Re: [Histonet] neutrophils

From:Barbara Bublava

another point coming to my mind - how does Chloracetate esterase react
when formalin gets acidic like expired folmalin could do?

Barbara Bublava wrote:
> Dear Robert, Dear Gayle
> trying to keep my Message short, maybee I ended up beeing too short
> I did not want to test if tissue can be fixed in 70% Ethanol. I did want
> to show what happens if tissue isnīt fixed enough (Dorothy mentioned
> expired formalin) and wents to the processor - often starting with 70%
> alcohol - or transferred, as Robert mentioned, to it for longterm storage.
> I tested the "extremes"!
> 1.) no fixation (70% alcohol, trying to simulate starting the processor
> with unfixed specimen)
> 2.) "half" or partial fixation (transferred to 70% before fixed
> completely - which means a rather big specimen for 4 hours, fixed at the
> edges, not in the center) - trying to simulate staring the processor
> with partial fixed specimen. The case one doctor "produced" by insisting
> on fast processing
> 3.) and "proper" fixation. (24 hrs - for a middle sized specimen, fixed
> in the center also) - correct procedure
> I did this short test to demonstrate how the incosistant staining the
> doctor claimed did happen and to show how important fixation is for a
> correct staining (nothing really new to us, but some doctors...)
> I did choose Chloracetate Esterase Kit because it was the stain burdend,
>  HE also did show differences but after all it could be interpreted.
> CAE did fail comletely!
> I donīt know if this is the solution for Dorothyīs problem, since I did
> my little test on human tissue and do not know if the stain will work on
> rodent, like other people considered.
> If alcohol and not autolysis is the problem - then it could maybe help
> to move the specimens to fresh formalin before processing?
> I know there are impairments in my little demonstration - for example
> the time specimens were left in 70% Ethanol (before moving to the
> processor) differs from 0 to 24 hrs (I wanted to process, cut and stain
> them together - so there can be no differences)
> a lot more testing could be done, and I would do it if I had the time
> for it...
> but at least and for my intention the test did what it should do. The
> doctor understood the overhelming importance of correct fixation.
> Barbara Bublava
> Medical University of Vienna
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