Anti-HA tag antibody [HA.C5] (ab18181)

Hi,

Thanks for the response. I did not receive your previous email for some reason.

I'd like to get a replacement antibody: ab18181

Please let me know how to proceed. Our mailing address should be in the order information I previously sent (the address below would also work):

ANSWER:

Thank you for getting back to me. I am sorry that my previous e-mail has not reached you.

This is to let you know that I have placed a new order for you - for one vial of ab18181 as a free of charge replacement (exchange for the original item: ab9110) and the new order number is 961284.

I hope the second vial will work as it is expected, and please do let me know how you are getting on with this product.

Good luck with your research!

Hello,

First, do you have any references for your N-ChIP protocol?

Second, we are interested in detecting DNA/transcription factor (TF)crosslinks. We would like to immunoprecipitate an epitope-tagged TF that has been incubated with and crosslinked to its binding sites on genomic DNA. We are now looking for an appropriate epitope tag to use, both for purifying the expressed TF and then for pulling down the DNA/TF conjugates. This is similar to, but somewhat simpler than, chromatin immunoprecipitation (which is usually an in-vivo experiment). I believe your X-ChIP protocol describes the conventional ChIP experiment, but your N-ChIP protocol looks like what we are interested in. Is this the case? Would any tag that is "IP" competent be suitable for such an immunoprecipitation experiment? Are any suitable for protein purification?

ANSWER:

Thank you for your enquiry.

NChIP involves the preparation of native, nuclease digested chromatin followed by the immunoprecipitation of chromatin using appropriate antibodies:

We have an excellent protocol for both the preparation of nuclease digested chromatin:

http://ops.abcam.com/index.html?pageconfig=resource&rid=31&pid=5

and the immunoprecipitation of chromatin:

http://ops.abcam.com/index.html?pageconfig=view_protocol&pid=293.

Both of these protocols were developed by Laura O'Neill and Bryan Turner. They have published their NChiP protocol in the following publication:

O'Neill LP, Turner BM. (2003) Immunoprecipitation of native chromatin: NChIP. Methods. 31(1):76-82. PMID: 12893176

Please forgive me if I am incorrect but given that you are using crosslinking in your detection of transcription factor interaction with genomic DNA your approach is more akin to XChIP. I would recommend an epitope tag that has been validated for its use by chromatin immunoprecipitation. We have an excellent antibody in the form of ab9110 HA, tag antibody - ChIP Grade. This has been shown to be capable of precipitating chromatin under cross linking conditions.

I hope this information helps, please do not hesitate to contact us if you need any more advice or information.