[Histonet] Question

[Jun, Kristine]

Topic - Using the vibrating microtome with frozen tissue

Question - Hello everyone. My lab recently purchased a Leica VT1000S with the intent to use it for 200 um sections of brain tissue. I am familiar with vibratome applications for fresh tissues, but my PI would like me to use it strictly for fresh frozen mouse brains (unfixed). I am having great difficulty with even obtaining one good section and the technical service representatives at Leica were unable to help me as well. I ordered injector blades from Ted Pella but, while my order is on its way, I was looking for any tips or suggestions. I keep my frozen brains on ice and fill the buffer tray with 4% PFA in PBS along with some ice around the edges, and keep dry ice around the buffer tray to keep it cold. My brain rapidly defrosts as I glue it to the stage and the moment that it hits the PFA, it starts to get gummy-looking in appearance. I have tried to section at various frequencies and speeds, but the results are still not great. If anyone has any tips or experience with this, please let me know!  Thank you!

Kristine Jun
UCSF Center for Cerebrovascular Research
1001 Potrero Avenue, Box 1371
San Francisco, CA 94110
phone: 415-206-3344
fax:  415-206-8907
junk <@t> anesthesia.ucsf.edu<mailto:junk <@t> anesthesia.ucsf.edu>