Epitope Tagging Questions...

hadi al-hasani hadia at bdg10.niddk.nih.gov
Fri Mar 28 22:20:00 EST 1997


Well, it depends... From my experience, there is no way to predict
which tag would be the best. It depends mainly on your construct: C-,
N-terminal tag (best) or internally located (tricky). Most of the
people use HA, FLAG or myc, but only because they have read that most
of the people use...

Concerning the HA epitope: there is a company (Babco =Berkely
Antibodies) which offers mono- and polyclonal anti-HA's. At least in
our hands the mAB works fine in IP and WB .
Recently there was a post from these guys which I repost here:

>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>
>Dear Colleagues,
>
>I will soon be embarking on a project to epitope-tag a putative
>membrane protein in mouse small intestine, to be expressed in Xenopus
>oocytes and mammalian cell lines.  I am aware of three different kinds
>of epitopes for which monclonals are available commercially:  HA (from
>influenza hemagglutinin), 9E10 (from human c-myc protein) and VSV-G
>(from esicular stomatitis virus glycoprotein).
>
>Are there other epitopes available?  What are the dis/ advantages?
>

Perhaps we could be of some help.

Our company sells a number of epitope tag antibodies, as
indicated below:

Name          Sequence 

HA           YPYDVPDYA
myc          EQKLISEEDL 
AU1          DTYRYI     
AU5          TDFYLK         
glu-glu      EEEYMPME    
KT3          KPPTPPPEPET
IRS          RYIRS
FLAG         DYKDDDDK

In the case of the HA antibodies, both mouse monoclonal and rabbit
polyclonal antibodies are available.  All of the other antibodies are
mouse monoclonals.  In each case, the peptide recognized by the
antibody is also available.  

We have a few other related antibodies as well, including an improved
anti poly-histidine antibody, and fluorescenated and biotinylated
antibodies, affinity resins, and antibodies against commonly used
fusion protein partners.

If any of these are of interest, please let me know.

We also have an annotated bibilography of the epitope tagging
literature that we will send along to any interested parties.

Tom Anderson
>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>>
(the guy's e-mail  adress was >babco at ix.netcom.com< if you are
interested. But I am sure, there are much more companies out there
selling these AB's.)

In our lab FLAG (AB purchased from Kodak) seems okay too, but
performed poorly with certain internally tagged proteins. However,  I
believe that if your tag is perfectly exposed to the solvent you could
use any AB - and if not.....well you have to try....

hope that helps 

/  hadi al-hasani
/  hadia at bdg10.niddk.nih.gov


On Fri, 28 Mar 1997 12:20:42 -0600, Simon Twigger
<simont at post.its.mcw.edu> wrote:

>Hi there,
>
>We would like to epitope tag our protein and Im trying to find out a bit
>more about the pros and cons of different tags. At the moment we are
>looking at the myc tag as it is small, ca. 10 residues, and there are
>commercially available antibodies for it. However these antibodies are
>rather pricy (ca. $200 for 100ug from Calbiochem or Pharmingen). I have
>tried to find information on HA tags but Im having trouble finding the
>epitope sequence and I can find noone selling antisera for it in our
>catalogues. 
>
>Does anyone know of a cheaper source of anti-C-myc antibodies?
>
>Can anyone point me to more information on the HA epitope/antibodies?
>
>Are there any other tags people would recommend (we dont really want to
>use a His tag)?
>
>Also, any words of wisdom on epitope tagging in general would be most
>appreciated. :)
>
>	Thanks in advance for any help with this.
>
>	Simon.
>
>
>Simon Twigger, 
>Medical College of Wisconsin, Milwaukee.
>simont at post.its.mcw.edu




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