3.3 Testing Crystals
for Diffraction in Batch
1. Go to a saved crystal coordinate in the Navigator using “Go to
XY”.
2. Center the crystal in the SA aperture.
3. Remove the current crystal coordinate.
4. Click “Add Stage Pos” to update the (x,y) position of the
current crystal.
5. Repeat steps 1 through 4 (Subheading 3.3), above for the
remaining crystal targets.
6. Set crystal targets to “Acquire” in the Navigator.
7. Adjust camera parameters as necessary for the Record beam.
8. Open a new file to save images.
9. Use the “Acquire at Items” function to collect the Record
image by choosing the Primary Task as “Just acquire and save
image or montage”. The microscope collects diffraction images
at the saved crystal locations and saves them into the opened
file for later review. Crystal targets with good diffraction are
kept in the Navigator for automated data collection.
3.4 Preparing
Crystals for Data
Collection (See Note 2)
1. Ensure crystals are at eucentric height for each target. Similar
to Subheading 3.3. above, this can be automated in SerialEM
using the “Acquire at Items” function.
(a) Exit the Low Dose Mode.
(b) Turn off diffraction mode if activated, and select an imaging magnification that clearly shows the crystal target
in view.
(c) In the Navigator, set crystal targets to “Acquire”.
(d) Adjust camera parameters as necessary for the
Record beam.
(e) Open a new file to save images if desired.
(f) In the “Acquire at Items” dialog, choose the Primary Task
to run the Eucentricity function for each target. Click
“GO” to start. The microscope goes to each crystal location, runs the automated Eucentricity routine, and
updates the Z-height for each target.
2. Check and decide on the tilt range for data collection.
(a) Go to a crystal target on the Navigator.
(b) Go to the defocused-diffraction beam set in step 4 (Subheading 3.1), set as the Low Dose View Mode.
(c) While using the phosphor screen, or live imaging, change
the stage alpha tilt as desired to check the maximum tilt
range for the crystal and note the imaging limits of the
crystal.
Automation of Continuous-Rotation Data Collection for MicroED
325
for Diffraction in Batch
1. Go to a saved crystal coordinate in the Navigator using “Go to
XY”.
2. Center the crystal in the SA aperture.
3. Remove the current crystal coordinate.
4. Click “Add Stage Pos” to update the (x,y) position of the
current crystal.
5. Repeat steps 1 through 4 (Subheading 3.3), above for the
remaining crystal targets.
6. Set crystal targets to “Acquire” in the Navigator.
7. Adjust camera parameters as necessary for the Record beam.
8. Open a new file to save images.
9. Use the “Acquire at Items” function to collect the Record
image by choosing the Primary Task as “Just acquire and save
image or montage”. The microscope collects diffraction images
at the saved crystal locations and saves them into the opened
file for later review. Crystal targets with good diffraction are
kept in the Navigator for automated data collection.
3.4 Preparing
Crystals for Data
Collection (See Note 2)
1. Ensure crystals are at eucentric height for each target. Similar
to Subheading 3.3. above, this can be automated in SerialEM
using the “Acquire at Items” function.
(a) Exit the Low Dose Mode.
(b) Turn off diffraction mode if activated, and select an imaging magnification that clearly shows the crystal target
in view.
(c) In the Navigator, set crystal targets to “Acquire”.
(d) Adjust camera parameters as necessary for the
Record beam.
(e) Open a new file to save images if desired.
(f) In the “Acquire at Items” dialog, choose the Primary Task
to run the Eucentricity function for each target. Click
“GO” to start. The microscope goes to each crystal location, runs the automated Eucentricity routine, and
updates the Z-height for each target.
2. Check and decide on the tilt range for data collection.
(a) Go to a crystal target on the Navigator.
(b) Go to the defocused-diffraction beam set in step 4 (Subheading 3.1), set as the Low Dose View Mode.
(c) While using the phosphor screen, or live imaging, change
the stage alpha tilt as desired to check the maximum tilt
range for the crystal and note the imaging limits of the
crystal.
Automation of Continuous-Rotation Data Collection for MicroED
325