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Include Page
_Micro-head-en
_Micro-head-en

Button Hyperlink
iconbuild
titleFrançais
typeprimary
urlhttps://wiki.umontreal.ca/display/Microscopie/Nikon + Ti2-EFura

Nikon Ti2-Fura microscope

Desmarais Building, Room 2236
Advanced Microscope Tier 1 usage price
Instrument awarded to the CI2B by the Canadian Foundation for Innovation (CFI #37439) in 2017
Advanced Microscope Tier 1 usage price

 

Applications

  • Inverted microscope
  • Widefield imaging
    • Brightfield

    • Fluorescence

  • Live imaging
  • Calcium ratiometric imaging
  • High-throughput imaging

Image Added

Tabs Container
directionhorizontal
Tabs Page
idDescription
titleDescription

Description

Light sources

  • LED lamp for transmitted light

  • CoolLed pE340-fura

    Expand
    titleComplete specifications
    ChannelSourceFilterFluorophoreNominal Power (mW)Measured Power (mW)
    1340nmBP340/20

    Fura

    ?

    		TBD
    2380nmBP380/20

    Fura

    		?

    TBD

    3435-645nm-
    		?

    TBD

    View file
    nameCoolLED_pE-340fura_User-Manual.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Quick Start Guidepdf.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Troubleshooting Guide.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Diagram.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Datasheet.pdf
    height250

  • Lumencor Spectra

    Expand
    titleComplete specifications
    ChannelSourceFilterBandwithFluorophoreNominal Power (mW)Measured Power (mW)
    Violet

    390

    390/22

    [379-401]

    DAPI, Hoechst, coumarins

    		236
    Blue

    437

    		437/26
CFP
[424-450]CFP, Alexa Fluor 430, Pacific Blue, BFP301
Cyan
473

  • 475

473/28
485/20[475-495]GFP, FITC, Alexa Fluor 488, Oregon Green, SYBR Green
FITC, GFP
  • 179
    Teal

    512

    		512/18[503-521]YFP, Alexa Fluor 514, SYTO dyes, Rhodamine 12360.6
    Green/Yellow542

    542/30

TRITC, Cy3

  • [527-557]

    Rhodamine, Alexa Fluor 546, 555, Cy3, TRITC

    		355

    Green/Yellow

    		575575/30

    [560-590]

    mCherry, DsRed, Alexa Fluor 568, Texas Red

    		293
    Red

    631

    		631/28

    [617-645]

    Alexa Fluor 633, Cy5, ATTO 647, mPlum, Alexa Fluor 647

    		243

    View file
    nameLumencor_Spectra_User-Manual.pdf
    height250
    View file
    nameLumencor_Specra_Certificate of Conformance.pdf
    height250
    Image Modified
    View file
    nameLumencor_Spectra X_Manual.pdf
    height250

Objectives

  1. 4x/0.2 Air
  2. 10x/0.45 Air
  3. 20x/0.75 Air
  4. 40x/0.75 Air
  5. 60x/1.4 OilEmpty
  6. -
    Expand
    titleComplete specifications
    PositionNameBrandFull nameIdentifierWorking distance (mm)Transmittance
    (% [nm])    		TechniquesCover glass thickness (mm)
    14x/0.2
    Air    		Nikon

    4x/0.2 Air
    CFI Plan Apochromat Lambda

    MRD00045

    		20.0
>80% [400-1000]

  		1. BF, Fluo0.17
    210x/0.45
    Air    		Nikon

    10x/0.45 Air
    CFI Plan Apochromat Lambda

    		MRD00105

    4.0

    		 BF, DIC N1, Fluo0.17
    320x/0.75
    Air    		Nikon20x/0.75 Air
    CFI Plan Apochromat Lambda    		MRD00205

    1.0

>80% [400-950]


  		1. BF
, Pol
  1. , DIC N2, Fluo

DIC N2
  1. 0.17
    440x/0.75
    Air    		Nikon40x/0.75 Air
    Plan Fluor     		MRH00400

