[3dem] Gatan 914 Holder Woes

[Radostin Danev]

Dear Cindi,

I could not see the pictures (for some reason could not connect to the FTP
server) but from what you describe I am observing a very similar effect with
our 914. The images look similar to what a "warmed-up" (re-crystallized) ice
looks like (cauliflower?). So at first I thought that the specimen does not
stay cool enough during the transfer. I tried cooling the holder to below
-190degC by filling up the cryo-station with liquid nitrogen to above the
holder tip level, but this does not seem to help.
I think Bob may be on the right track suggesting that there is a problem
with the cryo-shield. If you look carefully at the tip of the holder you
will notice that there is an open space between the cryo-shield (shutter)
and the holder even when the shutter is closed. This opening may be the
culprit.
My tomographic reconstructions show that the roughness (cauliflower) appears
only on one side of the ice. This further suggests that the effect is
probably due to poor cryo-shielding.

Could you please forward any tips/suggestions that you may get off the list?

Best regards,

    Rado


=====================================
Radostin Danev, PhD
Okazaki Institute for Integrative Bioscience
National Institutes of Natural Sciences (NINS)
5-1 Higashiyama, Myodaiji-cho
Okazaki, Aichi, 444-8787, Japan
E-mail：rado at nips.ac.jp
TEL：+81-564-59-5563
FAX：+81-564-59-5564
=====================================







-----Original Message-----
From: 3dem-bounces at ncmir.ucsd.edu [mailto:3dem-bounces at ncmir.ucsd.edu] On
Behalf Of Cindi Schwartz
Sent: Wednesday, October 08, 2008 06:55
To: 3dem at ncmir.ucsd.edu
Subject: [3dem] Gatan 914 Holder Woes

Hello List,

We are having problems with plunge-frozen cryo grids only when using  
our 914 holder. Basically, we are getting what I like to call, the  
cauliflower effect. Sometimes it's very subtle and sometimes it's  
blatant. Here are the facts:

1. This is happening to multiple people with multiple sample types
2. This happens when using the vitrobot or by hand-blotting
3. Grids from the same freezing session viewed with a Gatan 626 holder  
do not have bad ice
4. Temperature measurements of the 914 holder during all phases of  
loading the holder and inserting it into the microscope never get  
above -170C
5. Grids have this regardless of which of 2 tanks of ethane we use

Any ideas of what may be causing this would be greatly appreciated.

The pictures were too big to send to the list (although quite small by  
my standards). They are posted on our ftp site:

ftp://bio3d.colorado.edu/914

Thanks,
Cindi



**********************************
Cindi L. Schwartz
University of Colorado, Boulder
Dept. MCDB
BL3DEMC
O: 303-492-7980
F: 303-735-0770
**********************************

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