Re: Fixation time for mouse skin

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From:amos brooks <>
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    Wow! "dipped in paraffin"? I think that's the problem. The infiltration should
be the longest part of the process (next to fixation of course). The paraffin needs
to replace the xylene in the tissue. (hence infiltration) Even if you happen to get
a section of a poorly infiltrated tissue, it will not stain well. After the section
is taken it is desecrated (xylene) and rehydrated (alcohols: 100% to water). If the
xylene has not been cleared out of the tissue with paraffin, your stain will show
the effects of the mixing of xylene and water.
Amos Brooks wrote:

> I am asking this question for a graduate student here in our lab...Some mouse
> skin was recently processed and when trying to cut the sections it was found
> that the tissue was too brittle.  The skins were fixed in 10% nbf for 16 hours
> and then dehydrated in a series of ethanol dilutions(70%, 85%, 90% and 100%) for
> 20 minutes each.  Skin was then cleared in xylene for 20minutes, dipped in
> parrafin and then embedded.  We think that perhaps the skin was fixed for too
> long a period.  Does anyone have any suggestions regading times for the
> fixation, clearing or parrafin infiltraion?
> I realize that this question has probably been asked many times before, but I am
> relatively new to the field of histology and am still learning where to find
> good accurate information.
> Thanking you in advance,
> I remain sincerely yours,
> Christine Klein
> MD Anderson Cancer Center Houston Texas

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