    0.72

    		 
    		0.17
    5

    60x/1.4
    Oil

    		Nikon60x/1.4 Oil
    CFI Plan Apochromat Lambda    		MRD016050.13
>80% [475-725]

  		1. BF
, Pol
  1. , DIC N2, Fluo
DIC N2
  1. 0.17
    6

  1. -

    		----- --

Empty

 

Filters

  1. DAPI

  2. GFP

  3. Cy3

  4. Cy5
  5. Fura
  6. EmptyQuadBand DAPI/FITC/TRITC/Cy5
Expand
titleComplete specifications
PositionNameBrandIDExcitation FilterDichroic mirrorEmission FilterComments
1DAPISemrock96370 M352024-409LP447/60Semrock Brightline 96370 M352024 DAPI-3060A
No excitation filter
2GFPSemrockGFP-4050B

466/40x

495LP

525/50mSemrock Brightline 96372 M380163-17 GFP-4050B
3Cy3Semrock

LED-TRITC-A

554/23573LP609/54Semrock Brightline 96374 M380162-10 LED-TRITC-A
4Cy5Semrock

Cy5-5070A

618/50652LP698/70

Semrock Brightline 96376 M351081 23 Cy5-5070A

5FuraChroma

79001-ET-Fura-2

-400LP510/80

Chroma Fura 77074017 79001-ET-Fura2 C189116
Smaller cube (not 32mm). No Excitation Filter.

6
Empty
QuadBand
-
Semrock

-

----

Detector

  • Photometrics Prime 95B 25mm 

Expand
titleComplete specifications

Camera

Photometrics Prime 95B 25mm

Sensor Type

Back-illuminated sCMOS

Sensor Category

Monochrome

Nb Pixels

2.6 M

Pixel Layout

1608 x 1608

Pixel size

11.0 um

Sensor size

17.7 mm x 17.7 mm 

Sensor diagonal

25 mm

Bit depth

16-bit
Speed at full resolution30 images/s

Max QE

95 %
Readout noise1.6 e⁻

Cooling

-20°C Air

Dark Current

0.55 e⁻/pixel/sec

Full well capacity

80 000 e-

Dynamic Range

1: 50 000

Interface

USB 3.0

Mount

F-mount

Image Removed

View file
namePhotometrics_Prime-95B_Datasheet.pdf
height250
View file
namePhotometrics_Prime-95B_Manual.pdf
height250

Whitin Spectra

390/22 [379-401]
485/20 [475-495] 
540/30 [527-557]
631/28 [617-645]

FF409/493/573/652-Di02
Reflection - Transmission
350-404nm - 414-450nm
461-487nm - 499-530nm
543-566nm - 580-611nm
626-644nm - 659-850nm		F01-432/515/595/730-32 
432/36 [414-450]
515/30 [499-530] 
595/31 [580-611]
730/139 [661-800]

DAPI excitation can be improved by 375/40
FITC excitation should be replaced 474/24
TRITC excitation should be replaced with 554/22
Cy5 excitation filter can be improved with a 635/18


Detector

  • Photometrics Prime 95B 25mm 

Expand
titleComplete specifications

Camera

Photometrics Prime 95B 25mm

Sensor Type

Back-illuminated sCMOS

Sensor Category

Monochrome

Nb Pixels

2.6 M

Pixel Layout

1608 x 1608

Pixel size

11.0 um

Sensor size

17.7 mm x 17.7 mm 

Sensor diagonal

25 mm

Bit depth

16-bit
Speed at full resolution30 images/s

Max QE

95 %
Readout noise1.6 e⁻

Cooling

-20°C Air

Dark Current

0.55 e⁻/pixel/sec

Full well capacity

80 000 e-

Dynamic Range

1: 50 000

Interface

USB 3.0

Mount

F-mount

Image Added

View file
namePhotometrics_Prime-95B_Datasheet.pdf
height250
View file
namePhotometrics_Prime-95B_Manual.pdf
height250

