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_Micro-head-en
_Micro-head-en

Button Hyperlink
iconbuild
titleFrançais
typeprimary
urlZeiss LSM-900 Airyscan


Tabs Container
idInstrument Tabs
titleZeiss Axio-Imager Z2
directionhorizontal
Tabs Page
idDescription
titleDescription

Zeiss LSM900 confocal Airyscan microscope

Desmarais Building, Room 2234

Advanced Microscope Tier 2 usage price

Instrument awarded to Dr. Jean-Philippe Gratton by the Canadian Foundation for Innovation (CFI)

  • Applications
    • Transmitted light
    • Interference Contrast (DIC)
    • Phase Contrast
    • Fluorescence
    • Laser scanning confocal

  • Light sources

    • 12V 100W halogen lamp for transmitted light

    • X-Cite 120LED mini for visible fluorescence

      Expand
      titleX-Cite 120LED mini complete specifications

      Emission peak (nm)

      Power (mW)

      334

      5



  • Objectives

  1. 5x/0.15 Ph1 Air WD 5.30 N-AchroPlan 420931-9911

  2. 10x/0.25 Ph1 WD TBD N-AchroPlan 420941-9911

  3. 20x/0.80 Air WD 0.61 Plan ApoChromat 420650-9901

  4. 40x/1.4 Oil WD TBD Plan ApoChromat 420762-9900

  5. 63x/1.40 Oil WD 0.19 Plan ApoChromat 420782-9900

  6. 40x/0.95 Air WD TBD Plan ApoChromat 420660-9970 Variable coverslip 0.13-0.21
  7. Expand
    titleComplete lens specifications

    Position

    Nom

    Marque

    Nom complet

    Identifiant

    Grossissement

    Ouverture numérique

    Immersion

    Type

    Distance de travail (mm)

    Transmittance

    (% [nm])

    Technique

    Épaisseur du couvre-objet (mm)

    2

    20x/0.80
    Air

    Zeiss

    20x/0.8 DIC II

    Plan-Apochromat
    W0.8x1/36"

    440640-9903-000

    20x

    0.8

    Air

    Plan Apochromat

    0.55

    >90% [410-800]

    BF, DIC, Fluo

    0.17

    4

    63x/1.40

    Huile

    Zeiss

    63x/1.4 DIC III

    Plan-Apochromat

    M27

    420782-9900-000

    63x

    1.4



    Note
    Huile



    Plan Apochromat

    0.19

    >80% [440-710]

    BF, DIC, Fluo

    0.17

    BF: Bright-field
    DIC: Interference contrast

  • Filter cubes
  1. 38 GFP
  2. 49 DAPI
  3. 64 mPlum
  4. DIC Analyzer
  5. Empty
  6. Empty
    Expand
    titleComplete filter specifications


    Position

    Nom

    Marque

    Identifiant

    Filtre d'excitation 

    Miroir dichroïque

    Filtre d'émission

    Commentaire



    1

    DAPI

    Filter Set 49

    Zeiss

    488049-9901-000

    365/50

    [340-390]

    395LP

    445/50

    [420-470]




    2

    GFP

    Filter Set 38

    Zeiss

    000000-1031-346

    470/40
    [450-490]

    495LP

    525/50

    [500-550]


    FT 495

    BP 525/50



  • Detector
    • 2 GaASP PMT


Tabs Page
idUser Guide
titleUser Guide


UI Expand
expandedtrue
titleStartup
  1. Remove the dust cover from the microscope
  2. Turn on the computer (#1)

  3. Turn on the System (#2) and Components (#3) switches in the rack on the left of the microscope

  4. Turn on the laser key (#4) in the rack on the left of the microscope

  5. Use your UdM credentials to log in to Windows

    Note
    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 software


UI Expand
titleFirst Use

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.

Note

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

  1. If open, close the Zen software and wait for it to close completely (up to 30 seconds)
  2. On the Desktop open the Documentation folder
  3. Double-click Settings for LSM800
  4. A script will run and a black window will appear briefly
  5. You can then reopen the Zen software


UI Expand
titleLoading samples

This procedure puts the microscope in a safe configuration and performs a focus calibration. At the end of this procedure the microscope will be ready for acquisition.

