[3dem] Request for help to find source of resolution loss in cryo-EM

[Mike Strauss]

Hi Henning,

We have a similarly equipped system and encountered the following issues
that could lead to something like this:

1. A mild pulsing of the HT, visible at spot 1 in probe mode near
crossover.  This was made worse by the Keithley power supply for our Phase
Plate.  We resolved this by moving the power supply farther from the
column, and plugging it onto the microscope powered outlets.  Since the LS
Quantums are not coupled directly to the HT, this makes the problem even
worse when going through the GIF.
2. Vibration caused by pump (either turbo, or rotary, depending on what was
happening in vacuum-land).
3. Improperly "tuned" GIF.

I would also do a drift test to see if you have systematic motion along Y.
If you have a pre-GIF camera, check if you have a similar attenuation,
thereby possibly excluding the GIF as the source. Check to make sure the
phase plate is properly grounded, if you have one.

Let us know what it was once you figure it out.

Regards,

mike

On Fri, May 12, 2017 at 5:20 PM, Shi, Dan <shid at janelia.hhmi.org> wrote:

> Hi Henning,
>
> There is one more thing you may check, the comma free alignment is
> important to the high resolution, beam tilted too much would distort images.
>
> Dan
>
> Dan Shi
> Janelia Research Campus, HHMI
> 19700 Helix Dr, Ashbirn
> VA 20147, USA
>
> From: 3dem <3dem-bounces at ncmir.ucsd.edu> on behalf of Henning Stahlberg <
> henning.stahlberg at unibas.ch>
> Date: Thursday, May 11, 2017 at 9:08 AM
> To: 3dem <3dem at ncmir.ucsd.edu>
> Subject: [3dem] Request for help to find source of resolution loss in
> cryo-EM
>
> Hi,
>
> We have a strange problem on one of our instruments (Titan Krios with
> Quantum-LS GIF and K2 Summit in counting mode).
> When recording images at low mag (1.04 A/px for the 4k images) and
> low-dose conditions of a partly devitrified sample, and compute the FFTs,
> we can see the 3.7A ice ring and also the second-order 2.1A ice
> ring, sometimes a third-order ring in the corners at 1.7A resolution.
> We recently observed a resolution loss visible in the FFTs only in the
> vertical direction, shown by an amplitude decay in the upper and lower
> quarter of the FFTs.
> However, while the 3.7A ring shows that anisotropic resolution loss, the
> 2.1A ring is still isotropically present.  An example image is attached.
>
> Does anybody have an idea what could cause such a behavior?
>
> Thanks,
>
> Henning.
>
>
>
> Henning Stahlberg, PhD
> Prof. for Structural Biology, C-CINA, Biozentrum, University Basel
> Mattenstrasse 26 | D-BSSE | WRO-1058 | CH-4058 Basel | Switzerland
> http://c-cina.org | Tel. +41-61-387 32 62 <+41%2061%20387%2032%2062>
>
>
>
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>
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