  • If not already done, turn on the computer (#1) and log in to Windows using your UdeM credentials
  • Remove the dust cover from the microscope
  • Turn on the microscope power bar (#2) on the desk between the microscope and the computer
  • When using the instrument for the first time, it is necessary to import the microscope configuration into the software. See the First Use section below before starting the software
    • Start NIS-Elements

    Tabs Page
    titleUser Guide

    User Guide

    Tabs Page
    titleUser Guide
    UI Expand
    expandedtrue
    titleStart-up
    UI Expand
    expandedtrue
    titleFirst UseStart-up
    1. If not already done, turn on the computer (#1) and log in to Windows using your UdeM credentials
    2. Remove the dust cover from the microscope
    3. Turn on the microscope power bar (#2) on the desk between the microscope and the computer
    4. When using the
    When using the
    1. instrument for the first time, it is necessary to import the microscope configuration into the software. See the First Use section below before starting the software
    2. Start NIS-Elements
    UI Expand
    titleFirst Use

    When using the instrument for the first time, it is necessary to import the microscope configuration into the software. This procedure is usually carried out during the training session. However, This procedure is usually carried out during the training session. However, it can also be used to reset the software if it does not display correctly, for example. 

    Note

    Running this procedure will erase all your experiment protocols and reset the software to its original settings. If you are not sure, ask for support.

    1. If NIS-Elements is open, close it and wait until it has completely shut down (this may take up to 30 seconds)

    2. On the Desktop, open the Softwares folder

    3. Open NIS Settings Utility
    4. Click on the Import tab
    5. Click on Browse
    6. Navigate to your C:\Users\Public\Documents
    7. Select the file Nikon_Ti2-Fura_NIS Settings.bin
    8. Click Select
    9. Select all items
    10. Click Import
    11. Click OK
    12. Close the NIS Settings Utility
    13. You can now reopen NIS-Elements
    UI Expand
    titleLoading samples

    During this procedure, you will:

    • Set the microscope in a safe configuration

    • Load your sample

    • Find and adjust the focus

    Once completed, your sample will be ready for acquisition.

    UI Expand
    titleInitial Focus

    1. On the microscope, gently push the transmitted light arm backward

    2. In NIS-Elements, click Escape Z to move the objectives to a safe position

    3. If not done already, click on the lowest magnification objective

      Info
      The lowest magnification is the safest to use due to its long working distance: the sample will appear in focus well before the objective lens gets close to it. It is recommended to always perform the initial focusing with the lowest magnification objective. Since the objectives are parafocal, focusing with le safest objective will make it easier to locate the sample when switching to higher magnification objectives

    4. Place the test slide on the microscope stage, with the coverslip toward the objective

      Tip

      Using a test slide will significantly reduce the time needed to set up the instrument

    5. If necessary, use the joystick to move the stage so the sample is centered under the objective

    6. Gently return the transmitted light arm to the vertical position

    7. In NIS-Elements, click Escape Z again to return the objectives to their normal position

    8. Click on the desired optical configuration (BF, DAPI, GFP, etc.)

    9. Click Live (») to activate the light and display the camera image

    10. Adjust the light intensity and exposure time to obtain a well-exposed image

    11. Adjust the focus until the image is perfectly sharp.

    12. Click Stop to stop the Live or on Off to turn off the light
    Tip

    The focus is around Z = 2100 µm. The Z-position value is displayed: 

    • On the microscope remote control: Use the Display button to navigate the display menu
    • In NIS Elements: Ti2 Pad Tab under Z value
    UI Expand
    titleScanning an overviewCreate a preview

    It is possible to scan an overview create a preview image to then easily navigate your sample. It is strongly recommended to use the lowest magnification objective and whenever possible use the BF optical configuration not to bleach your sample.

