[3dem] Low Dose Focus/Exposure

[Eric Gilles Hanssen]

HI Tommi,
If the defocus series is unticked then your issue comes from the alignment file. I am guessing you have a different magnification for Focus and Acquisition. If that is not the case , then I don’t know. If that is the case then check that the Alignment parameter “Image Shift and Focus” at the magnifications (I am guessing either MH and SA or just SA) of interest are correct and redo them if they are not.
Regards
Eric


-----------------
Dr Eric Hanssen

Senior Research Fellow
Head - Melbourne Advanced Microscopy Facility – Bio21 Institute node
Adjunct Senior Fellow – Department of Biochemistry and Molecular Biology

Bio21 Molecular Science and Biotechnology Institute
30 Flemington Road - The University of Melbourne - Victoria 3010 - Australia
email: ehanssen at unimelb.edu.au<mailto:ehanssen at unimelb.edu.au> | Office: +61 3 83442449 | Microscope: +61 3 83442509 | Fax: +61 3 9347 8265
Web: www.microscopy.unimelb.edu.au<http://www.microscopy.unimelb.edu.au/>

[BANNER_foremail]

From: 3dem [mailto:3dem-bounces at ncmir.ucsd.edu] On Behalf Of Grassucci, Robert
Sent: Tuesday, 28 April 2015 8:45 AM
To: Tommi White; 3dem at ncmir.ucsd.edu
Subject: Re: [3dem] Low Dose Focus/Exposure

Hi Tommi
There is an option in low dose to take a defocus series.  Make sure that that is not activated.  It is designed to take an exposure at a certain defocus from where you set it in focus mode.  That would me my best guess.  Good luck.
Bob


From: 3dem [mailto:3dem-bounces at ncmir.ucsd.edu] On Behalf Of Tommi White
Sent: Monday, April 27, 2015 5:53 PM
To: 3dem at ncmir.ucsd.edu<mailto:3dem at ncmir.ucsd.edu>
Subject: [3dem] Low Dose Focus/Exposure

Dear 3DEM.

Perhaps the community can assist with this issue we are struggling with.  In Low Dose on our Tecnai F30, the Low Dose focus image has a different defocus value (using FFT) versus Low Dose Exposure.

I have been checking this on a relatively flat cross-grating (not cryo) specimen.  I set LD exposure to be zero defocus (with focus knob), but when I switch over to LD focus (~ 3 micron away on tilt axis) the image is defocused (~ -2 micron).

I believe there is some 200 kV procedure alignment that must not be set properly - but am not quite sure which one...any suggestions on what to tweak to get this exposure/focus area reflecting the same defocus?
Thanks for any suggestions...
Tommi
Tommi A. White, Ph.D.
Associate Director, Electron Microscopy Core Facility
Assistant Professor of Biochemistry
University of Missouri
W125 Veterinary Medicine
Columbia, MO 65211
573-882-8304
http://emc.missouri.edu
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