[3dem] Thinner ice after several microscope uses

[Chipman, Paul R]

Yes it is important to cool the trap prior to inserting the cold holder,
though I've never tried to quantitate the contamination.  We cool the
cryo-blades at least 30 minutes prior to inserting the cryo-holder.  We
wait an additional 45 minutes before opening the shield on the
cryo-holder.  The cryo-cycle is run every time the cryo-holder is used;
almost every night.  I've not seen any correlation between ice thickness
and insertion number.

Cheers,
Paul

Eduardo Sanz García wrote:
> Henning Stahlberg wrote:
>> Hi,
>>
>> We had a similar phenomenon: Ice contamination on cryo-EM grids was 
>> sometimes but not always quite bad, but the second inserted grid was 
>> often less contaminated than the first one. This was on a JEOL 
>> JEM-2100F.
>> I believe the problem might have been related to some weird 
>> air-pocked formation in the goniometer, potentially between two 
>> consecutive O-ring sealings. When we left the goniometer without 
>> sample holder over night, air might have slowly accumulated somewhere 
>> and was inserted into the column during the first sample transfer. We 
>> tried leaving a holder in the goniometer over night, but that did not 
>> significantly improve the situation, as if the "leak" only occurred 
>> during sample insertion.
>
> I was thinking in the same lines than you, a problem with the 
> goniometer-airlock, but yesterday, I had another idea of what could be 
> wrong.
>
> If for some reasons the cool trap takes a lot of time to cool off, 
> like  1 or 2 hours, the cryoholder could function like a cool trap 
> building contamination in my grid. On the other hand, if the 
> microscope have been working all day long, then all the contaminant 
> have been trapped during the day, then I will not have problems of 
> contamination.
>
> Two questions for everybody:
>
> * How long do you let the cool trap to cool off before inserting the 
> cryoholder?
> * Is it necessary to do a cryocycle every night?
>
> Thank you very much.
> _______________________________________________
> 3dem mailing list
> 3dem at ncmir.ucsd.edu
> https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem


-- 
Paul Chipman
Director, Biological Electron Microscopy Facility
Purdue University
765-494-1487