[3dem] Gatan 914 Holder Woes

[Dennis C. Winkler]

Dear Cindi and Rado,

We have the same problem here with our 914 holder - what you call  
cauliflower, I was calling cobblestones.  We would see this with the  
914, but not a 626, even when the grids from the same freezing session  
were looked at on the same microscope.

I noticed, as Rado did, the gap between the cryo-shield/shutter and  
the holder.  Suspecting this as the problem, I spoke with Mark Dods at  
Gatan.  At the time, Mark told me that all 914 holders have this gap  
and that there were no reports of users with this problem.

I would also appreciate hearing any information you may get on this.

Thanks,
  - Dennis

-----------------------------
Dennis C. Winkler, PhD.
National Institutes of Health
NIAMS/LSBR
50 South Drive
Bldg. 50, Room 1513, MSC-8025
Bethesda, MD  20892-8025, USA
Phone: (301) 496-0131
Fax:   (301) 480-7629
Email: Dennis_Winkler at nih.gov


On Oct 7, 2008, at 10:01 PM, Radostin Danev wrote:

> 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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