Anti-BACE2 antibody (ab8025)

Hello everyone! I am contacting you because of 2 antibodies we purchased and tested by immunohistochemistry. We tested anti-BACE1 (AB62034) and anti-BACE-2 (AB8025) on pancreas from different species. With both antibodies we see a clear staining of the pancreatic islet (different pattern for each antibody). Since we could not find any publications that used this antibodies for immunohistochemistry, we further tried to verify whether the antibody binds specifically. For this purpose we used BACE-1 KO mice. To our surprise we got the same staining pattern in the pancreas of KO and wild type mice. The staining pattern in the brain also remained the same. From this, we reason that the antibody binds to something else than BACE-1. Unfortunately we do not have yet the BACE-2 KO to verify this antibody. Could you please let us know, how sure you are that these two antibodies are binding to BACE-1 and BACE-2 respectively. We need this information, since we are planing to publish some of our investigations. Thanks in advance for your help! Kind regards,

ANSWER:

Thank you for contacting us and letting us know about this situation. Thank you also for your patience,as I have been in touch with the labs that produce these BACE1 and BACE2 antibodies. The specificity of the BACE1 antibody ab62034 was determined by a pathologist whovalidated the staining pattern in positive and negative control human tissues. The antibody has not been validated on knockout or wild type mouse tissues. The BACE2 antibody ab8025 wastested in Western blot for expected band size, and results were compared to other BACE2 antibodies raised against different regions of the protein. Ab8025 has also not been tested against knockout mouse samples, though it has been tested with wild type murine heart lysate. What species were your pancreas samples from? Are the staining patterns in wild type tissue expected based on literature using other antibodies?Has the BACE1 knockout been confirmed by PCR? I would recommend also trying a BACE1 wild type negative control, such as thyroid, skeletal muscle, or epidermis (BACE2 should be positive in these tissues). Since the BACE1 antibody has not been characterized in murine tissue, it is possible that it is cross-reacting, so additional tests may be necessary to determine the specificity in mice. I am sorry I don't have more data regarding the specificity of the antibodies, but I hope this information will be useful. Please let me know if you have any questions or if there is anything else that we can do for you.

The customer expressed concern that the product is said to cross react with rat and mouse but the immunogen sequence is only found in human.

ANSWER:

Thank you for your enquiry.

The sequence does not exist as an exact match of the 16 residues, but at least four mouse proteins have enough homology with the sequence to indicate that cross-reactivity is possible, having a band of the same size.

Mouse BACE2 has 9 of the same amino acids as the immunizing sequence. In this case, the matching amino acids are enough to create cross-reactivity with mouse BACE2.

We hope this will help.