Re: biotin blocking and non-immune sera for negative control

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From:"Bonnie P Whitaker" <>
To:"Amos Brooks" <>, "Sonia Greco" <>
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The blocking Amos describes is the serum or protein block...... the a/b
block is at any point after any HIER and before secondary........ I always
do mine before protein block ........ and a/b should be well rinsed after
each (unlike protein block)..... just wanted to clarify that point.
Bonnie Whitaker

----- Original Message -----
From: Amos Brooks <>
To: Sonia Greco <>
Cc: 'HistoNet Server' <>
Sent: Saturday, May 20, 2000 7:49 PM
Subject: Re: biotin blocking and non-immune sera for negative control

> Hi,
>     The blocking is put on just before the primary antibody then it is
> or blown off (not rinsed). I agree the directions are vague. One might
think it
> is before H2O2 or for that matter before epitope retrieval. By the way it
> used after enzyme predigestion if you are using that.
> Amos Brooks
> Sonia Greco wrote:
> > Dear Histonetters,
> >
> > I just bought the DAKO Biotin Blocking System, which suggests to use
> > application of the first step of the staining procedure. Does this mean
> > after the secondary antibody, before a strepABC and DAB staining. or do
> > apply it before between the primary and secondary antibodies?
> >
> > Also: I am doing immunohistochemistry with a polyclonal goat anti human
> > antibody. For a negative control, I would have liked to use a goat
> > non-immune IgG, but didn't have any, so I have used normal goat sera.
> > you know the best dilution to use to make it a good negative control?
> >
> > Thanks in advance
> >
> > Sonia
> > QIMR, Australia

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