[Histonet] fresh-frozen mouse tail sectioning

[Rachel, Rivka (NIH/NEI) [E]]

Do you have to freeze or section the tissue at all?  If there is significant beta-galactosidase activity, you should be able to just put the tail tip in X-gal and see if it turns blue, having of course appropriate positive and negative control tails that are known to carry or not carry, respectively, the relevant transgene.

Rivka


--
Rivka A. Rachel, MD, PhD
Staff Scientist, National Eye Institute
Neurobiology-Neurodegeneration and Repair Laboratory
Tel: 301 443-4906



-----Original Message-----
From: Gaupp, Dina D  [mailto:dgaupp <@t> tulane.edu]
Sent: Fri 5/16/2008 12:04 PM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] fresh-frozen mouse tail sectioning
 
To Whomever This Applies:
 
I am having big problems sectioning mouse tail, approximately 2mm in thickness, vertical embedded in OCT for frozen sections.  The tissue is fresh-frozen because the principle investigator would like to detect an enzyme x-gal.  No fix, no cryopreservation - the tissue was not immersed in any type of solution.  He snipped the ends of a mouse tail & immediately gave me the tissues.  I embedded the tissue vertically in OCT & flash froze at -80C.  Upon sectioning, the tissue rolled.  I could not for the life of me get 1 section.  All the tissues rolled & its so tiny to begin with that it was hard for me to grab it.  I used the anti-roll plate, hoping it would hold the tissue in place but it still rolled.  I changed temperature settings(increase & decrease temp), thickness, angle, rubbed it with my fingers.  Everything I could think of to get a section.  I asked someone else in the lab to try & they couldn't get a section.  It was like the tissue completely separated from OCT.  The principle investigator will give me more samples & I don't want this to happen again.  
 
Can anyone in histoland help me, tell me what I didn't do or what I did wrong?
 
Dina D. Gaupp, BS, MT
Senior 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> 
 
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