Imaging and Flow Cytometry Core CPOS
LSM 800 SOP
I-13
ver. 2.1 (Updated: 11/13/2020)
I.
Z-stack set up
Pre-requisite: you have setup all necessary channels according to previous chapters including laser power, master
gain, pin hole size.
1.
**Note: image intensity might increase beyond
saturation if the z-position you chose to setup
laser power / master gain is not the brightest
structure in the z-volume. Decrease laser power
/ gain accordingly. **
2.
click checkbox to activate 'Z-stack' module.
3.
select one channel with the most representative
structures / most robust in the group.
4.
turn ON 'Live'.
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5.
hands on **Fine Focus** knob. Please note
current z-position value (greater numeric value =
higher focus position)
or
6.
cursor over preview window. Press 'Control' key
on keyboard. Scroll with mouse wheel while
'Control' key is pressed.
7.
decrease focus height until you cannot see your
cell / structure. stop and click 'Set First'.
8.
Increase focus height until you cannot see your
cell / structure. stop and click 'Set Last'.
9.
Click 'Stop'. to pause laser scanning.
10.
note on 'Range' value to confirm if your desired
structure is roughly the same size as stated.
Readjust 'First' and 'Last' section positions
accordingly.
11.
note optical thickness under pinhole slider in
'Channels' window. This is your maximum
interval if you wish to capture all structure in a
certain z-volume.
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