CM20
Alignment Procedures
Last Revised: 20070508
Gun Alignments
Gun Tilt
Gun Shift
Condenser System Alignments
Aperture Centering
Aperture Stigmation
Sample Alignments
Sample Height
Sample Orientation
Objective Lens Alignments
Pivot Points
Rotation Centers
Objective Aperture Centering
Final Alignments
Objective Lens Astigmatism
Coma Free Alignment
Sample Orientation
Imaging
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Gun Alignments
It is necessary to align the gun such that the electron beam leaves the gun
and travels along the optical axis. This will provide optimum imaging
conditions and maximum brightness.
Gun Tilt
1. Set the magnification to ~30kX.
2. Condense the beam using the SHIFT and INTENSITY knobs until the
beam is approximately half the size of the screen.
3. Press the ALGN button on the right panel and then Gun TILT using the
screen buttons
4. Use the MULTIFUNCTION knobs to maximize screen brightness
5. Exit the alignments menu by pressing ALGN
 Use the exposure time readout or your own eyes to obtain best result
 Should the beam leave the screen, bring it back to the center using the
SHIFT knobs
 Condense the beam to the smallest possible size
 Select “Gun Tilt” from the Direct Alignments menu. Use the
multifunction knobs to bring the bright spot to the centre of the beam.
It may be necessary to move the beam back to centre using the left
trackball.
Gun Shift
1. Check if the beam for spot size 3 to 9 stays in the same position, if not,
perform the alignment
2. Press ALGN and then Gun SHIFT
3. Select spot size 9 and center it using the SHIFT knobs
4. Select spot size 3 and center it using the MULTIFUNCTION knobs
5. Repeat steps 3 and 4 until spot movement is minimized
6. Exit the alignments menu by pressing ALGN
2
Condenser System Alignments
The condenser system determines the beam convergence angle of the
incident beam and therefore affects the spatial coherency envelope function
of the microscope. It also affects the intensity on the screen.
Aperture Centering
1. Select the correct C2 aperture – this is typically the second largest
2. Select SPOT SIZE 1or 2 for most imaging applications
3. Select emission 1 or 2 in the PARAMETERS menu
4. Leave this menu by pressing the READY button
5. Set the MAGNIFICATION to ~50kX
6. Center the beam using the SHIFT knobs
7. Center the aperture until the beam can expand and condense
concentrically when going through INTENSITY
Condenser Stigmation
1. Set the MAGNIFICATION to ~50kX
2. Expand and condense the beam using the INTENSITY knob. Make sure
that the beam stays round (not elliptic) and expands concentrically, if not:
3. Press STIG on the right panel and COND using the screen buttons
4. Adjust by using the MULTIFUNCTION knobs
5. Press STIG again to leave the menu
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Sample Alignments
Sample Height
1. Set the MAGNIFICATION to ~100kX
2. Position an area of interest in the center of the screen, an edge is strongly
recommended.
3. Expand the beam with the INTENSITY knobs until it covers the screen
4. Set the objective lens current to 9990mA
5. Adjust the sample height using the screw on the goniometer until the
image is in focus (minimum contrast). It should be possible to bring the
sample in focus +/- 100Å or better
 If the screw reached the end position before Gaussian focus is
obtained, the remove the sample holder, remount the sample upside
down and try again.
 The spherical aberration constant is extremely sensitive to variations
in the lens current (focus changes). This should thus be kept
reasonably constant. Coarse adjustments in focus should be performed
by changing the position of the sample in the microscope with fine
adjustments left to adjustments in the lens current.
Sample Orientation
1. Set the MAGNIFICATION to ~50kX
2. Position an area of interest in the center of the screen
3. Condense the illumination on that area using the INTENSITY knobs
4. Press D to obtain a diffraction pattern
5. Tilt the sample using the goniometer controls, to a low index zone axis of
your choice and orient it such that the intensity of the reciprocal spots is
evenly distributed on either side of the transmitted beam. (The low index
plane is then normal to the optical axis)

The sample usually moves while tilting unless it is rotated in the plane
of the sample (which it should be if sample height is at eucentric
height, see last alignment). This causes the illuminated area to change
and consequently, the diffraction pattern does not correspond to the
original selected area. Iterate by moving the sample in real space and
tilting in reciprocal space until the area of choice is aligned to the
optical axis.
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Objective Lens Alignments
Pivot Points
To ensure that shifting the beam does not also tilt the beam, the pivot points
must be aligned.
1. Set IMG SHIFT to ZERO with the screen controls
2. Set the MAGNIFICATION to ~50kX
3. Use the FOCUS knob to bring the sample to exact (Gaussian) focus
4. Condense the beam to a minimum and centre it using INTENSITY and
SHIFT
5. Press ALGN and Pivot Points X
6. Adjust MULTIFUNCTION knobs until the two spots overlap perfectly
7. Repeat for Pivot Points Y
8. Exit the menu by pressing ALGN

Should the beam leave the screen while adjusting the pivot points, use
SHIFT to bring the beam back or lower the magnification.