    After completing the initial focus:

    1. In NIS Elements, click on the desired optical configuration (BF, DAPI, GFP, etc.)

    2. Click Live (») to activate the illumination and display the camera image on the screen

    3. Adjust the light intensity and exposure time to obtain a properly exposed image

    4. Using the Joystick navigate to the top left corner of your sample
    5. In the the XYZ overview click + to add a point and save this position
    6. Using the Joystick navigate to the bottom right corner of your sample
    7. In the the XYZ overview click + to add a point and save this position
    8. In the XYZ Overview right click and under Large Image Area select Define Area
    9. Draw a rectangle around the two points
    10. Right click again on the created region of interest and select scan preview
    11. Wait until the preview acquisition is completed

    You can now directly click on the overview to navigate your sample !

    UI Expand
    titleSecondary focus
    Warning

    First perform the initial focusing with the safest objective before selecting a higher-magnification objective

    UI Expand
    titleFocusing with air objectives

    After completing the initial focus:

    1. In NIS-Elements, click on the desired air objective (10x, 20x, 40x)

      Info

      The 20x is the best air objective because it has the highest numerical aperture (0.75).

    2. Click on the desired optical configuration (BF, DAPI, GFP, etc.)

    3. Click Live (») to activate the illumination and display the camera image on the screen

    4. Adjust the light intensity and exposure time to obtain a properly exposed image

    5. Adjust the focus using the precision knob until the image is perfectly sharp

    6. Click Stop to stop the Live mode or click on the optical configuration Off to turn off the illumination

    7. Click Escape Z to move the objectives to a safe position

    8. On the microscope, you can remove the test slide and install your sample

    9. In NIS-Elements, click Escape Z once again to return the objectives to their normal position.

    Your sample is now ready for acquisition!

    UI Expand
    titleFocusing with oil objectives

    After completing the initial focusing:

    1. In NIS-Elements, click Escape Z to move the objectives to a safe position

    2. Click on the desired immersion objective (60x)

      Info

      The 60x is the best immersion lens because it has the highest numerical aperture (1.4).

    1. Remove the test slide

    2. Place a single drop of oil on the objective

    3. Place your sample with the coverslip facing toward the objective

    4. Click Escape Z again to return the objectives to their normal position

    5. Click on the desired optical configuration (BF, DAPI, GFP, etc.)

    6. Click Live (») to activate the illumination and display the camera image on the screen

    7. Adjust the light intensity and exposure time to obtain a properly exposed image

    8. Adjust the focus using the precision knob until the image is perfectly sharp

    9. Click Stop to stop the Live mode or click on the optical configuration Off to turn off the illumination

    Your sample is now 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 NIS-Elements
    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 objectives with lens cleaner and paper
    5. Select the lowest magnification objective and click Escape Z to place the objectives in a safe position

    6. Wait until the SpectaX fan is off and then turn off the microscope power bar (#2)

    7. Turn off the computer
    8. Cover the instrument with the protective dust cover
    Note
    • Take back your samples including ones in the microscope
    • Leave the microscope and the working area clean
    Tabs Page
    title
    idtitleLog

    Log

    UI Expand
    expandedtrue
    titleTo do
    • Fix camera adapter
    • Purchase multiband pass cube for Spectra
    • Fix stage inserts Ti2 S HU
    • Replace LLG Spectra
    • Add HCA Jobs and Plates user guide
    UI Expand
    title2026-02-24 Installation multiwell insert
    • Purchase Multiwell plate insert
      • Replace LLG Spectra
    • TI2-S-HW

    Image Added

    UI Expand
    title2025-10-09 Installation Spectra
    • Installation Spectra
    • Installation LAPP
    UI Expand
    title2025-03-10 Insallation Desmarais 2236
    • Complete installation
    • Complete cleaning
    Tabs Page
    idtitleTechnical Datasheettitle

