1x transmitted light Electronically Switchable Illumination and Detection module (ESID)
1x Airyscan detector for 63x objective
If not already done, turn on the computer (#1) and use your UdM credentials to log in to Windows
Remove the dust cover from the microscope
Turn on the System (#2) and Components (#3) switches in the rack below microscope
Turn on the laser key (#4) in the rack below the microscope
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.
Start the Zen software
When using for the first time, it is necessary to import the microscope-specific parameters into the software.This procedure is usually carried out during the training session.However, it is also possible to use it to reset the software if it is not displayed correctly, for example.
Please note, this procedure will delete all your experiment protocols and restore the software to its original settings.
If open, close the Zen software and wait for it to close completely (up to 30 seconds)
On the Desktop open the Documentation folder
Double-click Zen Settings for LSM900
A script will run and a black window will appear briefly
Click OK to close the message Settings for Zen have been imported successfully.
You can then open the Zen software
This procedure puts the microscope in a safe configuration and performs a calibration.At the end of this procedure the microscope will be ready for acquisition.
This step is required to calibrate the microscope in XY and Z. Doing a calibraiton will save you a lot of time to find your focus or your sample.
On the microscope touch screen::
If not already done, press Home>Microscope>Control>Objectives>5x to select the 5x objective
In the Zen Software
Click Calibrate Now
The microscope will lower the objectives perform a XY calibraiton and then a Z calibration
Once calibrated, the focus can be found at Z = 1.7 mm). The Z value can be found on the microscope touch screen Home>Z-Position
On the microscope touch screen:
If not already done, Press Home>Load Position to lower the objectives to the lowest position
Press Set Work Position to store this position
If necessary, move the focus slightly up to remove the Lower Z limit reached message displayed on the touchscreen
If not already done, press Home>Microscope>Control>Objectives>5x to select the 5x objective
If asked, tap Done to remove the oil lens cleaning warning
Press Home>Microscope>XYZ>Position>Z-Position>Set zero>Auto to perform focus calibration
Press OK to start the calibration procedure
Wait a few seconds for the calibration to be completed
Press Home>Microscope>XYZ>Position>XY-Position>Set zero>Auto to perform a stage calibration
Press OK to start the calibration procedure
Wait a few seconds for the calibration to be completed
Once calibrated, the focus can be found at Z = 1.7 mm). The Z value can be found on the microscope touch screen Home>Z-Position
Important
Make sure to calibrate the focus before performing the first focus.
On the microscope touch screen:
If not already done, press Home>Microscope>Turret>Objectives>5x to select the 5x objective
The 5x objective is the safest because it has the longest working distance (12mm).The sample will appear perfectly sharp long before the lens approaches it.It is recommended to always first focus with the safest lens.The objectives are parafocal, focusing with the safest objective will then allow you to easily find your sample with another objective. The 10x objective is also safe because its working distance is 6.5 mm.
If not done already, press Home>Load Position to lower the objective to the lowest position and press Set Work Position to store this position
If necessary, move the focus slightly up to remove the Lower Z limit reached message displayed on the touchscreen
Place the test slide on the microscope stage with the coverslip toward the objective
Important
Always use the test slide to perform the first focus.
If necessary, move the stage so that the sample is centered on the objective
On the computer:
Open Zen
In the Locate tab, select BF or the desired fluorescence (DAPI, GFP, mPlum) to activate the configuration
Adjust the focus with the main dial while looking through the eyepieces until the image is perfectly sharp
Once calibrated, the focus can be found at Z = 1.7 mm). The Z value can be found on the microscope touch screen Home>Z-Position
In the Locate tab, select Off to turn off the illumination
Important
First focus with thesafest objective before selecting another lens and continuing with secondary focus.
After performing the first focus, on the microscope touch screen:
Press Home>Microscope>Control>Objectives, press 10x, 20x or 40x to select the desired objective
The 40x objective is the best Air objective because it has the greatest number of optical corrections (Plan Apochromat) and the largest numerical aperture (0.95) but has a smaller field of view. The 20x/0.8 objective offers the best compromise between Resolution and Field of View
There are two (2) 40x objectives, make sure you select the Air 40x
In Zen software :
In the Locate tab, select BF or the desired fluorescence (DAPI, GFP, mPlum) to activate the configuration
Adjust the focus with the precision dial while looking through the eyepieces until the image is perfectly sharp
In the Locate tab, select Off to turn the illumination off
Your sample is ready for acquisition!
