Actually a truly humanized antibody has human Fc and human Fab.  The only
murine part of the original mAb that's left is are the 6 CDRs that determine
epitope binding.  There's not really anything for a mouse secondary to bind
to (since the only murine part is presumably bound to the antigen).  Mouse
secondaries will only work with mouse/human chimeras (mouse Fab/human Fc) or
hemichimeras (mouse/human or human/mouse light and heavy chain swaps).
Nick's suggestion is probably the best way to go--this is esentially the
same process that the DAKO Animal Research Kit uses for mouse mABs on mouse
tissue and it works quite well.

Scott Turner
DNAX Research, Inc
Palo Alto, CA

-----Original Message-----
From: Carrie Kyle-Byrne [mailto:ckbyrne@exelixis.com]
Sent: Friday, April 11, 2003 7:53 AM
To: joyce judge
Cc: 'HistoNet@Pathology.swmed.edu'
Subject: Re: human Ab on human


joyce,
there is a difference between a true human anti-human antibody and a
humanized antibody.  i'm assuming that your antibody is a humanized murine
monoclonal.  humanized antibodies are a fusion of human Fab and murine Fc
(there's more to it but this is a simplified explanation).  the only part of
the antibody that is human is the part that will bind to the tissue.  the
part of the antibody that you will be binding secondary to is MOUSE.
therefore, you need to use an anti-mouse secondary.
Carrie Kyle-Byrne, BHS, HT(ASCP)
Assoc. Research Scientist II
Antibody Core Lab
Signal Transduction Research

Exelixis, Inc.
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----- Original Message -----
From: "joyce judge" 
To: 
Sent: Friday, April 11, 2003 5:00 AM
Subject: human Ab on human


>
> I have gone through the histonet archives to look for answers regarding
> Human Ab on Human tissue. I was wondering how people have  resolved this
> issue.  I will need to do IHC on a "humanized" antibody in human tissue
and
> we can not biotinylate or conjugate the primary at all.  Can anyone offer
> any help?
>
> Joyce Judge
>
>
>
>
>
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