[Histonet] Mouse Tissue Processing Problems Help

From:"Donna Harclerode"

My first suggestion is turn off the heat in your processor.  I
personally do not like tissues stored in 70% alcohol but others think it
is great. Having heat on in 70% alcohol and your other solvents would be
very drying to most tissue.  Clinical labs use heat because they are
pressed for time (not optimal, but it works). I recommend never having
heat on a processor with animal tissue. I also suggest using a xylene
substitute and not xylene.  I like the aliphatic hydrocarbon substitutes
but the limonene (orange smell that some people get sensitized to) also
more gentle than xylene.

I use a 1 hr schedule for most of my tissues- I use Anatech's ProPar
(aliphatic hydrocarcarbon xylene substitute) and Prosoft (alcohol
combination) on a VIP Processor (with pressure and vacuum but no heat).
All my tissues are well fixed before I put them on the machine.  I got
into the habit of washing tissues when I was using the Zinc formalin and
regular formalin tissues to keep the phosphate crystals out of the
tissues. It also keeps the formalin out of my alcohols and keeps the
processor lines cleaner.  Anatech site is
http://www.anatechltdusa.com/index.html  They make great products and
have the best technical support!  Graded alcohols will work (I have used
ethanol only for mice and isopropyl for rats, dogs and monkeys in the
past), but I find the ProSoft to be easier and not as drying as any of
the plain alcohols.  I do not make dilutions of ProSoft, but just rotate
the containers per Anatech recommendations.  

I still face off all my blocks and put them back on ice to rehydrate
them a bit before I section them.  When I do chrondocyte pellets, I drop
the time in each station to 20 minutes since they are so tiny.

I currently use the Surgipath embedding and processing paraffin but will
be switching to Richard Allan Type 3 for processing and Type 9 for
embedding from VWR or elsewhere. 


1.	NBF                  hold till process
2.	tap water           30 min
3.	ProSoft             1hr
4.	ProSoft             1 hr
5.	ProSoft             1 hr
6.	ProSoft             1 hr
7.	ProPar              1 hr
8.	ProPar              1 hr
9.	ProPar              1 hr
10.	Wax                 1hr
11.	Wax                 1 hr
12.	Wax                 1 hr


Good luck


Donna Harclerode, HT, (ASCP), HTL, QIHC


Cytori Therapeutics

6740 Top Gun St.

San Diego, CA 92121

858-458-0900 ext 322



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