Bioimaging Wiki

Microscope overview

The 3i Marianas SDC spinning disk confocal microscope is a fast imaging system which provides a combination of confocality, resolution and speed, well suited for the observation of dynamics in living cells and small organisms. The Marianas SDC provides fast acquisition (up to ~100fps, with limited ROI, ~30FPS full frame), and with lower phototoxicity and bleaching. A spinning disk with multiple small pinholes is installed between the light source and specimen to generate point-like illumination covering the whole area “simultaneously”. Each small aperture serves as a detecting pinhole to remove out-of-focus light. This system was built to be versatile, and allow imaging of different sample sizes. It includes both high and low magnification objectives (which require different pinhole sizes, 25 or 50um), DIC, a removable heating stage with CO2 delivery, and photomanipulation modules (Phasor and Ablate!). The scanhead allows 1x or 2x magnification (for extra magnification and to allow Nyquist sampling and deconvolution).

Microscope: Nikon Ti2 Eclipse
Confocal scanner: Yokogawa CSU-W1 + Uniformizer
Camera: Photometrics Prime 95B

Data files older than 1 month will be automatically deleted on this system, please copy your data to the GIMM server using the desktop link.

System components

Lasers

Laser line	Wavelength	Maximum Power
405	405 nm	100 mW
445	445 nm	150 mW
488	488 nm	150 mW
561	561 nm	100 mW
594	594 nm	100 mW
637	637 nm	140 mW
Ablate!	355 nm	60 μJ/200 Hz

Emission Filters CSU W1 unit

Position	Filter name	Transmission (nm)	Dichroic	Reference
1	Quad (c###q)	440/40 521/21 607/34 700/45	405/488/561/640	Semrock FF01-440/521/607/700
2	c405s	445/45	405/488/561/640	Semrock FF01-445/45
3	c488s	525/30	405/488/561/640	Semrock FF01-525/30
4	c561s	617/73	405/488/561/640	Semrock FF02-617/73
5	c594s	642/80	445/515/594	Chroma ET642/80
6	c637s	692/40	405/488/561/640	Semrock FF01-692/40
7	Dual (c###d)	525 642	405/488/561/640	Semrock EM01-R488/568
8	c445s	482/25	445/515/594	Semrock FF01-482/25
9	c488/YFP	542/27	405/488/561/640	Semrock FF01-542/27
10	cBF-DIC	-	any	-

Nikon Ti2 filterset cubes

Turret Position	Filter cube	Excitation	Dichroic	Emission
4 (lower deck)	DAPI	378/52	409	BP 447/60
2 (lower deck)	GFP	472/30	495	BP 520/35
3 (lower deck)	Texas Red	562/40	593	BP 624/40
6 (lower deck)	CSU (Confocal)	-	empty	-
3 (upper deck)	Phasor/Ablate	-	<380	-

Objectives

Magnification	Model	Immersion	NA	WD (mm)	Reference
10x	Plan Apochromat	Air	0.45	4	MRD70170
20x	Plan Fluor	Oil, W, G, Sil	0.75	0.51–0.33	MRH07241
40x	Plan Apochromat	Air	0.95	0.21	MRD70470
40x	Plan Apochromat	Silicone Oil	1.25	0.3	MRD73400
60x	Plan-Apochromat	Water	1.20	0.31–0.28	MRD07602
100x	Plan Apochromat	Oil	1.45	0.13	MRD71970

Upon request:

Magnification	Model	Immersion	NA	WD (mm)	Reference
40x	Apochromat	Water	1.25	0.2–0.16	MRD77400

Camera

Model	Frame Size	Pixel Size (µm)	Quantum Efficiency
Photometrics Prime 95B	1200 x 1200	11 x 11	> 95%

Microscope Turn On Procedure

Turn on the master power bar (left side of the table)

Turn the laser key to the On position

If your using temperature and/or CO2, turn on OkoLab power bar (left ide of the table)

After 2 minutes, turn on PC (left side of the table)

Login in to Windows with your Agendo credentials
Wait 2 min to allow for system components detection
Start the SlideBook software

Microscope Turn Off Procedure

If there is another user for this microscope during the day:

Exit the SlideBook software, leave the microscope on
Log off the computer
Clean up immersion objectives

Else:

Exit the SlideBook software
Shut down the computer
Turn off incubation/CO2 on the powerbar (if used)
Switch laser key off
Turn off the master power bar

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