Hi,
Iwork with marin cyanobacterial cells (Synechococcus and Prochlorococcus) and I fix samples with glutaraldehyde and keep them in -80c until analysis. On the day of analysis I need to wash out the glutaraldehyde since it interferes with the assay I work with, then I restore the samples in -80c until other analyses. Is glutaraldehyde fixation reversible so I need to re-add glutaraldehyde before the second freezing (in -80c)?
Glutaraldehyde fixation is irreversible because the molecules from a crosslink with the proteins. It is a reaction that needs a lot of energy to be reversed, so you don't need to put your samples again in glutaraldehide.
You could check the following links for more precisions
http://synapses.clm.utexas.edu/lab/howto/cross-linking%20fixatives.pdf
http://www.leicabiosystems.com/pathologyleaders/fixation-and-fixatives-2-factors-influencing-chemical-fixation-formaldehyde-and-glutaraldehyde/
After you fixed your sample (killed everything inside), you don't need to fix it again. Different question is how will you preserve your sample with double freezing. If you don't need to count them than it's ok to freeze the sample again
I need to count the cells again after I thaw them (after the second freezing), so do you think it is not good to subject the cells to two freeze-thaw cycles?
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