anti Tag antibody cross reactivity

Jason via (by jason.d.fernandes from
Mon Oct 26 23:17:02 EST 2009


Adding a small tag makes it easy to purify, IF, or id by western.  In
general a small flexible linker is a good idea GGGS is a standard
linker but it will depend on how you clone it.  N or C terminus will
depend on your protein but in general there is not much of a
difference (global TAP studies show little difference between N-
terminal tagging and C-terminal, but if you know an important
interaction occurs at one end you may want to avoid tagging it
there).  FLAG is considered a very clean tag, it is also very
expensive reagent wise and depending on what you do you might denature
the antibody (i.e no denaturing purification).   His is a workhorse
tag that can take a lot of abuse but you will purify polyhistidine
proteins from mamallian extract. It is also very cheap. Strep II from
IBA is a nice small clean tag that I use frequently. HA and myc are
also fairly well established and generally considered cleaner than his
but not as clean as FLAG (also in general I use 3xFLAG).


On Oct 25, 7:16 am, Azam Rahimpour <rahimpou... from> wrote:
> Hi
> does anybody use different epitope tags for proteins expressed in mammalian cells? I have some questions about this:
> 1. Is it important to add small tags to either C or N terminal of proteins?
> 2. Is it necessary to put an spacer or specific sequence between tag and protein coding region?
> 3. which anti tag (FLAG, His, myc, HA ...) has lower cross reactivity to cellular proteins?
> thanks in advance

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