1. Turn on the computer (#1)
2. If required, turn on the incubation power bar

3. Turn on the camera and laser power bar on the left of the microscope (#2)

4. Turn on the microscope power bar on the right of the microscope (#3)

5. Use your UdeM credentials to log in to Windows
   

   Remarque
   When using for the first time, it is necessary to import the microscope-specific parameters BEFORE starting the software. See the First Use section below.

6. Start the Zen Blue software

Please note, this procedure will delete all your experiment protocols and restore the software to its original settings.

On the microscope touch screen:

1. Press Home>Load Position to lower the stage to its lowest position
2. Press Set Work Position to store this position
3. If necessary, move the focus slightly up to remove the “Lower Z limit reached” message displayed on the touchscreen
4. Press Home>Microscope>Turret>Objectives>10x to select the 10x objective
5. If asked, tap Done to remove the oil lens cleaning warning
6. Press Home>Microscope>XYZ>Position>Z-Position>Set zero>Auto to perform focus calibration
7. Press OK to start the focus calibration procedure
8. Wait a few seconds for the calibration to be completed

Make sure to calibrate the focus before performing the first focus.

Once calibrated, the focus can be found at Z = 5 mm). The Z value can be found on the microscope touch screen Home>Z-Position

First focus with the safest lens before selecting another lens and continuing with secondary focus.

This microscope does not have additional air objectives. However, if there were, the procedure would be as follows:

After performing the first focus, on the microscope touch screen:

1. Press Home>Microscope>Turret>Objectives
2. Press on the desired lens

In Zen Blue software:

1. In the Locate tab, select BF or the desired fluorescence (CFP/GFP/DsRed ou GFP/RFP/Cy5, etc…) to activate the configuration
2. Adjust the focus with the precision dial while looking through the eyepieces until the image is perfectly sharp
3. In the Locate tab, select Off to turn the illumination off
4. Your sample is ready for acquisition!

The 63x objective is the best oil objective because it has the most optical corrections (Plan Apochromat) and the largest numerical aperture (1.4). It offers a lateral resolution of 240nm at a wavelength of 550nm.

• Files can be saved temporarily (during acquisition) on the local C: drive (desktop)
• At the end of each session, copy your data to your external drive and delete it from the local C: drive
• You can store your files on the D: drive (Data Storage). If you do, please create a folder per laboratory using the principal investigator last name. Within, create one folder per user (Firstname_Lastname).

1. Save your data
2. Close the software Zen Blue
3. Transfer your data to the D: drive (Data Storage) or to your external drive and delete it from the local C: drive
4. If used, turn of the incubation power bar
5. If used, clean oil lenses with lens cleaner and paper

6. Turn off the microscope power bar on the right of the microscope (#3)

7. Turn off the camera and laser power bar on the left of the microscope (#2)
8. Turn off the computer

• Zen quick start guide.pdf

(FIXED) Problem with the FRAP module:

At first could not modulate the laser intensity (yet could not find the spot on the field), rebooted and then the FRAP fiber does not carry any light (even 488)

==> Laser manipulation module was too far in x and y (noticeable as a tilt and a gap between both parts of the module).

Corrected Trigger Module configuration for the BackCam

• reassigned each camera with its own trigger module in MTB config 

• DualCam calibration parameters set, using beads
• Realigned Lasers:
  • Laser output measured in spinning disk mode at the sample with 10x objective, 1x Extended tubelens, 100% laser power and ALL lasers ON:
    
    • 405nm: 0.70 mW before => 0.66 mW after
      • with laser 488 OFF: 0.81 mW before => 0.83 mW after
    • 488nm: 3.75 mW before => 4.17 mW after  
      • with laser 405 OFF: 3.95 mW before => 4.33 mW after
    • 561nm: 2.1 mW before => 2.37 mW after 
    • 639nm: 0.9 mW before => 1.1 mW after 
• NOTE: AOTF2 for FRAP

• Error message when switching between FRAP & SD illumination

When both 405nm and 488nm laser are ON simultaneously, the output power is decreased by 11%.

• Error when the definite focus is used (unexpected error the definite focus did not respond)
• To solve it:
  • Turn off the Zen Software
  • Turn off the microscope power bar (#3)
  • Turn off the definite focus by pressing and holding the definite focus button
  • Turn on the definite focus by pressing once the definite focus button
  • Wait until the definite focus display shows "Detecting stand"
  • Turn on the microscope power bar (#3)
• Confirm that the definite focus is showing in the microscope tactile display
• Open Zen software and run an experiment using the definite focus to confirm it is fully functional

• Added to wiki

• AOTF1 changed

The spinning disk is a multipoint confocal. Outside the focal plane the sample disappears completely. It is therefore easier to find your sample through the eyepieces. Follow the tuning procedure to achieve this.

Yes. It is an inverted microscope designed for the observation of living specimens. The spinning disk is particularly appreciated for its limited phototoxicity. This is an inverted microscope designed to look at specimen in a dish or a multi-well plate. The objectives are optimized to image through thin glass bottom multi-well plates. You may also image specimen mounted between a slide and a 0.17mm thick coverslip.

Image Removed