    Technical Datasheet

    Stand

    • Nikon Ti2-E inverted Serial 540749
    • Pillar for transmitted illumination Ti2-D-PD Serial 599941
    • Manual condenser Turret TC-C-TC
    • Perfect Focus Unit with motorized nosepiece Ti2-N-ND-P Serial 128285
    • 6 positions motorized epi filter turret Ti2-F-FLT-E Serial 510911
    • EPI-FL module TI2-LA-FLL Serial 547589
    • Ti2-LAPP System Ti2-LA-BM-E Serial 548536
    • Camera adapter Ti2-BDTV2
    • Ti2 Controller Ti2-CTRE Serial 451316

    Light sources

    • LED Lamphouse for dia illumination Ti2-D-LHLED Serial 557988
    • CoolLED pE340-Fura Serial AY1035
    • Lumencor Spectra7-API-IB Serial 3719 with LLG adapter 82-10012 Rev B Applied precision part number 34-100926-000

    Condenser

    • LWD Condenser lens (O.D.=30 mm, NA=0.52)

    • Filter turret 7 manual positions

      1. Neutral Density ND

      2. Empty
      3. Empty
      4. Empty
      5. Empty

    Objectives

    • 4x/0.2 MRD00045
    • 10x/0.45 MRD00105
    • 20x/0.75 MRD00205
    • 40x/0.75MRH00400
    • 60x/1.4 MRD01605
    • Empty

    Stage

    • Motorized stage Encoded Ti2-S-SE-E Serial 960166
    • Remote control joystick Ti2-S-JS Serial 128408
    • Inserts
      • Combo slide 3cm dish Ti2-S-HU with tilt adjustment (no incubation)

    Filters

    1. DAPI Cube Semrock Brightline 96370 M352024 1 No excitation filter
    2. GFP Cube Semrock Brightline 96372 M380163-17 GFP-4050B Ex 466/40 Em 525/50 Di 495
    3. Cy3 Cube Semrock Brightline 96374 M380162-10 LED-TRITC-A Ex 554/23 Em 609/54 Di 573
    4. Cy5 Cube Semrock Brightline 96376 M351081 23
    5. Fura77074017 79001 ET Fura2 C189116 (smaller cube not 25mm) No Excitation Filter
    6. Empty

    Detector

    • Photometric Prime95B 25mm Serial A18C203010

    Workstation

    • HP Z440 Workstation
    • I155439-CIB
    • Intel Xeon E5-1620 v4 @ 3.5GHz
    • Motherboard HP 212B Chipset Intel C612
    • BIOS M60 V02.61 2023-03-23
    • RAM 32 GB DDR4 1200 MHz ECC (4 x 8 GB)
    • OS 1 TB SSD 530 MB/s
    • 4 TB HD Data Storage (2 x 2 TB spanned volume) 212 MB/s
    • Video Card nVidia Quadro P4000 8GB GDDR5 5.3 TFLOPS FP32
    • Monitor HP Z27n display 27' 2560x1440
    • Software NIS-Elements AR v5.2 HASP 53A125C5  NIS Elements AR
      • ND (6 dimensions)
      • Quick Piezo Z
      • Quick MultiExcitation
      • Shutter
      •  Wavelength Switcher
      • HC
      • JOBS
      • General Analysis 
      • JOBS View
      • Remote DB for JOBS
      • General Analysis 3

    Consumables

    • Liquid Light Guide

    Manuals

    Anti-vibration Table

    • -LA-BM-E Serial 548536
    • Camera adapter Ti2-BDTV2
    • Ti2 Controller Ti2-CTRE Serial 451316

    Light sources

    • LED Lamphouse for dia illumination Ti2-D-LHLED Serial 557988
    • CoolLED pE340-Fura Serial AY1035
    • Lumencor Spectra7-API-IB Serial 3719 with LLG adapter 82-10012 Rev B Applied precision part number 34-100926-000

    Condenser

    • LWD Condenser lens (O.D.=30 mm, NA=0.52)

