
10ug per well
- At what dilution the antibody was used? Could you provide the details about the troubleshooting performed. It will help us to recheck the quality of this antibody
I tried the following dilutions
1:1000
1:6667
1:3333
1:2000
1:5000
We tried with both 5% BSA and 5% marvel. We also tried different times from 1hour to overnight.
We also tried preclearing the lysates.
We also exposed the blots up to 45 minutes to try and see any bands.
After this we went back to our 96 well assay we are developing as this has always worked with the previous batch of antibody. Running samples in parallel with only the primary antibody batch changing showed clearly that the second batch is not working.
I do have another batch of antibody, which I haven’t tested yet. I was hoping to buy a few vials of this antibody as the first batch has enabled us to establish a 96 well assay.
Finally I am happy providing a replacement from different lot could you confirm if you are interested in trying the new vial.
I would be very happy to receive a replacement vial as so far your company is the only one which had an antibody which worked.
I have some aliquots of the batch we are having problems with if you require any for testing.

Read More
Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused.

I am happy sending you a free vial from different lot; could you confirm the purchase order number of lot GR66059-1. We currently have many orders from ICR so the correct order number will be helpful.

When testing this antibody we heated lysates for 10 minutes at 100C under reducing and denaturing conditions. Although we have used 10ug of lysates I can however recommend trying 20-30ug of lysates. I would also recommend running LS174T lysates as positive control along with xenograft lysates. You can try these suggestions in your next experimental set up.

Read More
Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry free floating
Sample
Chicken Tissue sections (HH 24 trunk section)
Specification
HH 24 trunk section

Abcam user community

Verified customer

Submitted Nov 10 2011

Answer

I am unable to send you both antibodies as replacements but what I can do is send you one as a free replacement and also, as whichever one you receive has not been tested in IHC, I can also send a testing discount code. What that means is that you test the first antibody in IHC, and then write an Abreview on it, either positive or negative and then the testing discount code becomes active. You can then use the code to get any antibody from our catalogue free of charge. I realize that it is not most convenient way but hopefully the first antibody you test will prove to be successful in IHC, then you can use the testing discount code for another antibody. I look forward to your reply.

Read More

1-10 of 20 Abreviews or Q&A

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