[Histonet] Problem with liver fixation

[Geoff McAuliffe]

Formalin fixes slowly, even more slowly at 4 degrees C. Even though the 
fix may penetrate, fixation will not be complete.
I suggest a formalin+alcohol+acetic acid fix at room temp for 24 hours.
There are many, many variations of this fix. Lillie recommends 85 ml of 
95% EtOH, 10 ml conc. formalin (37-40% formaldehyde), 5 ml glacial 
acetic acid.
I agree with Rene, Bouin will not make a difference. Alchoholic Bouin is 
also worth a try.

Geoff

Itai Moshe wrote:
> Dear All,
> I'm trying to use liver sections in paraffin blocks for IHC, but get no
> signal at all, doesn't matter what antibody i'm using.
>
> I'm using section about 3mm thick (the natural thickness of mouse liver) and
> about 5-7mm Width and Length.
>
> 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, 24Hr 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.
>
> Before doing IHC:
> 1) Xylen x2 - each for 10Min at RT
> 2) ETOH 100%x2, ETOH 96%, ETOH 80%, ETOH 70% - each for 2Min at RT.
> 3) IHC protocol...
>
> Will Bouin's solution be better ?
>
> Thank you all very much in advance
>
> Itai M
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>
>   


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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
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mcauliff <@t> umdnj.edu
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