Re: human-human ihc

From:Barbara Wright


If Steve puts 40% human serum in his secondary reagent which, if I am understanding
correctly should be a anti-human IgG - the 40% human serum should bind to all of
the anti-human IgG in the secondary reagent leaving no binding for the primary
antibody.  Steve can put the 40% human serum in his protein blocking reagent and in
his primary antibody - but I would leave it out of the secondary.  Am I reading it

Barb Wright

rueggp wrote:

> what i would do for this is use lots of normal human serum to block with before
> the primary antibody and again before the secondary, i would also put human
> serum in my secondary reagent.  by a lot i mean about 40% solution.
> patsy ruegg
> "Crochiere, Steve" wrote:
> > Hi,
> > Does anyone out there have an IHC procedure for staining human tissue with
> > human Mab's? I have tried to do this by blocking endogrnous peroxidase with
> > H2O2,  IgG activity with a FAB fragment, protein with normal serum, and
> > biotin with the vectorlabs biotin blocking sysytem, and still have so much
> > background and non-specific staining in the negative controls (IgG whole
> > molecule) that the slides are unreadable.
> > Any suggestions would be greatly appreciated.
> > Thanx,
> > steve

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