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

[Itai Moshe]

Thank's,

Did you used it for mouse liver section ?
Can you please post a your fixation protocol ?

2011/7/7 Edwards, Richard E. <ree3 <@t> leicester.ac.uk>

>  I  have no  experience of using  Bouin’s fixed tissue  for  immunos, tho’
> it  is an excellent fixative for many tinctorial procedures providing the
> fixation time is carefully  controlled…I do not think that  the  methanol in
> the  formalin could affect  anything adversely, also  10% neutral buffered
> formalin is the  fixative  of choice  for  most  labs, I  have never  had
> a  problem with it.****
>
> ** **
>
> *From:* Itai Moshe [mailto:itai.moshe <@t> mail.huji.ac.il]
> *Sent:* 07 July 2011 10:10
> *To:* Edwards, Richard E.; histonet <@t> lists.utsouthwestern.edu
> *Subject:* Re: [Histonet] Leaving sample over night in ETOH 70% during
> fixation****
>
> ** **
>
> I had a problem with liver fixation with PFA, most of the time I've got no
> signal, so I've switched to Bouin's, and got good resalts, although it is
> harder to cut bouin's sections with microtom.****
>
> ** **
>
> Could it be because that I've switched from Paraformaldehyde powder to
> formaldehyde solution (37%) (that contains methanol as a preservative) ?**
> **
>
> I have one good PFA  powder experiment that I've done, and
> some excellent old experiments that someone else in my lab did many years
> ago, but i don't know how, all worked great with PFA fixation, but i could
> not replicate the that.****
>
> ** **
>
> I've tried to do a Formaldehyde 24Hr fixation R.T for the mouse liver
> samples, but the results was catastrophic and even the DAPI didn't worked.
> ****
>
> ** **
>
> So i thought that maybe that because that the working samples were in 70%
> ETOH over night.****
>
> ** **
>
> ** **
>
> 2011/7/7 Edwards, Richard E. <ree3 <@t> leicester.ac.uk>****
>
> Leaving  tissue overnight in 70% ethanol at 4C should have no detrimental
> effect n the tissues, why 4C overnight with Bouin's??****
>
>
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:
> histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Itai Moshe
> Sent: 07 July 2011 09:22
> To: histonet <@t> lists.utsouthwestern.edu
> Subject: [Histonet] Leaving sample over night in ETOH 70% during fixation
>
> 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****
>
> _______________________________________________
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> http://lists.utsouthwestern.edu/mailman/listinfo/histonet****
>
>
>
> --
> Itai Moshe
> Mark Pines lab
> Institute of Animal Sciences, Volcani Center.
> Dept. of Animal Sciences, School of Veterinary Medicine, The Hebrew
> University of Jerusalem
> Israel****
>


-- 
Itai Moshe
Mark Pines lab
Institute of Animal Sciences, Volcani Center.
Dept. of Animal Sciences, School of Veterinary Medicine, The Hebrew
University of Jerusalem
Israel