paraformaldehyde and colloidal iron staining of mouse tissue

From:Sharron Ladd

Dear Histonet,
I  work in a University research setting. I have been helping someone 
who knows absolutely nothing about histotechnique with Colloidal iron 
staining of various mouse tissues (including liver, spleen and kidney). 
The mice were perfused with 4% formaldehyde (made from paraformaldehyde) 
and the tissues were then stored in 10% neutral buffered formalin (made 
in the lab). He is using Van Gieson solution as a counterstain. He tried 
the stain several times and each time his sections turned green. So, we 
each ran the stain side by side and threw in human appendix that had 
been fixed in commercially made 10% neutral buffered formalin. 
Everything turned green except the appendix!!
I have read many of the past postings about paraformaldehyde vs. 
formaldehyde, especially Dr. Kiernan's postings. Many, many people here 
at the university make their own 4% formaldehyde (from paraformaldehyde) 
and that is all they will use. Dr. Kiernan said in his posting that the 
reason this is done is "tradition, dating from about 1960." Some 
professors claim morphology is better. I also think it is used because 
some of the tissues used to be set aside for EM and 4% formaldehyde 
(from para) worked for both EM and light microscopy. I have 10% formalin 
on my tissue processor only for running cell blocks, but some people get 
very upset when they see it there. They don't want 10% formalin touching 
their tissue.
In the colloidal iron staining situation above, I suggested perfusing 
the animals with 10% NBF instead. I got the same reaction as always, 
WHY?--they are both formaldehyde and what if the methanol in the 
commercial preps does something awful to my tissue??
This email is getting awfully long, so I guess what I want to know 
is...Can perfusion fixation in 4% formaldehyde (made from 
paraformaldehyde)  interfere with colloidal iron staining? Secondly, how 
can I convince people that  there is no reason to fear commercial 
preparations of formaldehyde (especially since no one in my department 
does EM anymore)?

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