Re: Signal Amplification

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From:Jochen Schuck <>
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Hi Zenobia,
I had very nasty background problems with mouse tissue and mouse antibodies.
What works nicely is if you put tissue specific Fab-fragments on your tissue
before the staining proceedure (I used them 1:50 for 1 hour in 5% serum,
goat anti-mouse Fab-fragments in goat serum...). Of course this only works
for antigens with have nothing to do with immunoglobulins biochemically.

Good luck and happy staining!

Jochen Schuck

> Von: Debbie Gae Pepperall <>
> Antworten an:
> Datum: Tue, 28 Mar 2000 14:02:29 +1000
> An: Marsha R Price <>
> Cc:
> Betreff: Re: Signal Amplification
> Hi Marsha
> I noticed you mention the TSA...Have you used the CSA from Dako? I have
> used it....but not impressed with it, as it picks up a lot of non-specific
> staining...beautiful punctate dots on the membrane of cells in both the
> positive and negative sections. I have contacted DAKO...they know of these
> "unfortunate" staining patterns...however cannot give me an answer yet. I
> have tried x-tra washing, blocking, avidin/Biotin Block you name it...even
> diluting the antibody to infinitum. just thought I would throw this curve
> ball into the mix. Hope we can overcome this.
> Have any suggestions? Thanx in advance.
> Cheers
> Zenobia Haffajee
> HAPS, Newcastle, NSW, "down Under".
> At 08:25 AM 27/03/00 -0500, you wrote:
>> Amos,
>> have you heard of Tyramide Signal Enhancement? It works great and is easy
>> to use.
>> Marsha Price
>> On Sun, 26 Mar 2000 09:14:54 -0500 amos brooks <>
>> writes:
>>> Hi,
>>> DAKO makes a Catalyzed Signal Amplification Kit. This amplifies
>>> the signal
>>> enough to cause an antibody that usually doesnt stain on FFPE
>>> (formalin fixed
>>> paraffin embedded) tissues. The procedure is a bit complicated and
>>> rather
>>> expensive but the results can be quite quite good.
>>> Ventana also makes a one step amplification reagent that I've
>>> heard is good
>>> although I've not used it much.
>>> Amos Brooks
>>> Jay Turner wrote:
>>>> I am staining a fractured rat femur with an immunohistochemical
>>> stain to
>>>> IGF-1.  The stain is a standard biotin-avidin peroxidase with DAB.
>>> The
>>>> samples were decalcified in Cal-Ex (Fisher), it was a little harsh.
>>> There
>>>> is stain in the section but it is very faint.  I don't want to re-do
>>> these
>>>> animals and re-decalcify.  I have tried to use a metal enhanced DAB
>>> (cobalt)
>>>> but the color is too close to the hematoxylin counterstain.  Is
>>> there any
>>>> other way to enhance DAB and the staining?  Any suggestions?  Is
>>>> fluorescence the way to go?
>>>> Thanks!
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