Rotation Centers
Current and voltage fluctuations should not produce a image shift or defocus
the image. The beam should be placed in the axis of rotational symmetry of
the objective lens. This is the same as aligning the incident beam to the
optical axis. Another benefit from this procedure is that lens aberrations are
less distinct near the optical axis.
1. Set the MAGNIFICATION to ~100Kx
2. Position an area of interest in the center of the screen, an edge is strongly
recommended.
3. Expand the beam with the INTENSITY knobs until it covers the screen
4. Use the FOCUS knob to bring the sample to exact (Gaussian) focus
5. Check that the objective lens current is still about ~9990mA, otherwise
return to “Sample Height”
6. Set FOCUS to step 3 or 4 (this gives the modulation amplitude)
7. Press ALGN and ROTATION CENTER
8. Choose either current or voltage but don’t do both (current is the more
suitable choice and voltage the easier…) Adjust MULTIFUNCTION
knobs until there is no movement in the image
9. Repeat for successively higher MAGNIFICATION up to ~300kX
10.Leave the menu by pressing ALGN
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Objective Lens Stigmation
Astigmatism causes images to appear diffuse and causes streaks in the
defocused image.
1. Set the MAGNIFICATION to ~300kX, if the astigmatism is bad, go to
lower magnifications
2. Move to an amorphous edge, preferably one that is thin (glue is not
amorphous)
3. Press STIG on the right panel and then OBJ on the screen buttons
4. Use step 3 for coarse adjustments and smaller steps for fine adjustments
5. Bring the image into focus using the FOCUS knob
6. Use the MULTIFUNCTION knobs to make the image sharper, remove
streaks or to ensure that the defocus is consistent around edges.
7. Increase the magnification to ~800kX and adjust again
8. Press STIG to leave the menu
 It is possible to use the CCD for this. Start the program “Digital
Micrograph” on one of the standalone computers. REDUCE THE
INTENSITY ON THE FLUORESCENT SCREEN UNTIL THE
SCREEN IS BLACK. Insert the CCD and press Start View in Digital
Micrograph. Increase the intensity SLOWLY until an image is
obtained. Use ROI Tools, hold down the ALT key on the keyboard
and mark out a square on the image. Then select Process -> Live ->
FFT to obtain a Fourier transform. Astigmatism is minimized when
the Fourier transform is round. Best focus is when the Fourier
transform is largest (and slightly white)
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Objective Aperture
To remove the high spatial frequency oscillations in the contrast transfer
function of the objective lens and to reduce the confusion in the image, we
introduce an aperture in the back focal plan of the objective lens, which
blocks electrons with high spatial frequency
1.
2.
3.
4.
Move the sample to an area that is both interesting and thin
Bring the sample to exact focus using the FOCUS knob
Press D to obtain a diffraction pattern
Using the diffraction pattern as a guide for the eye, insert and center the
second smallest objective aperture around the transmitted beam
5. Return to imaging mode by pressing D
6. Check with the astigmatism if the objective aperture is centered. If the
aperture is off axis the image will again appear astigmatic.
Note: Once alignment is completed, dark field imaging may be used by
centering the aperture over a non-central diffraction spot. High resolution
dark field imaging is obtained by first centering the aperture for bright field
imaging then tilting the beam using the DF button and placing a noncentered spot in the center of the aperture.
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Coma-Free Alignment
Coma is an aberration associated with objects near the optical axis. Previous
alignments are not sufficient to bring the incident beam close to the optical
axis since even beam tilts of 1 mRad can affect the image. This step ensures
that the beam is indeed aligned with the optical axis. Deviations from the
optimal position are found by inducing slight tilts to either side of the optical
axis. A beam tilt causes a slight change in focus of the image and this
change should be the same (symmetrical) on either side of the optical axis.
1.
2.
3.
4.
5.
Set the magnification to ~500kX
Move to an amorphous edge, preferably one that is thin
Bring the sample into focus using the FOCUS knob
Press ALGN and COMA FREE
If there is no image, reduce the magnification and perform the “pivot
points” correction again.
6. Press coma x
7. Adjust the MULTIFUNCTION knobs until the two images have the
same defocus and appear similar
8. Repeat for coma x
9. Change focus and check the images remain similar
10.Perform the Objective Lens Astigmatism correction again
 The Fourier transform should be the same for both images once coma
is properly corrected.
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Imaging
1. Move the sample to an area of interest
2. Use the handlebars for coarse adjustments and IMAGE SHIFT for fine
adjustments
3. Choose the best defocus for optimal imaging conditions (use “Scherzer”
for best point resolution)
4. Check (Objective) Astigmatism
5. Check Coma
6. Check Focus
7. Check the drift in the sample
8. Don’t condense the beam too strongly. A convergent beam gives poor
resolution
9. Use a smaller spot size (higher number) – intensity will unfortunately go
down
10.Use lower emission (less electrons)
11.Take Images BE CAREFUL WITH THE CCD
Good Luck!
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