Does formaldehyde crosslink completely kill cell? - (Feb/05/2008 )
Hi, dear everybody,
I am doing CHIP by standard procedures as formaldehyde crosslink and IP, to analyze the histone modification. However, I have to treat the sample additionally for about 20 minutes, after crosslink. I want to know whether histone modification still occurs in a formaldehyde treated cell. In other words, does formaldehyde crosslink completely kill cell, and lead to inactivation of histone modification enzyme?
Thank you very much.
Chaoyang
when doing histology we always thought of fixation (treatment with formaldehyde/ paraformaldehyde) as instant death.
this meant that all chemical reactions within the cell were frozen at the moment of fixation - the crosslinking acts as an additional imobiliser (as opposed to acetone fixation for example) holding everything in place.
i would say all enzymes would probably be innactivated - enzymes added after fixation (once the fix has been removed) may still do the job intended.
i know its a different setup from yours but i think the rules still apply
dom
I am doing CHIP by standard procedures as formaldehyde crosslink and IP, to analyze the histone modification. However, I have to treat the sample additionally for about 20 minutes, after crosslink. I want to know whether histone modification still occurs in a formaldehyde treated cell. In other words, does formaldehyde crosslink completely kill cell, and lead to inactivation of histone modification enzyme?
Thank you very much.
Chaoyang
I would agree that most enzymatic activity is dead upon fixation but I know that some enzymes still maintain some amount of activity (eg. proteases, phosphatases, RNases). This is probably due to the robust nature of these enzymes but I would guess that histone modifying enzymes would not fall into this category.