    • Filter turret 7 manual positions

      1. Neutral Density ND

      2. Empty
      3. Empty
      4. Empty
      5. Empty

    Objectives

    • 4x/0.2 MRD00045
    • 10x/0.45 MRD00105
    • 20x/0.75 MRD00205
    • 40x/0.75MRH00400
    • 60x/1.4 MRD01605
    • Empty

    Stage

    • Motorized stage Encoded Ti2-S-SE-E Serial 960166
    • Remote control joystick Ti2-S-JS Serial 128408
    • Inserts
      • Combo slide 3cm dish Ti2-S-HU with tilt adjustment (no incubation)

    Filters

    1. DAPI Cube Semrock Brightline 96370 M352024 1 No excitation filter
    2. GFP Cube Semrock Brightline 96372 M380163-17 GFP-4050B Ex 466/40 Em 525/50 Di 495
    3. Cy3 Cube Semrock Brightline 96374 M380162-10 LED-TRITC-A Ex 554/23 Em 609/54 Di 573
    4. Cy5 Cube Semrock Brightline 96376 M351081 23
    5. Fura77074017 79001 ET Fura2 C189116 (smaller cube not 25mm) No Excitation Filter
    6. Empty

    Detector

    • Photometric Prime95B 25mm Serial A18C203010

    Workstation

    • HP Z440 Workstation
    • I155439-CIB
    • Intel Xeon E5-1620 v4 @ 3.5GHz
    • Motherboard HP 212B Chipset Intel C612
    • BIOS M60 V02.61 2023-03-23
    • RAM 32 GB DDR4 1200 MHz ECC (4 x 8 GB)
    • OS 1 TB NVMe 5500 MB/s via PCie x4 to M2 Adapter Startech PEX4M2E
    • 4 TB HD Data Storage (2 x 2 TB spanned volume) 212 MB/s
    • Video Card nVidia Quadro P4000 8GB GDDR5 5.3 TFLOPS FP32
    • Monitor HP Z27n display 27' 2560x1440
    • Software NIS-Elements AR v5.2 HASP 53A125C5  NIS Elements AR
      • ND (6 dimensions)
      • Quick Piezo Z
      • Quick MultiExcitation
      • Shutter
      • Wavelength Switcher
      • Hight content
      • JOBS
      • General Analysis 
      • JOBS View
      • Remote DB for JOBS
      • General Analysis 3

    Consumables

    • Liquid Light Guide

    Manuals

    View file
    nameLumencor_Spectra X_Manual.pdf
    height250
    View file
    nameLumencor_Specra_Certificate of Conformance.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Datasheet.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Diagram.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Troubleshooting Guide.pdf
    height250
    View file
    nameCoolLED_pE-340fura_Quick Start Guidepdf.pdf
    height250
    Image AddedImage Added
    View file
    nameCoolLED_pE-340fura_Brochure.pdf
    height250
    View file
    nameCoolLED_pE-340fura_User-Manual.pdf
    height250
    View file
    namePhotometrics_Prime-95B_Manual.pdf
    height250
    View file
    namePhotometrics_Prime-95B_Datasheet.pdf
    height250
    View file
    nameNational Instruments_NIDAQ_Triggering Guide for NIS-Elements.pdf
    height250

    Anti-vibration Table

    • TMC #63-42352-03 Serial 218043
    FAQ
    Tabs Page
    Tabs Page
    id
    titleTroubleshooting & FAQ

    Troubleshooting

    UI Expand
    titleCamera Driver Failed

    This can happen if the acquisition software is turned on before the camera has fully initialized. The software should be started after the camera has fully initialized.

    • Turn off NIS-Elements
    • Ensure the Initialize light at the back of the camera no longer blinks
    • Start NIS-Elements

    FAQ

    UI Expand
    titleCan I use this microscope to look at cell in a dish?
    • Yes. 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
    • For long timelapse, be aware of photo-toxicity

    Include Page
    Plateformes:_Plat-foot-en
    Plateformes:_Plat-foot-en