Anti-HA tag antibody (ab13834)

Hi Technical,



We have a customer who is looking for a HA tag antibody and a matching secondary, I have sourced and found him the following;



http://www.abcam.com/HA-tag-antibody-ChIP-Grade-ab9110.html

http://www.abcam.com/HA-tag-antibody-ab13834.html

http://www.abcam.com/HA-tag-antibody-Agarose-ab27029.html

http://www.abcam.com/HA-tag-antibody-ChIP-Grade-ab71113.html

http://www.abcam.com/Proteinase-K-ab64220.html



The customer’s first email;

I received your email address from a colleagues at the school of biology sciences. I am looking for some things that you probably sell. First of all an rabbit anti-HA antibody and an anti-rabbit FITC-conjugated antibody (I think they are both from Abcam). And secondly proteinase K. Do you sell these and can you give me a quotation for each of them?



But as he is new to this, he has provide the following details, as he didn’t provide much in the first place.

The customer’s Reply;

Since I’m relatively new to this field of research (I do more behaviour than immunohistochemistry), I’m not entirely sure which of the antibodies is the best. However, perhaps I can explain the experiment and you might know better . We are currently transfecting HEK cells with a mutant receptor that has an HA tag and we want to see whether the receptor is nicely incorporated into the membrane or whether it is somewhere else (using confocal laser microscopy). So we need a good anti-HA antibody for that (and indeed the anti-rabbit conjugated antibody would then be the secondary antibody. I hope that help.

Are you able to help me out with this, as it is out my field?



Looking forward to your response.





Kind regards,



Melissa Gliddon

Customer Service and Technical Support Manager | mailto:melg@sapphirebioscience.com

________________________

Sapphire Bioscience Pty. Ltd.

126 Cope Street │ Waterloo NSW 2017 │ Australia

Ph: +61 2 9698 2022 │ Fax: +61 2 9698 1022 | http://www.sapphirebioscience.com/

ANSWER:

Thank you for your enquiry, I am pleased to help.

From what I understand here, the customer would liketo detect the HA tag on their recombinant protein expressed in cell culture. This is Immunocytochemistry (ICC). I have reviewed the selection you made:

ab9110 Rabbit polyclonal tested ICC-IF. This will be suitable

ab13834 Rabbit polyclonal tested WB not ICC.This one willnot be guaranteed in ICC-IF for their experiments.

ab27029 - this is conjuated to agarose beads for IP so will not be suitable.

ab71113 - Rabbit polyclonal tested ICC-IF. This will be suitable.

Proteinase K:
Could you confirm for what reason do they need proteinase K? Antigen retrieval is not usually required in ICC and I would suggest to try it only if there is no staining in order to help expose the antigen. In ICC, Fixation should be in just 4% PFA for 10 minutes. Antigen retrieval is usually used for IHC on paraffin embedded sections which are fixed in 10% PFA for at least 24 hours.

There are many suitable secondaries in the catalog. Here are two exmaples of anti-rabbit secondaries tested in ICC-IF conjugated to FITC as requested by the customer:

ab97050 Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (FITC)
Tested in: Flow Cyt, ICC/IF, IHC-P
http://www.abcam.com/index.html?datasheet=97050 (or use the following: http://www.abcam.com/index.html?datasheet=97050).

ab97199Goat polyclonal Secondary Antibody to Rabbit IgG - Fc (FITC)
Tested in: Flow Cyt, ICC/IF, IHC-P
http://www.abcam.com/index.html?datasheet=97199 (or use the following: http://www.abcam.com/index.html?datasheet=97199).

We also have an ICC webinar coming soon which they may like to consider reviewing. This is being held next week, regrettably not at a convenient time for Australia. However, a copy will be available on our blog (link at the bottom of the Abcam homepage)in early May.

The following link aboutthis webinaralso contains links to some ICC protocols (bottom left hand side)which I hope will be helpful:
http://www.abcam.com/index.html?pageconfig=resource&rid=14548

I hope this will be useful. If you have any further questions, please do not hesitate to contact me.