[Histonet] frozen section - spleen problems

[Gaupp, Dina D]

Histoland:
 
I am having problems sectioning spleen without artifact(holes) as well as pencil shaves.  A little bit of background.... mouse spleen fresh frozen embedded in OCT (plastic mold) at -80C freezer.  The tissue was then given to me already frozen in OCT mold.  Each specimen consisted of 2 spleens and 2 hearts in one mold.  The hearts cut fine, but the spleens cut like pencil shaves.  The border of the spleen stayed intact, but the entire center would either section with huge gaps in the tissue or it shredded into pieces.   Now the PI(Principle Investigator) wants to know what did I do wrong.  I've tried warming the tissue before sectioning it with my finger as well as increasing the cryostat temp to -18C.  Any warmer & the heart starts to malfunction upon sectioning.  I told the PI not to embed 2-different samples in one block.  I don't know what else to tell him or what else I can do to give him a better sample.  I've tried telling him - its a frozen section of tissue - morphology will not be well preserved, you have higher chances of artifact than paraffin processed tissue.  Again, remind you this tissue was given to me already frozen in OCT.  I wish there was a way we could soften the tissue other than rubbing my finger across it.  Anyone have any tricks up their sleeves to cut spleens perfectly??????
 
Do you think initially when freezing the spleen down, freeze it at a warmer temp. (-40C or -50C) & the tissue will become less brittle upon sectioning?  I've never frozen tissue warmer than -76C.   I know that freezing tissue any warmer than -80C can cause artifact & obstruct cellular/cytoplasmic morphology.....  I've been doing a little background reading on this matter & I've read that some freeze their tissue at -40C or warmer & are able to get a section without artifact......why this can't be meeee.
 
Histology seems to be a field of trial & error... what works for you might not work for someone else.  I am glad we have histonet, because we can gather everyones do's & don'ts & come to our on conclusions.
 
Thanking you all in advance for your advice....
Dina
 
Dina D. Gaupp, BS, MT
Histology Core Lab Supervisor
Center for Gene Therapy, SL-99
Tulane University Health Science Center
1430 Tulane Ave
New Orleans, La 70112
Lab: 504-988-1194
dgaupp <@t> tulane.edu <mailto:dgaupp <@t> tulane.edu>