I agree with Andi as I have also worked for tissue processor manufacturers
and assisted in designing two.  The heat and vacuum are often the biggest
problem and should be carefully used.  The placement of the paraffin ovens
can also generate heat into the process tank as the heat will rise to the
tank.  Thus what you think is ambient may be as high as 40 degrees C for all
reagents.  The addition of even more heat and vacuum will only cause more
damage.
Tony and Barry have the same thoughts I have on over fixation, over
dehydration and over clearing, it doesn't happen.  Under fixation with the
addition of heat to the alcohols just changes the way the proteins and
tissue are fixed leaving different artifacts of fixation.  Pam Marcum

> -----Original Message-----
> From: Andrea Grantham [mailto:algranth@u.arizona.edu]
> Sent: Thursday, January 17, 2002 10:43 AM
> To: histonet@pathology.swmed.edu
> Subject: Re: mouse processing my current schedule
>
>
> Renee,
> I process a lot of rodent tissue in my lab and don't have a problem with
> over processing. I have an old processor that doesn't use vacuum or heat
> during the processing and only some gentle swirling for agitation (Tissue
> Tek dip 'n dunker). This seems to be perfect for all the animal
> tissue that
> I process. My usual schedule is one hr in all stations - 2-70%
> ETOH, 1-80%
> ETOH, 2-95% ETOH, 3-100% ETOH, 2-Xylene, 2-Paraffin. I usually let the
> tissue set for a little longer in the last paraffin to make sure that it
> gets infiltrated well.
> When I worked for a company that manufactured enclosed tissue
> processors we
> often had complaints that the tissue was coming out looking over
> processed.
> If they turned off the vacuum in some or most of the stations they had
> better results. Also if you have variable agitation - like a stirring
> mechanism that you can program for different speeds you might
> want to turn
> it down a notch or two.
> Hope this helps.
> Andi
>
>
>
>
>
> At 08:19 AM 1/17/02 -0500, you wrote:
>
> >Some people have asked to see my processing schedule  to help
> determine if
> >in fact our mouse tissue is being over processed.  After fixing
> for 48hrs
> >and then being held in 70% ethanol until processing(could be a few
> >weeks)  mouse tissues (about 3mm in size ) are processed by the
> following
> >schedule:
> >
> >70% ethanol              15min             Ambient
> >temperature          Vacuum     *all solution are also
> >80% ethanol              15 min            Ambient
> >temperature          Vacuum       stirring.
> >95% ethanol              20 min            Ambient temperature
>        Vacuum
> >95% ethanol              30 min            Ambient temperature
>        Vacuum
> >absolute ethanol        20 min            Ambient temperature
>       Vacuum
> >absolute ethanol        30 min            Ambient temperature
>       Vacuum
> >Xylene                      30 min            Ambient
> >temperature          Vacuum
> >Xylene                      30min             Ambient
> >temperature          Vacuum
> >Paraplast                 45 min            Ambient
> >temperature          Vacuum
> >Paraplast                 45min             Ambient
> >temperature          Vacuum
> >Paraplast                 45min             Ambient
> >temperature          Vacuum
> >
> >
> >Thanks,
> >Renee
> >
> >
> >Renee Hoyle-Thacker
> >CIIT Centers for Health Research
> >6 Davis Dr.
> >Research Triangle Park NC 27709
> >Voice: (919) 558-1322
> >
> >
> >
>
> .....................................................................
> : Andrea Grantham, HT(ASCP)     Dept. of Cell Biology & Anatomy     :
> : Sr. Research Specialist       University of Arizona               :
> : (office:  AHSC 4212)          P.O. Box 245044                     :
> : (voice:  520-626-4415)        Tucson, AZ  85724-5044    USA       :
> : (FAX:  520-626-2097)          (email:  algranth@u.arizona.edu)       :
> :...................................................................:
>            http://www.cba.arizona.edu/histology-lab.html
>
>
>