UI Expand
titleFocus Calibration Z

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>Control>Objectives>5x to select the 5x 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
Note

Once calibrated, the focus can be found at Z = 1.6 mm). The Z value can be found on the microscope touch screen Home>Z-Position


UI Expand
titleFirst focus
Warning
titleImportant

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

On the microscope touch screen:

  1. Press Home>Microscope>Turret>Objectives
  2. Press 5x to select the 5x lens
    Info

    The 5x objective is the safest because it has the longest working distance (TBD mm). 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 para-focal, focusing with the safest objective will then allow you to easily find your sample with another objective.

  3. Press Home>Load Position to lower the stage to its lowest position
  4. Press Set Work Position to store this position
  5. If necessary, move the focus slightly up to remove the “Lower Z limit reached” message displayed on the touchscreen
  6. Place the test slide on the microscope stage with the coverslip toward the objective
    Note
    titleImportant

    Always use the test slide to perform the first focus.

  7. If necessary, move the stage so that the sample is centered on the objective

On the computer:

  1. Open the Zen
  2. In the Locate tab, select BF or the desired fluorescence (DAPI, GFP, mPlum) to activate the configuration
  3. Adjust the focus with the main dial while looking through the eyepieces until the image is perfectly sharp

    Note

    Once calibrated, the focus can be found at Z = 1.6 mm). The Z value can be found on the microscope touch screen Home>Z-Position


  4. In the Locate tab, select Off to turn off the illumination


UI Expand
titleSeconday focus
Warning
titleImportant

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


UI Expand
titleFocusing with air objectives

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

  1. Press Home>Microscope>Turret>Objectives
  2. Press 10x, 20x or 40x to select the desired lens
    Info
    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). It offers a lateral resolution of 420nm at a wavelength of 550nm.
    Note

    There are two (2) 40x objectives, make sure you select the Air one


  3. Adjust the focus with the precision dial while looking through the eyepieces until the image is perfectly sharp
  4. In the Locate tab, select Off to turn the illumination off
  5. Your sample is ready for acquisition!


UI Expand
titleFocusing with oil lenses

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

  1. Press Home>Microscope>Turret>Objectives

  2. Press 63x Oil, 40x Oil to select the desired lens. The microscope will automatically lower the stage so that the sample is accessible.

    Info

    The 40x 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.

    Note

    There are two (2) 40x objectives, make sure you select the OIL one


  3. Place a drop of oil on the objective
  4. Press Done. The microscope will automatically return the sample to its original position

In Zen software:

  1. In the Locate tab, select BF or the desired fluorescence (DAPI, GFP, mPlum) 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!
UI Expand
titleStorage management
  • 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).
Note

In any case, your files should be removed from the C: drive.

UI Expand
titleShutdown
  1. Save your data
  2. Close the software Zen
  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, clean oil lenses with lens cleaner and paper
  5. Select the 5x objective and press load position to place the objectives in a safe position
  6. Turn off the laser key (#4) in the rack on the left of the microscope
  7. Turn off the System (#2) and Components (#3) switches in the rack on the left of the microscope

  8. Turn off the computer

  9. Cover the microscope
Note
titleImportant Reminders
  • Take back your samples including ones in the microscope
  • Leave the microscope and the working area clean
Tabs Page
idLight Path
titleLight Path

The following diagrams allow you to follow the light path in transmitted light (bright field, DIC) and in reflected light (fluorescence).



