Anti-AKAP12 antibody [JP74] (ab49849)

Ab tested in human and mouse
positive control: Rat brain cytosolic S1 fraction
can Ab be used with rat samples?

ANSWER:

Thank you for your patience.

I sincerely apologize about this delay, but I have good news:

Yes, rat is a tested species and I have updated our datasheet to reflect that. We therefore guarantee this antibodyto work with rat samples.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Ab tested in human and mouse
positive control: Rat brain cytosolic S1 fraction
can Ab be used with rat samples?

ANSWER:

Thank you for contacting us.

I have received the following information from the lab so far:

Wedid a Blast comparison of the rat and human proteins and although the overall identity was only 68%, there are long stretches of >90% identity throughout the two sequences, probably accounting for recognition of the rat protein by the antibody.

We are inquiring about why rat is not listed.

I will keep you informed once I receive more information.
Thank you for your patience.

Ab tested in human and mouse
positive control: Rat brain cytosolic S1 fraction
can Ab be used with rat samples?

ANSWER:

Thank you for contacting us.

The lab is looking into this and will let me know if there is a reason why rat was not listed as tested species. I will get back to you when I have more information.

Thank you for your patience and understanding.

Dear Sir or madam,
months ago we purchased AKAP12 antibody from Abcam (ab49849). In the antibody's datasheet is written that apart from WB and IP the antibody can be used for ICC (IF) as well. We have performed IF (fixation with paraformaldehyde (PFA) and permeabilization with Triton). This IF was compared with another IF where self-made antibody was used and it showed totally different staining! Since you haven’t shown any ICC pictures, neither you listed any references for IF I wonder whether the antibody was tested for ICC? I would appreciate any comment and/or recommendation.
Otherwise I have successful WB and IPs!
Looking forward for your reply

ANSWER:

Thank you for contacting us and for alerting us to the problem you are experiencing with our Anti-AKAP12 antibody ab49849. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

Indeed, this antibody has been tested in ICC/IF, and as we have received no complaints about this product, I am concerned it is not working for you. Thus, I would like to investigate this particular case further for you. In order to proceed with this, I have enclosed our questionnaire so that we can gather further information regarding the samples you tested and the protocol you used. I would appreciate if you could complete this, and also provide some information about the self-made antibody (e.g. which part of AKAP12 it recognises and which species it was raised in). I would also appreciate if you could also provide an image which would help us to assess the results.

Once you have submitted the form to us, we will look at the protocol you used and see if there are any suggestions we can make to improve your results. If we determine that the antibody is not working as stated on the datasheet, and as you have purchased it in December (i.e. in the last six months), we would be happy to replace or refund the antibody for you.

We look forward to receiving your reply.

Dear Sir, I am using your antibody Akap12. With this mail i am attaching western blot figure which was developed using ab49849 antibody. I would like to confirm from you which is the exact Akap12 band. In your product details you have mentioned the molecular weight of Akap12 is approximately 300Kda (predicted is 191Kda). I have used high range molecular weight marker in one side and the other side is colour marker. In this figure which band is the exact Akap12 band (Whether first red arrow from the top or second arrow from the top). Figure 1 and 2 are same with different time exposure time. I have used 7% gel for this study. The sample used is cell lysate, not animal tissue.    I will be happy if the reply is at the earliest.

ANSWER:

Thank you for contacting Abcam. I have looked at the western blot image that you attached and I believe that the Akap12 band is actually the large band a the top of the image, not marked by a red arrow. If you would like my opinion of which band is the closet to 191kDa, then I would chose the top red arrow, but again I believe the actual Akap12 band is higher. Based on the datasheet and the references listed on the datasheet (PubMed: 21831305), the band that is of interest is at 300kDa. To achieve an image that has better resolution, you could rerun the gel and this time let the gel run longer, so that you get rid of all the low molecular weight products and gain a greater separation of the higher molecular weight proteins. Also using a molecular weight marker, that goes to 300kDa would also allow more accurate sizing. Please let me know if there is anything else I can help you with.