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The key on the side of the keypad must always be left in the horizontal position (see image below), even when the system is powered down. However, occasionally I find that it has been turned to the vertical position, which means it is switched off. Make sure it is in the horizontal position, otherwise the lasers and anything else connected to the "Components" switch will not come on. This includes the realtime controller under the table, so you will
If you get an error in Zen that says:
"No connection to realtime controller. If the ZEN boot does not continue after 1 min, open the door of the RT PC and press the Reset button. Wait for about a minute for the ZEN boot to continue"
If you see that error and it seems there is no power to the realtime controller (no lights on, fan not rotating) then the key is probably in the wrong orientation. You message saying "Unexpected error during hardware control task. An attempt to move the filter-set "ShutterArgon" to position 1 failed", it is because the key is in the wrong position.
If this happens, you should quit Zen, turn on the key and then open Zen again.
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- Turn on the "Main switch" on the keypad
- Turn on "Systems/PC" on the keypad
- Wait for computer to boot up and log in to LSM-User account
- Turn on the "Components" switch on the keypad.
- Click the "Start System" button
Click the" Acquisition" tab and expand the "Laser" menu. Turn on the lasers you need. (Some of the lasers might show a red background initially, which means it is not ready to be used. This will go away after a couple of mintues when the laser is warmed up)
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Locating your Specimen
- Click the "Locate" tab and expand the "Microscope Control" drop-down menu
- Select the objective lens you'd like to use - either via the menu in locate', or via the touchscreen controller next to the screen.
- Select a suitable fluorescence filter. Only the first three should be used; the rest are special positions for the confocal and multiphoton
- Put your specimen on the microscope and open either the transmitted light shutter or the fluorescence shutter depending on which mode you want to use to find your sample
- Turn on the lamp by clicking its icon and pressing the On button
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- When your specimen is in focus, close the shutter and click the "Acquisition" tab.
Imaging Setup
- Expand Imaging Setup
- Choose LSM and Channel
- (Non Descanned is for multiphoton only)
- (Lambda Mode and Online Fingerprinting are for spectral imaging)
- Use Smart Setup if you want to easily set up your experimental settings (e.g. filter and detector combinations) depending on your dyes.
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- Expand Acquisition Mode. In here you can change a variety of parameters but the main ones are the scan settings:
- Scan Mode: Whether the scan switches channels in Frame or Line mode (frame by frame or line by line)
- Frame Size = Scan Format
- Line Step: Increases scan speed but lowers the resolution in one dimension by skipping lines in the scan
- Scan Speed: the line frequency. Expressed in pixel dwell time as opposed to in Hz
- Averaging, including number of scans and whether it should be done line by line or frame by frame
- Method: Mean = Average, Sum = Accumulate
- Bit Depth: 8 bit is default
- Direction: unidirectional or bidirectional (Stick to unidirectional)
- Scan Area: Zoom and Scan Field Rotation
- Expand Channels
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The function Match Pinhole sets the pinhole to keep this optimal interval and in addition sets the slice thickness for all detection channels approximately the same. This typically results in slightly thicker slices for channels detecting the longer wavelength range. In case the channels are assigned to different tracks and a Frame wise Multitracking scheme is applied, the pinhole diameter is set for each track such that the values of the resulting optical sections from the different channels are identical and have double the value of the optimal interval.
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