[Histonet] RE: Inconsistent Sections with Cryostat

[Lee & Peggy Wenk]

I just attended Jan Minshew's workshop on cryostats at the NSH Symposium in 
Providence, RI, and she brought up something I had never thought of that 
causes thick and thin.

If the handle that tightens the blade in the blade holder is over-tightened, 
the blade will become bowed, and that will cause thick-and-thin during 
sectioning.

According to Jan, the handle should be tightened to be parallel to the angle 
of the blade holder "slope". A lot of times, the handle can be pushed 
further back towards the back of the cryostat, beyond being parallel with 
the slope. The thought seems to be, if tight is good, then tighter is 
better, but not in this case.

Peggy A. Wenk, HTL(ASCP)SLS

-----Original Message----- 
From: Manfre, Philip
Sent: Thursday, October 03, 2013 7:54 AM
To: Perow, Elliott S (PEROWES10) ; histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] RE: Inconsistent Sections with Cryostat

I believe you may need to have the unit serviced.  It sounds like something 
is not tight enough, perhaps the stage or blade holder unit.  You said you 
secured everything which makes me think you have some issue with the 
cryostat itself.  If a sharp blade, tightened blade and specimen, and 
varying the speed doesn't work, then you may need professional assistance 
form a service technician, especially if it is an older unit that has been 
moved around.  Have you tried different temperatures?  I don't think that is 
the problem but it is worth a try.

Good luck! Cryosectioning can be an art in itself. Those decent sections are 
likely thicker than you intend, by the way.  Classic "thick and thin".


Philip Manfre, B.A., HT (ASCP)
Associate Principal Scientist
Merck Research Laboratories
WP45-251
PO Box 4
West Point, PA 19486

215-652-9750
215-993-0383 (fax)
philip_manfre <@t> merck.com




-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu 
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Perow, 
Elliott S (PEROWES10)
Sent: Thursday, October 03, 2013 12:33 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Inconsistent Sections with Cryostat

Hello All,

I have a Leica Cryocut 1800 with a Reichert-Jung 2020 microtome that has sat 
for 8 years and went through a move from one academic building to another. 
I am trying to section brook trout brain, but am having difficulty getting 
consistent sections.
I will get a good section and then the next section will be a very very thin 
shaving of usually just the top portion of the OCT block.  I have tried 
adjusting the clearance angle, made sure the blade and specimen is secured, 
replaced the blade with a new one, have tried different sectioning speeds, 
different thicknesses of the section and still continually only get a decent 
section every other turn.  I was wondering if anyone has had any similar 
experiences or if maybe the machine just needs a maintenance check due to it 
being an older machine.  I wasn't sure if the advancement mechanism may be 
off or if it is another issue that doesn't seem apparent to me.  Any help 
would be greatly appreciated.

Thanks,

Elliott Perow
Juniata College
Biology POE


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