9. Blank the beam and switch the microscope into diffraction
mode that has been set up to collect low-dose diffraction
patterns.
10. Unblank the beam and collect a still exposure diffraction pattern from the single crystal.
11. Assess the quality of the diffraction pattern. If the reflections
are sharp, well separated, and extend to high resolution (see
Fig. 2), proceed to the next section to collect a continuous
rotation data set from this crystal. If the crystal shows poor
diffraction, or no diffraction, switch the microscope back to
imaging mode, remove the selected area aperture and beam
stop, and return to step 5.
3.3 MicroED Data
Collection
1. Once a crystal has shown high-quality diffraction from a single
exposure. Return the microscope to imaging to visualize the
crystal on the grid square again (500–1000Â magnification).
Remove the selected area aperture and beam stop.
2. Tilt the stage in both the positive and negative directions to
ensure the crystal remains centered while rotating and to determine the maximum possible tilt range that can be collected
without the crystal being obscured by other crystals or the bars
of the grid. If the crystals move while rotating, correct the
Z-height to return the crystal to the eucentric position.
3. Once the tilt range to be collected has been confirmed in step
2, tilt the grid to the maximum tilt angle of this range. Put in
the selected area aperture and beam stop. Blank the beam.
4. Based on the manufacturer and model, set up the stage rotation
such that when data collection begins, the stage can be rotated
slowly from the starting tilt angle to the end tilt angle (see Note
7).
5. According to manufactures direction, set the high-speed detector up for data to be collected in a movie mode.
6. Return the microscope to diffraction mode used for collecting
low-dose diffraction patterns.
7. Unblank the beam and begin the rotation of the stage. Once
the stage begins to rotate, begin recording with the high-speed
detector.
8. Once the data set has been collected, return the stage to 0
tilt,
remove the beam stop and the selected area aperture.
9. Repeat the procedures presented in Subheadings 3.2 and 3.3 to
collect data sets from multiple crystals on the grid. Generally,
multiple crystals are needed for structure determination.
Therefore, it is recommended to collect as many high-quality
data sets as possible from grid that shows high-quality crystals.
Following the successful use of these methods on suitable
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Guanhong Bu and Brent L. Nannenga
mode that has been set up to collect low-dose diffraction
patterns.
10. Unblank the beam and collect a still exposure diffraction pattern from the single crystal.
11. Assess the quality of the diffraction pattern. If the reflections
are sharp, well separated, and extend to high resolution (see
Fig. 2), proceed to the next section to collect a continuous
rotation data set from this crystal. If the crystal shows poor
diffraction, or no diffraction, switch the microscope back to
imaging mode, remove the selected area aperture and beam
stop, and return to step 5.
3.3 MicroED Data
Collection
1. Once a crystal has shown high-quality diffraction from a single
exposure. Return the microscope to imaging to visualize the
crystal on the grid square again (500–1000Â magnification).
Remove the selected area aperture and beam stop.
2. Tilt the stage in both the positive and negative directions to
ensure the crystal remains centered while rotating and to determine the maximum possible tilt range that can be collected
without the crystal being obscured by other crystals or the bars
of the grid. If the crystals move while rotating, correct the
Z-height to return the crystal to the eucentric position.
3. Once the tilt range to be collected has been confirmed in step
2, tilt the grid to the maximum tilt angle of this range. Put in
the selected area aperture and beam stop. Blank the beam.
4. Based on the manufacturer and model, set up the stage rotation
such that when data collection begins, the stage can be rotated
slowly from the starting tilt angle to the end tilt angle (see Note
7).
5. According to manufactures direction, set the high-speed detector up for data to be collected in a movie mode.
6. Return the microscope to diffraction mode used for collecting
low-dose diffraction patterns.
7. Unblank the beam and begin the rotation of the stage. Once
the stage begins to rotate, begin recording with the high-speed
detector.
8. Once the data set has been collected, return the stage to 0
tilt,
remove the beam stop and the selected area aperture.
9. Repeat the procedures presented in Subheadings 3.2 and 3.3 to
collect data sets from multiple crystals on the grid. Generally,
multiple crystals are needed for structure determination.
Therefore, it is recommended to collect as many high-quality
data sets as possible from grid that shows high-quality crystals.
Following the successful use of these methods on suitable
292
Guanhong Bu and Brent L. Nannenga