[3DEM] Gatan cryopluge

[Ulrich Gohlke]

Dear Takashi,

Although I haven't used the Vitrobot myself, I understand that the 
gradient of ice thickness is actually an advantage. It means that even 
if if the blotting time is not the same in each case (if done manually), 
there is always an area on the grid which has just the right tickness. 
Exactly as you described it, too thick or too thin at either end but 
optimal somewhere in between. The disadvantage is that you might need 
several grids in order to be able to collect enough data, but if the 
grid preparation is automated, what's the problem? Several grids also 
means that each of them spends less time in the microscope, so (perhaps) 
there is less  contamination.
The slope of the ice gradient is also determined by the time between 
blotting and freezing, i.e. the longer you wait the thicker the ice at 
the bottom of the grid. If this time can be adjusted for the Vitrobot, 
you might get a wider usable area for your grids.

Regards,

 Uli

***********************************************************************

Dr. Ulrich Gohlke
Birkbeck College
School of Crystallography
Malet Street
London WC1E 7HX
United Kingdom
Tel.: +44(0)207 631 68 09       Email: ubcg16e at mail.cryst.bbk.ac.uk
Fax.: +44 (0)207 631 68 03     Web: http://www.cryst.bbk.ac.uk/~ubcg16e


Ishikawa, Takashi wrote:

> Dear Oscar, Bill, Norm, members,
>
>  
>
> We are also happy with the FEI Vitrobot, basically. Students without any
>
> experience of cryo can make a good freezing on the first day. However, we
>
> realized one problem: Frozen grids have gradient of thickness quite 
> often. In
>
> other words, ice is too thick at one end and the grid is totally dried 
> at the
>
> other end, while the area (a few to 10 meshes) in between has optimum
>
> thickness. I guess this is because the grid is blotted by tilted 
> filter paper
>
> attached on pads. Undoubtedly, It would be nicer if we have even 
> thickness
>
> of ice.
>
>  
>
> Did anybody have the same problem as we have and solve this issue ? Or
>
> anybody has the idea how this problem happened ?
>
>  
>
> Regards,
>
> Takashi
>
> Takashi Ishikawa
> Research Scientist / Group Leader
> Department of Biology
> Swiss Federal Institute of Technology (ETH Zürich)
> ETH Hönggerberg, HPK F7
> CH-8093 Zürich, Switzerland
> phone: +41 1-633-2462
> Fax:  : +41 1-633-1073
> e-mail: takashi.ishikawa at mol.biol.ethz.ch
>
>  
>