Tabs Page
idManuals
titleManuals


Tabs Page
idLog
titleLog
UI Expand
titleTo do
  • Quality control for Illumination, Liquid light guide and filters quality.
UI Expand
title2024-09-19 Joystick replacement
  • Joystick replaced
UI Expand
title2024-09-03 Airyscan upgrade
  • Installation Airyscan upgrade. New workstation. New detector. New casing.
UI Expand
title2024-07-08 Joystick issue
  • 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.
UI Expand
title2024-02-14 Added to wiki
  • User guide added to Wiki French and english version

UI Expand
title2021-10-20
  • Zen 2.6 updated hotfix 12
  • Microsoft Windows updated

UI Expand
title2021-09-27
  • Added to wiki


Tabs Page
idTechnical Datasheet
titleTechnical Datasheet

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

    1. H Empty

    2. Ph1
    3. Ph2
    4. Ph3
    5. DIC II #426702
    6. DIC III #426706

Objectives

  1. 5x/0.15 Ph1 Air WD 5.6 N-AchroPlan 420931-9911-000

  2. 10x/0.25 Ph1 WD 6.0 N-AchroPlan 420941-9911-000

  3. 20x/0.80 Air WD 0.61 Plan-Apochromat 420650-9901

  4. 40x/0.95 Air WD 0.25 Plan-Apochromat 420660-9970 Variable coverslip 0.13-0.21
  5. 40x/1.4 Oil WD 0.13 Plan-Apochromat 420762-9900-000

  6. 63x/1.40 Oil WD 0.19 Plan Apochromat 420782-9900-799

Stage

  • Motorized stage Marzhauser Scan IM 130x100 Serial 14032630
  • Remote control joystick Marzhauser  2-Schsen-Joystick CZ Serial 2420142128 Article 90-76-200-0820 Zeiss Serial 432903-9011-000
  • Inserts
    • Slide combo
    • 6-well plate
    • 35 mm dish
    • Multi-well plat

Filters

6-Position motorized Filter Turret 424947

  1. GFP Zeiss Filter Set 38 cube 424931 000000-1031-346 BP 470/40 450-490 FT 495 BP525/50 500-550
  2. DAPI Zeiss Filter Set 49 cube 424931 489064-0000-000 G365 FT 395 BP 445/50 335-383 395 420-470
  3. mPlum Zeiss Filter Set 64 HE cube 424931  489064-0000-000 BP587/25 FT605 BP647/70 574-599 605 612-682
  4. DIC Analyzer Zeiss 424937-9901
  5. Empty
  6. Empty

Scanner

LSM900 Scan head

  • 8x Motorized Filter wheel 1
  1. Empty
  2. SP 470 000000-2090-296 410-470 93% 480-750 0.001%
  3. SP 545 000000-2090-297 410-546 93% 557-750 0.001%
  4. SP 620 000000-2090-298 410-617 93% 630-750 0.001%
  5. Empty
  6. Empty
  7. Empty
  8. Empty
  • 8x Motorized Filter wheel 2
  1. Empty
  2. LBF 640 0000000-2090-299 410-617 93% 630-644 0.001%
  3. LP 575 0000000-2090-294 576-750 93% 410-565 0.001%
  4. LP 655 0000000-2090-295 656-750 93% 395-643 0.001%
  5. SP 620 000000-2090-298 410-617 93% 630-750 0.001%
  6. Empty
  7. Empty
  8. Empty

 Main Beam splitter

MBS 405+488+561+640 T10/R90

400102-9600-000

Transmission

  • 420-471 93%
  • 502-546 93%
  • 575-618 93%
  • 656-800 93%
  • 631-642 10%

Reflection

  • 399-410 99%
  • 481-492 99%
  • 558-564 99%
  • 631-642 90%

Detector

  • 2 GaASp PMT
  • 1 Airyscan 2 63x Serial 2657000116
  • 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
  • VNvidia RTX A4000 16 GB GDDR6
  • Monitor HP Z32x
  • Software Zen 3.10

Incubation

Consumables


Tabs Page
idFAQ
titleTroubleshooting & FAQ

Troubleshooting

UI Expand
titleI don't see any fluorescence!

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.

  1. Open the light path file
  2. Starting from the light source and moving towards the detector, verify that there is indeed light after each component of the microscope

FAQ

UI Expand
titleCan I use this microscope to observe cells in culture?
Yes it is an inverted microscope. Point scanning confocal induce damage so care should be taken while imaging live cells. a coverslip (thickness 0.17mm).




Tabs Container
idDemo Image
titleZeiss LSM-800
directionhorizontal


Tabs Page
idDemo Image
titleDemo Image




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