Re: Ready to use LSAB Primary Antibody

From:Connie McManus <conmac@cc.usu.edu>

Jenny,

You mentioned that your pos ctrl works great.  Why do you think your tissues
should be as good as the control? I don't know if  if the presence of HBV in
the tissues I work wtih (this is what I work with in IHC) is going to be a
strong one, a weak one or nonexistent.  Do you run a negative control as well
as a positive control?  You need to run both controls.

How long do you incubate in the antibody?  I use the LSAB peroxidase kit, not
the prediluted antibodies you're talking about, but I have found that by
increasing incubation time from the recommended 10 minutes to 30 minutes in the
core antigen, link antibody and streptavidin steps, my tissues stain very
nicely.

Do you do antigen retrieval?  I don't on my tissues, except on one occassion
where someone placed the tissues in lq N2 with no cryopreservant or other
fixation, then they were inappropriately thawed.  Those tissues were a
MESS...freeze artifact ruled.  I did antigen retrieval on these tissues and
amazingly I got some staining.  I thought is was artifact at first, but when
compared side by side to the same sections without the AR, I had to conclude
the staining was real. They were not reliable for the project, but it was
interesting to see how AR can work wonders.

Best wishes and Happy New Millenium!

Connie McManus
Veterinary Diagnostics Laboratory
Utah State University
Logan, Utah
USA
ph (435) 797-1891

Jenny Molde wrote:

> Dear Histonetters
>
> Is anyone using the ready to use prediluted Dako LSAB antibodies? I am
> using them with the Dako LSAB 2 kit and am getting not too good a staining.
> Very weak and sometimes nothing at all although my positive control works
> like a bomb. Your expertise would be greatly appreciatted. Many thanks in
> advance.
>
> Jenny Molde
> University of Cape Town
> Cardiovascular Research Unit
> Anzio Road
> Observatory
> Cape Town
> 8001




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