Re: [Histonet] 4% PFA

From:"J. A. Kiernan"

You do not say what "PFA" is. If the concentration is
4% a fair gues is that it is formaldehyde, perhaps
generated by depolymerizing paraformaldehyde. 

If so, the fixation time is exactly the same for a
2 mm mouse's ovary as for a 10 mm cube of elephant's
liver: 24 hours for reasonable structural preservation,
but a week or two will stabilize both intra- and
extracellular details more thoroughly. 

Formaldehyde (whether generated from the low polymers 
in formalin or the high polymers of paraformaldehyde) 
penetrates tissues nearly as rapidly as water, but 
its chemical reactions with proteins are quite slow. 

Minimal fixation (minutes to a few hours) by formaldehyde 
is necessary for some purposes, and is followed by
cryoprotection, quick freezing and cutting of frozen
sections. The structure, especially in cytoplasm
viewed at high magnification, is rather poorly
preserved. This doesn't matter for many investigations,
especially with the central nervous system. 

Probably your Histonet enquiry will collect many answers 
like this one, and they won't all give you the same
advice. Fixation has to be matched to the needs of the
work being done. There are plenty of books that 
explain about fixatives and how to choose the right
one for a particular job. Every histo lab should have 
a shelf with at least a dozen. 

In a research lab (you did say mouse ovaries!) you cannot 
be far from a library, full of textbooks and also the 
journals that contain the papers cited in textbooks.  
Go to the original source if you are repeating 
someone else's technique and hope to get similar results.

                            John Kiernan.
                            London, Canada.
Petia P Stefanova wrote:
> Hi to all,
> Does anybody know the duration of fixation with PFA?
> My samples are mice ovaries and testis so I guess it won't take long.
> I appreciate your hepl!
> Petia
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