[3dem] Regarding Image Quality from FEI Tecnai G2 Polara

Wim Hagen hagen at embl.de
Tue Sep 6 11:41:47 PDT 2016


Hi Sayan,

Log 6 column vacuum does not rule out a vacuum leak as vacuum pumping means balancing the always present leakage vs pumping and the Polara is a massive cryo-pump. 
Al Polara's are shipped with a mass-spectrometer measuring head in the column vacuum, so ask your engineer to do a helium leak test.
I demand this after each service action as good vacuum levels do not rule out vacuum leaks!
Massive amounts of ice on the cryo parts causes all kinds of charging problems. I've seen this by keeping a Polara cold for ten weeks (just to see how far one can go...).

Could also additionally be a temperature problem when e.g. the objective aperture holder touches the cryo-box, also have this checked by service engineer.

I've also had issues here from users who did not clip the grids properly. Without the clip-ring fully pushing the grid into the cartridge, contamination was horrible after just one night. But that was clearly visible in the image, not just in the power spectra.

Sounds to me you need FEI over to do some massive cleanup/checkup of your system. 
What's the D-number? Just want to know if I build this one myself :-)

Best,

Wim Hagen
EMBL Heidelberg

> On Sep 6, 2016, at 19:35, Sayan Bhakta <19sayanbhakta89 at gmail.com> wrote:
> 
> Hi Bob,
> Thanks for your suggestion. I would surely convey the message to the FEI engineer.
> Apart from this, we also have another issue and that is constant ice formation inside the column. The engineer who has come, kept a grid inside the column in the weekend and on Monday there was ice contamination formed onto the grid. After that when we inspect, we see continuous decrease in quality of images taken subsequently on the first day itself. This might be because of the fact that the ice starts growing from the day 1 itself. We don't see it on the screen because of the beam may be??? As suggested by many there must be some leakage in the column. But the vacuum of the system is constant at 6. May be that minor leakage is somewhere undetectable. If you share your view on this, It would be highly appreciated.
> Sayan
> 
>> On Tue, Sep 6, 2016 at 8:25 PM, Grassucci, Robert <rg2502 at cumc.columbia.edu> wrote:
>> Hi Sayan,
>> 
>> I believe what you are seeing is terracing or ring collapse.  This happens when the feg tip shape goes from a sharp 9high intensity) tip to a more blunt (dim) tip.  Sometimes you will even see it split and look like 2 sources.  This happens at our facility about once a month so we monitor the intensity very carefully.  The good news is that it generally recovers quickly and then remains stable for a long while.  FEI service engineers know about this phenomenon and have some things to do to make it less frequent.  One is to decrease the tip temperature and the other is to increase the extraction voltage.  You can also reshape the tip by bringing down the Ht and extraction and then bringing it back up  but this does not always work well.  If your tip is damaged then it just may need to be changed. Speak to your engineer so he can correct this.   I hope that helps.  Good luck.
>> 
>> Bob
>> 
>>  
>> 
>>  
>> 
>> From: 3dem [mailto:3dem-bounces at ncmir.ucsd.edu] On Behalf Of Sayan Bhakta
>> Sent: Saturday, September 03, 2016 6:39 AM
>> To: 3dem at ncmir.ucsd.edu
>> Subject: [3dem] Regarding Image Quality from FEI Tecnai G2 Polara
>> 
>>  
>> 
>> Hi,
>> 
>> We are facing many problems with our Cryo EM facility. I am giving some details of the machine and one of the main problems we are facing. It would be very helpful if people comment on this and help us to find the actual problem.
>> 
>> Machine:
>> 
>> FEI Tecnai G2 Polara
>> 
>> 300Kv
>> 
>> Have been running for last two years.
>> 
>> Camera:
>> 
>> FEI Eagle 4k*4K
>> 
>>  
>> 
>> Problem:
>> 
>> On the first day when we are putting our sample into the Cryo EM, we are getting good images. From the second day the quality of the images deteriorates.
>> 
>> 1.  If we look at the images taken on the first day and second day, the intensity values of the images decreased on the second day.
>> 
>> 2.  From the FFT, we see nice thorn rings from the images taken on the first day. But for the images taken in the second day, the FFT is very bad. Most of them are having drifts.
>> 
>> 3. Irrespective of the days, there are some images for which the 1D spectrum is having a cut in between. What I mean by that is there is spectrum upto certain distance in the 1D spectrum, then there is no waves, then again there are some waves.
>> 
>> 4. Irrespective of the days, images are sometimes with blakish shadow on one side and with whitish shadow on other side. We need to do gain correction more frequently than usual- a number of times in a day.
>> 
>>  
>> 
>> For your information:
>> 
>> 1. The vacuum is stable on the second day.
>> 
>> 2. We are giving Cryo-cycle each weekend.
>> 
>> 
>> If anyone has ever faced the same problem or has any advice on this regard, it would be very helpful. We have been struggling very hard to standardise many issues with our microscope.
>> 
>> --
>> 
>> Sayan Bhakta
>> Research Scholar
>> Group of Dr. Jayati Sengupta
>> CSIR - Indian Institute of Chemical Biology
>> Kolkata, West Bengal
>> India
> 
> 
> 
> -- 
> Sayan Bhakta
> Research Scholar
> Group of Dr. Jayati Sengupta
> CSIR - Indian Institute of Chemical Biology
> Kolkata, West Bengal
> India
> _______________________________________________
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> 3dem at ncmir.ucsd.edu
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