Hi All,
This is an interesting discussion that has relevance to all of our microscopy. I have a couple of comments:
1. I have always used formaldehyde fixative freshly prepared from paraformaldehyde on the same day it was needed.
2. When I was tempted to freeze aliquots to save time later on, upon thawing the aliquot I regularly saw small amounts of white precipitate on the wall of the container – the longer the storage, the worse the precipitate became. I interpreted this to be repolymerised formaldehyde (newly formed paraformaldehyde). While I have not checked, the chemistry of this repolymerisation process suggests that the pH of the solution should decrease – something that would not be good for fixation.
3. If commercial formaldehyde solution (37% formaldehyde gas in water) needs to have methanol to prevent polymerisation, why do we think that freshly prepared formaldehyde (from paraformaldehyde) should be any different? Surely this suggest that fresh formaldehyde solution is unstable? Maybe freezing makes this even worse?
4. In my older EM days, all of our fixatives were stored in a different fridge/freezer to where our antibodies were stored. As was explained to me at the time and as Pippa Hawes has pointed out, the activity of the antibodies could be affected by fixative vapours.
5. As many people have already suggested, maybe, with super-resolution microscopy, artefacts from good or bad fixation will start to become more obvious.
Just my thoughts/recollections…
Paul Rigby
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Martin Wessendorf
Sent: Tuesday, 20 September 2016 10:43 PM
To: [hidden email]
Subject: Re: aliquoting 4% formaldehyde
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Dear Doug--
My understanding is that if you cool solutions containing formaldehyde gas, you accelerate its re-conversion back to paraformaldehyde.
However, I don't have a reference for this at my fingertips.
--Did the solution freeze solid? If so, the solubility of FA gas might've decreased when the phase changed and you might've lost some, but that's a guess.
Good luck!
Martin Wessendorf
On 9/20/2016 9:34 AM, Cromey, Douglas W - (dcromey) wrote:
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I was talking with a student yesterday about their cells which did not look particularly good under the microscope. The discussion eventually came around to fixation. The student told me that he made up some 4% formaldehyde fixative and then froze back (20 deg C) aliquots for later use. This was something he had seem on an online research forum. In my fairly lengthy experience I have never heard of this being done and I am suspicious that this is NOT a good idea. Thoughts?
Doug
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