[3dem] Insertion of Holder 626

[Wim Hagen]

Dear Ariel,

In order of priority:

1) Have your airlock pumping and holder O-ring checked/cleaned by customer service.

2) When you bake the holder, it uses 750 mA current to get to 100° C and then slowly decreases this current to maintain the 100° C. As the holder Dewar vacuum gets better, less current is needed, which makes it a good indicator of bake-out ‘quality’. We aim for <400 mA.

3) Frozen ethane evaporates in vacuum, the more ethane is frozen on the grid, the more likely it will not evaporate fully during airlock cycle, which then leads to a vacuum crash when going into the column as the column’s ion getter pumps definitely don’t like Ethane or Propane/Ethane mix.

Best,

Wim Hagen
EMBL Heidelberg 


> On 24. Feb 2020, at 14:13, TALAVERA PEREZ Ariel <Ariel.Talavera at ulb.be> wrote:
> 
> Dear all,
> 
> I am using a Gatan 626 holder on a side entry TALOS microscope. I am 
> getting problems with very frequent crystalline ice formation on the 
> grids. Some times I get a mix of vitreous and crystalline water, but 
> must of the time I only get crystalline water. I have been extra careful 
> during sample preparation, and transferring the grind on to the holder, 
> and I have done 48 hours holder bake out before insertion, but I still 
> get ice on the grid.
> 
> I monitored the temperature of the holder during the insertion process, 
> including 3 minutes for the Airlock pump. During those 3 minutes the 
> temperature rises up to -152.2. Can this be the reason of the unwanted 
> ice formation? How high can be the temperature of the holder to ensure a 
> nice vitreous water?
> 
> I am also getting vacuum problems while inserting the holder. This 
> process has resulted to be extremely, extremely sensitive. After the 3 
> minutes Airlock pumping (I cannot give longer time because of the 
> aforementioned temperature issue) as soon as I open the column port the 
> vacuum either rises to 99 Log or completely crashes the column vacuum. 
> This has happened with two different holders either cooled with liquid 
> nitrogen or at room temperature. So, my question: Is this insertion 
> process always that  picky or it can be that there is something wrong 
> with the holders/Airlock pump?
> 
> Thanks a lot in advanced.
> 
> Best regards.
> 
> Ariel
> 
> -- 
> Ariel TALAVERA PEREZ, PhD
> Center for Microscopy and Molecular Imaging (CMMI)
> Laboratory of Microscopy
> Université Libre de Bruxelles
> Rue Adrienne Bolland, 8
> B-6041 Gosselies, Belgium
> Phone +32 (0)26509866
> 
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