tSEM Imaging

Before you start your work, go through this checklist:

Vac

Gun status

Current/Voltage

Location of STEM detector and stage

Exchanging grids: 

To remove grids from the SEM

1. Turn the EHT off
2. Double click on 'Safe Stem'
3. Click 'Stem out'
4. Under the Vacuum tab, click on 'Vent'
5. Turn on the N₂ gas
6. Wait for the chamber to completely vent, then open the chamber door
7. Turn off the N₂ gas
8. Remove grid holder and be sure to wear gloves. Do not touch the grid holder or anything inside the chamber without wearing gloves
9. Exchange grids and be sure to document what grids were removed and what grids were added

To add grids to the SEM

1. Load grids into chamber and close the door

2. Under the Vacuum tab, click on 'Pump'

3. Double click 'Safe Stem'

4. Once there is a green check mark next to Vacuum, click 'Stem In'

5. Double click on a position in the Stage Point List

6. Check vacuum levels to see if they are steady. If so, turn EHT on

Four routine gun alignment checks:

Gun alignment

1. Scroll to an area on the aluminum flat surface of the grid holder. Set detection mode to SE2. Focus the image

2. Under the Aperture tab, click on 'gun alignment'

3. Click 'Emission'

4. Set scan speed to 9

5. Go back to the aperture tab and center beam if necessary by using the sliders on the Gun Alignment grid

Aperture alignment

1. Scroll to a tissue section on the grid

2. Click on STEM and then click 'Normal' 

3. Click on aperture alignment

4. Click  'focus wobble on' in the SmartSEM Status window

5. Adjust alignment accordingly with the aperture x and y knobs on the keyboard

6. Click 'focus wobble off' in the SmartSEM Status window

Objective current wobble

1. Click 'objective current wobble on' in the SmartSEM Status window

2. Adjust wobble with aperture x and y knobs accordingly

3. Click 'objective current wobble off' 

Stigmation calibration

1. Click and expand side bar
2. Click on calibrate stigmation
3. Adjust stigmation with the stigmator knobs accordingly
4. Adjust stigmation further with the calibration window if necessary