HA tag detection

Muriel Paste mpaste at dbm.ulb.ac.be
Thu Sep 28 05:57:03 EST 2000


I have been trying to tag my favorite protein with an Heamaglutinin (HA)
tag, however
this tag does not seem to be recognised by antibodies capable of
recognising other
HA tagged proteins.

The tag is clearly in frame, and I can detect protein expression through
metabolic
labelling, so the protein is clearly expressed (in a reticulocite lysate
system) and
I assume that my tag is expressed with it.

I have inserted three repeats of the HA tag at the C terminus of the
protein (which
is not normally cleaved), but there is no spacer between the C terminus
of the my
protein and the HA tag. Furthermore I have noticed that on ocassion HA
tags are separated
from each other by a glycine.

Now I was wondering if these differences are enough to make the
detection of my tag
impossible, or if there are other factors which I have not considered
and which should
be taken into account.

If it is my tag which is not recognised, suggestions as to how to
express the tag with
a higher chance of it being correctly recognised by my antibodies would
be most welcome.

Thanks

Muriel.






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