[Histonet] Re: Antibody problems

From:"PW Howorth, Physiology"

Hi folks,

Recently, I have been enduring a problem which is undoubtedly familiar to 
some and I was wondering how have you have overcome the dilemma that I'm 
facing.  The problem is that I need to do some double labeling, however, 
the current antibodies that I use are derived in rabbit and in both 
methods, I use a biotinylated secondary step, followed by strept-avidin 
Cy3.  The obvious step is to use different species with conjugated 
fluorophore secondary, but having spent 2 months in frustration, I am 
willing to consider other options, namely; FAB fragments to change 1 
antibody species from rabbit to say, goat; and avidin-D kit to block 
endogenous biotin (but in this case, it would be spare biotin sites 
remaining from first antibody detection).

Do any of you good folks, use either of these methods or have tried them 
and would be willing to share your thoughts ?

Many thanks


Patrick Howorth,
Dept of Physiology,
University of Bristol,
University Walk,
BS8 1TD.

+44 (0)117 33 17112 (office)
+44 (0)117 33 17579 (lab)

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