[3dem] EM sample in high salt

[Florian Hauer]

Hi Peijun,

sorry to bother you all again with the GraFix protocol, but it seems to  
help in this case, too. If you run your sample in a GraFix gradient  
containing 1 M salt, you can do a buffer exchange afterwards using a  
desalting column (e.g. Pierce Desalt Spin column, GE PD MiniTrap G-10  
column etc.). During this buffer exchange, you can choose the salt  
conditions you need for EM. The GraFix protocol will additionally help to  
further stabilize your complex. We have lowered the salt conditions for  
different complexes this way in various ranges and it has always worked  
well without compromising the particle itself.

The reference to the GraFix method is: Kastner B et al. GraFix: sample  
preparation for single-particle electron cryomicroscopy. Nat Methods. 2008  
Jan;5(1):53-5. Epub 2007 Dec 23.

Should you have any more detailed question regarding the procedure, do not  
hesitate to contact me personally.

All the best for your experiments,

Florian.

On Tue, 08 Jul 2008 00:15:44 +0200, Peijun Zhang <pez7+ at pitt.edu> wrote:

> Hello all,
>
>
> I have an EM specimen which only stabilizes in high salt buffer (1M  
> NaCl).
> I would like to carry out cryoEM imaging at lower salt conditions.  Does
> anyone have a good protocol to reduce salt to 100mM or less without
> compromising the specimen?
>
>
> Thanks so much,
>
>
> Best,
>
>
> Peijun
>
>



-- 
Florian Hauer

3D Cryo Electron Microscopy Group
Abt. 10710
Max-Planck Institute for biophysical Chemistry
Am Faßberg 11
37077 Göttingen

Phone: +49 551 201 1302
Fax:   +49 551 201 1197