AW: [Histonet] overfixation with formalin

Gudrun Lang gu.lang <@t>
Sun Nov 4 02:02:52 CST 2012

This margin effect was especially seen in needle biopsies. In 2 mm cores.
But he showed also a lymphnode slide, I would estimate 0,5-1 cm, with a
margin-effect, that causes bad cutting in the center (loosened aereas). In
this context he said, that a lymphnode sitting in higher concentrated
formaldehyde over the weekend isn't properly fixed in the center because of
the hardened surface.

I'm not with his opinion.



Von: Charles.Scouten <@t>
[mailto:Charles.Scouten <@t>] 
Gesendet: Samstag, 03. November 2012 23:20
An: gu.lang <@t>; histonet-bounces <@t>
Betreff: Re: [Histonet] overfixation with formalin


Is this due to the well known effects that formaldehyde penetrates tissues
very slowly, at a rate of about 18mm/25 hours.  And that autolysis (tissue
breakdown) begins to occur  immediately with extraction.   If so, then in
your tissue the center decayed before the fixative reached it.  How large
was the starting piece of tissue, how far was the center negative area from
any direct exposure to formaldehyde?  Was it precut in slices less then a mm
thick, before being dropped in fixative? 


Charles W. Scouten, Ph.D 
Product Specialist 
Leica Biosystems Richmond, Inc.
5205 Route 12
P.O. Box 528
Richmond, IL 60071
United States of America 
Telephone 630 964 0501 
facsimile +1 630 964 0576 

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From:        "Gudrun Lang" <gu.lang <@t>> 
To:        <histonet <@t>> 
Date:        11/03/2012 01:42 PM 
Subject:        [Histonet] overfixation with formalin 
Sent by:        histonet-bounces <@t> 


Hi histonetters!

I'm just attending a histo-course, where the teacher told us his opinion
about overfixation.

For him overfixation takes place in any formaldehyde solution with a
concentration above 5%. This should cause the margin-artefact, that leads to
false-positive IHC at the margins of the tissue and to false-negative
results in the center. The higher concetrated fixative should harden and
shrink the surface, so it cant be penetrated any more by the fixative.

I told him about the publication of Cecil Fox, who saw shrinkage only in
solutions with formaldehyde concentration above 30% (I think) and said, that
the methanol-part is responsible for that.

I believe, that these margin-artefacts are due to drying at the time of
biopsy or an effect of the needle-shot itself. (But believing is no

In our lab we use 8% formaldehyde as standard fixative, buffered with
low-molar phosphatebuffer. There are no complains from the doctors about

Please help me with the histonet-wisdom. What's your opinion? 


Gudrun Lang

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