After performing the first focus, on the microscope touch screen:
Press Home>Microscope>Turret>Objectives
Press 63x Oil, 40x Oil to select the desired objective.The microscope will automatically lower the stage so that the sample is accessible.
The 40x and 63x oil objectives provide the same spatial resolution because they have the same numerical aperture (1.4). The 40x oil objective offers a larger field of view and transmits light slightly better beyond 700nm. The 63x oil objective transmits light slightly better in the visible spectrum (440-710 nm) and has a better Strehl ratio (90%). It is particularly suited for super-resolution imaging, but its field of view is smaller.
There are two (2) 40x objectives, make sure you select the 40x Oil
Place a single drop of oil on the objective
Press Done.The microscope will automatically return the objective to its original position
In Zen software:
In the Locate tab, select BF or the desired fluorescence (DAPI, GFP, mPlum) to activate the configuration
Adjust the focus with the precision dial while looking through the eyepieces until the image is perfectly sharp
In the Locate tab, select Off to turn the illumination off
Your sample is ready for acquisition!
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).
In any case, your files should be removed from the C: drive.
Save your data
Close the software Zen
Transfer your data to the D: drive (Data Storage) or to your external drive and delete it from the local C: drive
If used, clean oil lenses with lens cleaner and paper
Select the 5x objective and press load position to place the objectives in a safe position
Turn off the laser key (#4) in the rack below the microscope
Turn off theComponents (#3) and System (#2) switches in the rack below the microscope
Turn off the computer
Cover the microscope
Important Reminders
Take back your samples including ones in the microscope
Leave the microscope and the working area clean
Quality control for Illumination, Liquid light guide and filters quality.
Joystick replaced
Installation Airyscan upgrade. New workstation. New detector. New casing.
Issue when the joystick is at the rest position. If it is slightly to the left, the stage will move while the image is acquired. Temporary fix: Make sure the joystick is slightly to the right when it is resting.
User guide added to Wiki French and english version
Zen 2.6 updated hotfix 12
Microsoft Windows updated
Added to wiki
Stand
Zeiss AxioObserver LSM900 Serial: 2633000272
Manual Field diaphragm for transmitted light
Manual polarizer
Left imaging port with LSM900 confocal
Manual Aperture diaphragm
Manual Fluorescence field diaphragm
6x motorized nosepiece
6x Motorized reflector changer
3x Motorized sideport turret (100% Visible, 100% Left (confocal), 100% right (empty)
TL Motorized shutter
RL Motorized shutter
Light sources
Transmitted LED light
X-Cite 120LED mini Serial XT120LM-0279 RS232 COM Port 6
Laser URGB 400102-9300-000 Serial 2631000466 Toptica iChrome-ZLE-4_40407 v3.1 Aug 2016
Condenser
Manual condenser
Condenser Lens NA 0.55 WD 26mm
Manual polarizer
Filter turret 6 positions manual
H Empty
Ph1
Ph2
Ph3
DIC II #426702
DIC III #426706
Objectives
5x/0.15 Ph1 Air WD 5.6 N-AchroPlan 420931-9911-000
10x/0.25 Ph1 WD 6.0 N-AchroPlan 420941-9911-000
20x/0.80 Air WD 0.61 Plan-Apochromat 420650-9901-000
40x/0.95 Air WD 0.25 Plan-Apochromat 420660-9970-000 Variable coverslip 0.13-0.21
1 transmitted ESID Motorized Left TL ESID LED, Right ESID Detector
Workstation
HP Z2 G9 Serial CZC4027PF7 829W6EC#ABB
I155440-CIB
Motherboard HP 895C
BIOS version 2024-09-03
1 x Intel Core i7-12700K 3.6 GHz
RAM 128GB DDR5 4 x 32GB no ECC
OS 500 GB NVMe 6 GB/s
7 TB HD Data Storage 213 MB/s
Nvidia RTX A4000 16 GB GDDR6
Monitor HP Z32x
Software Zen 3.10
Incubation
Consumables
Troubleshooting
The best way to solve a problem in Microscopy is to follow the light path.You will find in the Light path tab of this page, the diagrams which will allow you to follow the light all along its path through the microscope.
Open the light path file
Starting from the light source and moving towards the detector, verify that there is indeed light after each component of the microscope
FAQ
Yes. It is an inverted microscope designed for observing specimens in culture. The objectives are optimized for viewing through glass-bottom multi-well plates. It is also possible to observe samples between a slide and coverslip (with a thickness of 0.17mm).
