Cryostat help!

Paul L Pearson rambis at iastate.edu
Thu May 5 10:56:02 EST 1994


In <1994May4.224609.25276 at ohsu.edu> nishir at ohsu.edu writes:

>In article <pataky-270494154832 at steevlab.generes.ca> pataky at bdc.ubc.ca (Dave
>Pataky) writes:
>>Anybody out there know why Tissue-Tek, presumably designed for embedding
>>samples for cryostat cutting, is so $##@***%* annoying to work with?  The
>>problem:  I'm cutting chick embryonic brain tissue, anywhere from 10-40
>>microns, at -20 and no matter what I try the sections won't stop curling
>>up, rolling into tubes as soon as I lift the antiroll plate.  I've tried
>>adjusting the blade angle, the "anti-roll" (that's a joke) plate, the
>>temperature, speed of cutting, new blade (disposable), nothing works. 
>>Ideally I'd like to see nice flat sections that stay that way, preferably
>>"ribboning" off the blade so I can mount several at once.  Know any tricks
>>or alternatives to Tissue Tek which may work better?  Any ideas how to deal
>>with static electricity? (sometimes the sections stick to the "anti-roll"
>>plate).  Feels like I'm battling the antichrist (and losing!),
>>
>>
>>-- 
>>Dave Pataky
>>Dept of Zoology, UBC
>>pataky at bdc.ubc.ca
>>
>I don't think your problems are with the TissueTek. When I've had problems with
>rolling I can usually fix it by adjusting the angle of the anti-roll plate, or
>by slightly warming the sample by quickly pressing my thumb against it before
>cutting.  It also helps to keep the cover closed as much as possible so that
>the knife doesn't warm up too much relative to the sample. One other
>suggestion: minimize the amount of Tissue tek around the sample.  Embed in as
>small a drop as possible so that most of what you're cutting is tissue and not
>big globs of Tissue Tek.

>Rae Nishi
>Cell Biology & Anatomy
>Oregon Health Sciences University
>Portland, OR 

We use Lipshaw M-1 embedding matrix instead of Tissue-Tek.  It seems to have
less effect on the section with regard to rolling.  As in the above comment,
use as little around the block as possible.  Try cutting at a lower
temperature as well.  I'm cutting embryonic pig heads at -24 or so and getting
pretty good results.  Other days, the weather conditions aren't favorable and
you can't get a section to come off well.  Constatnly adjusting the anti-roll
guide is a must, as well as making sure the guard is at the same temp as the
stage.  YOu might have to change religions or develop new ritual incantations
in some cases, but these suggestions might help.

Paul L. Pearson, 

.x,

-- 



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