Questions of Cryostat Finesse

From:"R.J. Rushmore" <> (by way of Histonet)

      Hello Histonet folks,   I cut fixed frozen coronal sections of
brain.  I cut at -20, warm on a slide warmer with nice looking  results. 
However, at a certain level, I seem to have problems.  At  this level, the
thalamus is separated from cortical grey matter by a substantial  amount of
white matter.  Cutting a section results in a differential  expansion - the
grey matter on either side of the white matter expands more than  the white
matter itself, so ruffles of tissue form.  I use subbed  coverslips to
collect these sections, and upon collection, the ruffles of tissue  melt to
the slide with bubbles underneath the tissue on either side of the white
matter.  At the very least, there are notable folds.  In order to get
around this, I have altered the following variables:  knife angle, roll bar
angle, chamber temperature, velocity of cut and more.  I have also tried
pressing the coverslips onto the sections, delicately attempting to paint
the  ruffles away, warming coverslips, using slides, warming the collected
sections  at a slower rate than on the slide warmer.  I have even lightly
exhaled on  the section to give it more form, as a web site recommends. 
All to no  avail.     If you have any hints, or have faced this  problem
before, could you please let me know.   Yours Sincerely,   RJ Rushmore
Department of Anatomy and Neurobiology Boston University

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