[Histonet] Re:sectioning problems

[Gayle Callis]

First and foremost, if you are having the ribbon pulling up as chuck with 
block (Point #3) you need to make sure  knife angle setting on the 
microtome is correct.  Check your manufacturers instructions on HOW to set 
a correct knife angle for your particular microtome.  it sound as though 
the clearance angle is incorrect.  I don't think the paraffin is the 
problem, see last comments.

You did not say what tissues you are sectioning animal?  human? mouse? nor 
how the tissue is processed?  If the tissues are dehydrated, cleared and 
infiltrated with paraffin - optimally for your particular tissue? then you 
should experience good sectioning with correct microtome settings.  Beware 
of infiltrating tissues with excessively hot paraffin, 60C is all that we 
allow with all your animal tisues, over that - I think and with long 
infiltration times, you dry out and create some sectioning problems.

64C is pretty hot in my estimation for embedding, with ANY paraffin.  We 
set ours at 60C only.  You have hot and cold surfaces on your embedding 
center to help you out, these are usually preset at factory although we can 
change that with our Sakura embedding center.  Too hot is an enemy, as you 
cook the tissues even more.  Even the kind of blades you use will affect 
how your sectioning goes, and we all have our favorites - be sure to try 
others on the market!!!

Dirrerent brand paraffins will have different sectioning qualities 
depending on what additives they add to make sectioning easier, etc.  Some 
paraffins are sticky, others are harder, some you have more compression, 
but with care and correct knife angle setting, one can usually learn to 
work with ANY paraffin.   These additives will cause the paraffins to look 
different too - opacity, clarity, hardness, ribboning ability, tackiness as 
you have noticed.

Although I like flowers, I favor the brownies!!!





At 11:20 PM 9/22/2005, you wrote:
>I recently have been having problems pulling ribbons from my microtome and 
>was
>wondering if anyone has advice.
>
>1.) I was having problems pulling ribbons from newly embeded blocks, but 
>ones that had
>been embeded previously were very easy to pull ribbons from.  The new ones 
>would simply
>curl on the blade and wouldn't even grab on to the adjascent sections.
>
>2.) I changed the paraffin in the embedding station because I noticed all 
>the temeratures
>were set to 64 C when the melting point for the Surgipath paraffin formula 
>'R' was 56-67C.
>Could this have been a problem?
>
>3.) I switched to an old lot of Fisher embeding medium and now all the 
>section come off, but
>on occasion when the chuck on the mictomome comes back up it seems to (via 
>static
>electricity?) pull the ribbon from the microtome blade and interupt the 
>ribbon.
>
>4.) Also, visually the Fisher paraffin blocks look almost transparent 
>whereas the Surgipath
>blocks seem opaque and white.
>
>I'm relatively new to the histology game and I realize that there is both 
>art and science
>involved, but this is clearly unacceptable behavior from the paraffin, 
>something is surely
>wrong.  I'm sure you are all experienced and maybe you have advice?  If 
>you do I would
>apreciate it so much.  The first and best advice get their choice of FTD 
>flowers and/ or
>homemade brownies.
>
>Thank you--Tyler Wellington
>
>
>
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Gayle Callis
Research Histopathology Supervisor
Veterinary Molecular Biology
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
406 994-6367
406 994-4303 (FAX)