[Histonet] Help! Gelatin embedded mouse brains cracking!

From:"Joshua Berman"

I've been using 10% gelatin/1xPBS to embed mouse brains prior to quick 
freezing in a cold isopentane bath.  I like the gelatin embedding because I 
can see the orientation of the brain in the block and trim it to optimize 

The only problem I've had is that sometimes (with varying frequency) the 
block (and sometimes the brain with it) cracks in the freezing bath.

I have isopentane in a metal container surrounded by a dry ice methanol 
bath.  I put one piece of dry ice in the isopentane.
The 4% para perfused mouse brains are infiltrated with 25%, then 30% sucrose 
after postfixing in 4% para. The gelatin does not contain any sucrose and 
was not placed in para.  The brain is placed in 37*C geatin in a cryomold, 
oriented, and then goes into the refrigerator for the gelatin to harden 
prior to trimming the block and freezing it.

Is this temperature control? Differential rates of expansion?  Do I need to 
add something to the gelatin or use more or less of it?
Any suggestions?

Thanks in advance.

Joshua Berman
Columbia University 

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