RE: Neg Immunocontrols

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From:brian chelack <> (by way of histonet)
To:histonet <>
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Hello John,

I was just lurking for the weekend to see if anyone would have any other ideas
regarding a true negative control for polyclonal antibodies.

In my mind the best true control for a polyclonal antibody is the very same
with the addition of the specific antigen it is directed against.  We have
used this
method in liquid phase tests using insulin and anti-insulin antisera.  I
know that
there are bound to be problems with this method on tissue sections due to
conformational changes in antigen structure in a solid phase compared to its
immunizing form, but absorbtion controls seem to me to be the best
alternative for
a negative control.

Anybody who agrees, disagrees or just wants to argue because its Monday
morning is invited to comment.


BJC - Brian
On Thu, 15 Oct 1998 08:10:34 -0700 Tarpley, John wrote:

> From: Tarpley, John <>
> Date: Thu, 15 Oct 1998 08:10:34 -0700
> Subject: RE: Neg Immunocontrols
> To: "A. Mark Briones" <>,
>      'brian chelack' <>
> Cc: HistoNet Server <>
> The point on polyclonal antisera is well taken and that's why, whenever
> possible, we use preimmune serum from the same animal or pool of animals
> used to produce the antibody diluted to the same Ig concentration as the
> primary antibody. By using this control you can see if any "background"
> antibodies already present in the serum will affect your results.
> Unfortunately, this is rarely possible when using commercial antibodies. Can
> anyone suggest a true negative control for polyclonals in this case?
> John Tarpley 15-2-B
> Specialist Image Analysis & Immunohistochemistry
> Amgen Inc
> One Amgen Center Drive
> Thousand Oaks, CA  91320
> > ----------
> > From: 	brian chelack[]
> > Sent: 	Wednesday, October 14, 1998 5:07 PM
> > To: 	A. Mark Briones
> > Cc: 	HistoNet Server
> > Subject: 	Re: Neg Immunocontrols
> >
> >
> >
> > Just a quick observation regarding the use of "non-immune sera" as a
> > negative
> > control.  Investigators should be aware that non immune sera will always
> > contain
> > antibodies to other common pathogens that the negative control animal has
> > experienced in its past.  A case in point in our lab was the use of
> > "non-immune
> > rabbit sera as a control alongside a rabbit antisera for Bovine
> > respiratory syncitial
> > virus.

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