[Histonet] Leaving sample over night in ETOH 70% during fixation

[Rene J Buesa]

If you have an overnight step it is better to prolong the formalin fixation instead of leaving the tissues in 70EthOL.
Since formalin will require at least 24 hours to completely bind and 48 to crosslink, if you leave your tissues overnight in 70EthOL the unfixed tissue will be fixed by the alcohol instead and that can cause difficulties in the subsequent tests.
René J.

--- On Thu, 7/7/11, Itai Moshe <itai.moshe <@t> mail.huji.ac.il> wrote:


From: Itai Moshe <itai.moshe <@t> mail.huji.ac.il>
Subject: [Histonet] Leaving sample over night in ETOH 70% during fixation
To: histonet <@t> lists.utsouthwestern.edu
Date: Thursday, July 7, 2011, 4:22 AM


Dear All,
Does leaving the samples (diaphragm, liver) over night in ethanol 70% at 4C
during the fixation process, will be better for the tissue fixation, and
does not harm the sample ?
Does the fixation process should be done straight forward from the first
step to the last one without any over night stops (except from the PFA,
Bouin's step) ?


My fixation protocol is like this:
1) immediately after killing the mouse i'm putting the sections in a
fixation solution that is made from: 10ml formaldehyde 37%+5ml PBSx20+85ml
DDW  - pH 7, Or bouin's solution over night at 4C.
2) ETOH 70%, ETOH 80%, ETOH 96%, ETOH 100% x2 - each for 1Hr at RT
3) Xylen x2 - each for 1Hr at RT.
4) Paraffin x3 - each at 60C for 1Hr.

Thank you all very much in advance

Itai M
Doe
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