Anti-APC antibody (ab15270)

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Dear tech,

A customer would like to know thatab15270 recognizes full length protein of APC or truncation of the APC product. Would you please help to check it ? Thank you for your help!

ANSWER:

Thank you for contacting us.

ab15270 should only detect the full length APC protein.

I hope the information is useful to you. Please do not hesitate to contact me if you have further questions.

If you could arrange for the alternate antibody to be sent that would be ideal,    

ANSWER:

As requested, I have issued a free of charge replacement with the order number 996071 (one unit of ab15270).

To check the status of the order please contact our Customer Service team and reference this number.

Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

I wish you the best of luck with your research.

What is a good negative control for me to use in a Western blot?

ANSWER:

I am sorry that I don't know which cell line does not express APC. The antigen of this antibody is derived from the C-terminal end of human APC, and therefore you can pick a colon cancer cell line (with a known APC mutation and truncated C-terminal) as your negative control.

Please contact us again if you have any additional questions.

What is the staining pattern with this antibody?

ANSWER:

Thank you for your enquiry. The originator of this antibody has said that the staining pattern is cytoplasmic. I did quickly read in the following article that APC protein is located in both the cytoplasm and the nucleus and "APC is a nucleocytoplasmic shuttle protein whose predominantly cytoplasmic localization requires NES function and suggests that APC may be important for signaling between the nuclear and cytoplasmic compartments of epithelial cells."

Proc Natl Acad Sci U S A. 2000 October 24; 97(22): 12085–12090.

I hope this helps, please let me know if you have any additional questions.

In this paper, Carcinogenesis 21(11): 1935-1940, November 2000, the authors state "In the normal colonic mucosa APC immunoreactivity was present in all 150 of the samples examined" and "APC immunoreactivity was totally absent in 83% of the 83 colorectal cancers examined". In addition, "the frequency of detectable APC mutations is similar in colonic adenomas and carcinomas (~60%) has suggested that APC mutations may be an early or even the initiating event in the process of colonic carcinogenesis" and almost all of the mutations found in the APC gene are insertions, deletions or nonsense mutations that lead to a stop codon and result in the production of a truncated protein that has lost the C-terminal end."

My question, then, is if this paper is correct and this antibody is made from the c-terminus, how can colon carcinoma be an effective positive control?

ANSWER:

Thank you so much for your patience with this enquiry. I have been in touch with the antibody's originator to obtain some clarification regarding this antibody.

Your customer is correct - colon carcinoma should not be recommended as a positive control. The originator tested it on both normal colon and colon carcinoma and subsequently found that in the literature most colon carcinoma is negative for APC expression.

So we are updating the datasheet to state that normal colon tissue is recommended as a positive control, and I'm also requesting an image of staining with ab15270 in normal colon tissue. Once I have that, I will add it to the datasheet.

If you have any more questions, please let me know.