UCSF/IND Titan Krios 2 pixel size & sign-up guidelines

 

 

Nominal Magnification

Pixel Size (Å/pix)

Super Resolution

Pixel Size (Å/pix)

Counting mode (or after 2x FT bin)

Dose Rate 

(e-/pix/s); 

Total Dose (e-/A2)

Exposure time (s)

Dose (e-/A2/frame)

80 frames 

Magnification of 5 μm physical pixel (at super res)

64,000 

0.673 1.346

16.0 ; 52

6.0

0.66 (0.075s frame)

36284
105,000  0.417 0.834 15.0 ; 43 2.024 0.53 (0.0255s frame) 117232

130,000 

0.326

0.652

14.0 ; 66

2.024

0.82 (0.0255s frame)

148764

33,000  1.33 2.66        

 

  • Use ilab to book time for Krios2 usage and grid location/details. Please email [email protected] and [email protected] for any further modifications to grid loading and microscope/camera related issues. You can contact any Eric at any time for troubleshooting.
  • Magnifications calibrated for image shift vs coma at specified dose rate, exposure time, specific illumination area. If another magnification or dose rate necessary please contact [email protected] to modify.
  • At the beginning of each users session, reference settings files for desired magnification will be loaded and saved onto a ‘n’kx_run_new_SerialEMsettings.txt file that the user can then modify for their specific image shift, template and paths
  • energy filter tuning will be taken care of weekly, and should not be touched. energy filter zero loss peak is handled in the serialem script but if there are issues with it drifting far off users should make sure that the beam is unblanked and coordinates over an empty hole are updated in the script. If the serialem script isn't working or the zlp is lost, please contact Eric and he will handle it remotely.
  • alignments should not be touched, currently LM mags should be within a few microns with the low dose mags. If this is not the case, contact Eric
  • Scipion online drift workflows have been set for 105kx and 130kx, superres and counted respectively. Let Eric know if you have a non-standard dose/mag need.
  • 16 eps settings tested at 105kx nominal mag for 5x5 image shift (up to ~4-5 μm shift), total exposure and frame exposure adjusted. test dataset for apoferritin goes to 1.8 Å with standard processing of FTbin 2 data. This has also been done with 3x3 with 3 shots per hole. 

 

Last updated: 07/20/21