Re: [Histonet] Bouins fixation problem

From:Geoff McAuliffe

Hi Kim:

    Hmm. Certainly sounds like an infiltration problem. My first though 
is that you are not getting all of the alcohol out of the medulla so 
xylene (or substitute) is not getting in so the wax is not getting in.  
When you say that you have checked all of the processing solutions and 
they are fine, how did you check them? What is the criteria for deciding 
that a solution is good? I would dump ALL of  the processing solutions 
and ALL of the wax and start fresh. Run some kidneys, cut them, and see 
if that solves the problem. There is a small chance that the 
manufacturer of your wax has changed the formula but with modern waxes 
and additives such a change seems unlikely to cause problems.
    As for reagents going bad, picric acid last forever, formaldehyde is 
fine as long as there is no ppt. on the bottom of the bottle, I don't 
think glacial acetic acid goes bad. Even if the fixation was incomplete 
(you did not say how long you fix the kidneys, if you perfuse fixative, 
or if you slice them open) the alcohols should finish the fixation.
    Good luck!


Kim O'Sullivan wrote:

>Hi everybody,
>We have used bouin's fixative for fixing mouse kidneys for years with no problem, and have recently been experiencing problems cutting them once they have been processed and wax embedded. The  kidney tissue has holes in it in the medulla region, is very difficult to cut(tissue brittle) and it appears almost like the wax has not infiltrated the kidneys properly. Additonally once the sections have been mounted on glass slides they appear fine but when we go to PAS stain them half of them come off the slides in solution (this has never happened before in the last 3 years)I have checked the processing schedule and that is fine (as is all the solutions) I have checked the temperature of the wax (which is fine). Does anyone out there know what our problem may be?? Additionally is there any way that any of the solutions used to make bouin's (picric acid, formalin and glacial acetic acid)can go off? And how would you check for this.
>Would appreciate any help!!
>Kim O'Sullivan
>Histonet mailing list

Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029

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