Biotinylation of MoAbs

David L. Haviland, Ph.D. haviland at KIDS.WUSTL.EDU
Wed Apr 17 17:41:06 EST 1996


On Wed, 17 Apr 1996, Mick N. Mulders wrote:

Nick:

> I am having problems with labelling my mouse-MoAbs with biotin. Anybody 
> has a good kit that really works.  Already tried Sigam and Amersham. 
> With the first one I get a coupling efficieny of 0.4 per IgG molecule. 
> With the second one 0.0

I've had some experience doing this.  I opt for the modification reagents 
provided by Pierce.  Most of my experience is in using the NHS-Biotin 
reagent wherein my only advice is to use fresh DMSO in which to dissolve 
the NHS-biotin and not to have any Azide or Tris in the solution 
containing the antibody.  I have it work reasonably well for general 
ELISA use - haven't tried with flow work yet.

In theory, biotin LC-hydrazine should work better than the NHS-biotin as 
with the NHS, one does run risk of biotinylating primary amines that may 
be involved in Ag binding.  So far, I've yet to "over-biotinylate" an 
antibody.

Hope this helps,
David

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