Improving Autofocusing

Because MSI-raster is normally used for acquiring images on negatively stained particles on continuous carbon, the intermediate mag preset (hl) often lacks contrast. To reduce focusing failure, we recommend a different focusing sequence for the Z focus node.

  • Use Stage Alpha Tilt to determine defocus at the low magnification sq preset to do a rough stage-z adjustment to U-center

  • Refine stage-z position at the high magnification fa preset and Use beam-tilt to determine the required correction.

For "Stage_Wobble" Step:

  • Enable = Yes

  • Image registration= phase correlation

  • Fit limit = 1000 (not used)

  • Correct for delta Defocus/Z between 0 and 2e-4 meters

  • Correction type = Stage Z

  • Check drift greater than 3e-10 meters/s = No

  • Stigmator correction = No

  • Stigmator Defocus Max/Min = N/A

For "Z_to_Eucentric" Step, change these:

  • preset = fa

  • Correct for delta Defocus/Z between 0 and 2e-5 meters

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