[Histonet] Curling floating mouse brains

[Julie Ritchie]

Hello everyone.  

I am slicing mouse brains that were perfused with 4% paraformaldehdye in
PBS, then sunk in 30% sucrose in PBS.  I want to cut the brains in 40 um
sections, collect in wells of PBS, and do IHC.  BUT my sections keep curling
into little cannolis (or even tighter, like those pirouette cookies).  

Just a few weeks ago I sectioned 24 rat brains in this manner without any
problem at all.  I did the perfusions for all of these animals, so it is not
an issue of inadequate perfusion.  

I asked someone if it could be related to size of tissue (mouse vs. rat),
and it was suggested that 30% scucrose was too high for mouse brains, so I
melted them out of frozen OCT (kept in a frig overnight in PBS), resunk
brains in 20% sucrose/PBS and refroze in OCT again.  They did indeed slice
better at this conc of sucrose (no streaks, shredding, etc. when slicing)
but the brains sure did curl still in the PBS!!! I thought the sections were
too thick, so dropped to 30 um, and still there is curling.  Even 20 um
sections curl up in the PBS.  

It is not a humidity problem, or a blade problem, for I have tried to cut on
another machine, and tissue still curled. 

Cutting temp is 13-14 degrees, same as used for rat brains.

Tried different PBS---> same curling.

I am at a loss. 

Any advice would be very helpful. 

Thank you.