[3dem] Gatan 914 cryotransfer holder summary

[Frank Polzer]

Dear all,

 

Thanks a lot for your great support. To make sure everyone in the mailing
list has access all your good advice, I will copy all your answers into this
email. I hope that is ok for everyone.

 

I think Valorie was right with the worn out O-ring since I observe a lot of
ice on the outside part of the shaft. 

 

I will try to replace it and see if I can get rid of that problem.

 

Thanks again for all your help!

 

Cheers,

Frank

 

 

 

I think this is because of lN2 getting into the neck of the transfer
station, so the o-rings freeze. If you find it is stuck you can sometimes
free it up by holding the neck with your hand to warm it up, but of course
it is better not to freeze the o-rings in the first place.  

Generally I cool our holders in the column and then just cool the transfer
station a minute before doing the transfer, you need to make sure that the
plug is in the neck in this case so that you dont get lN2 in there, which
would cause the same problem....

 

D.

 

Dear Frank,

 

We do a couple of things to try and minimize this happening. First, we cool
the holder in the microscope (less frost on the rod). Second, we tilt up the
back end of the workstation while cooling the work area in order to keep the
liquid nitrogen from reaching the o-ring seal point. We also work in a
humidity controlled room (< ~30% relative humidity). Finally, I sent the
whole system back to Gatan for them to check it over
they were very helpful
and 'shaved' off a little of the workstation rod interior diameter to
minimize the contact points between it and the cryo-holder rod. I am not
sure how many holders/workstations you have, but we generally segregate them
(i.e. matched sets).

 

I hope this information helps.

 

Sincerely,

 

Liz

----

Elizabeth R. Wright, PhD
Assistant Professor
Director, The Robert P. Apkarian Integrated EM Core
Georgia Research Alliance Distinguished Investigator
Emory University
 <mailto:erwrigh at emory.edu> erwrigh at emory.edu
Ph: (404) 727-4665

 

Dear Frank,

 

Probably you will hear the same answer from others, but for us, the only
answer is to work quickly.  

 

For the first loading, we make sure the nitrogen stays below the level of
the rod while the station and the holder get cold.

 

Then we warm the transfer station after each loading (put a hot metal
cylinder wrapped in cloth into it and put it into a warm but not hot oven.

To load the holder cold from the scope, we fill the transfer station with
LN2, wait only for it to stop boiling, keep the nitrogen below the rod
level, and do the transfers within a few minutes.  If you let the holder sit
in the station too long, it will get stuck.

 

good luck,

Esther

 

Hi Frank. This can be associated with several issues. The simplest is an old
dry o-ring. try to replace it. Otherwise it may be the holder is slightly
bent or there is a leak in the transfer station. The latter you can
recognize if frost builds around the tube. 

A control to isolate the problem is to test performance using another holder





Good luck

 

Dear Frank,

We had this problem at Purdue as well and Paul was able to figure it out.

Is the body of the workstation frosting up where it joins the shaft?  If so
it, the O ring has worn out.  There is an O ring at the joint between the
shaft and the cup.  Replace it and you should be back to normal.

 

Cheers,

 

Valorie

 

 

Valorie Bowman

Senior Research Electron Microscopist,

 

Dept. of Biology

Purdue Univ.

W. Lafayette, In 

    47906

 

 

ph. 765-494-5643

fax 765-496-1189

vdb at purdue.edu

 

Dear Frank,

 

Have you tried cooling the work station and the cryoholder separately? You
can cool the holder in the microscope.

 

Cheers

Benoit

 

Hi Frank.
I have a 914 holder, and it has gotten stuck as well. The most likely reason
is that there are still traces of water, even though you think you’ve gotten
rid of it. I tend to really blast the transfer station with the “hair dryer”
for a LONG TIME, and then its okay.
The other reason may be that the transfer station got a “bump” and the shaft
is not really straight (i.e., horizontal). 

Hope this helps.

Sharon

 

 

Hi Frank,
I have used cryoem for several years and change grids hundreds of times,
never had such problem, but several person in our lab were stuck many times.
I think the difference is that most people put in too much LN2 in the
station, which flow to the junction and freeze the holder. I never put in
LN2 higher than the grid level. 

When I first use cryoEM, someone suggests always keep grid under LN2. In
fact, after cool down to -160 degree, the air inside the station is cold
enough. Grid would move if the LN2 is higher, hardly kept it in the center. 
Best,
Jim

 

 

 

 

 

 

Dr. Frank Polzer 
TEM Group 
Institute of Physics 
Humboldt Universität zu Berlin 
Newtonstraße 15 
12489 - Berlin 
  
Tel.: +49 30 2093-4995 (office) 
Tel.: +49 30 2093-7829 (TEM) 
Fax: +49 30 2093-7760 
  
 <mailto:frank.polzer at physik.hu-berlin.de> frank.polzer at physik.hu-berlin.de

 

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