Epalinges Instruments

Acquisition systems

Confocal

Confocal microscopy, most frequently confocal laser scanning microscopy (CLSM) or laser confocal scanning microscopy (LCSM), is an optical imaging technique for increasing optical resolution and contrast of a micrograph by means of using a spatial pinhole to block out-of-focus light in image formation.^[1] Capturing multiple two-dimensional images at different depths in a sample enables the reconstruction of three-dimensional structures (a process known as optical sectioning) within an object.

Zeiss LSM 800

Stage	Zeiss AxioObserver, Inverted
Illumination	HXP 120V
Lasers	405nm 488nm 561nm 640nm
Objectives	Plan Apochromat 10x / 0.45 NA DIC II Plan Apochromat 20x / 0.80 DIC II Plan Apochromat 40x / 1.30 NA / oil immersion DIC II Plan Apochromat 63x /1.40 NA / oil immersion DIC II
Stage	Motorized XY
Contrast	DIC
CO2 \| T° control	No
Detector type	3x GaAsP
Optional modules	Multiposition and Tiling
Software	Zen Blue 2.6
Room	CLF-212a

Zeiss LSM 880 with Airyscan

Stage	Zeiss AxioObserver Z1, Inverted
Illumination	HXP 120V
Lasers	405nm 458nm, 488nm, 514nm 561nm 640nm
Objectives	EC Plan Neofluar 10x/0.3 NA Ph1 DIC I EC Plan Neofluar 20x/0.5 NA Ph2 DIC II EC Plan Neofluar 40x/1.3 NA oil immersion DIC III Plan Neofluar 63x/1.25 NA oil immersion DIC III EC Plan Neofluar 100x/1.3 NA oil immersion DIC III
Stage	Motorized XY
Contrast	Phase (10x & 20x)
CO2 \| T° control	No \| No
Detector type	GaAsP detectors Airyscan add-on (enhanced resolution and SNR)
Optional modules	Multiposition (Mark and Find) Time-Lapse
Software	Zen Black
Room	CLF-212a

Leica SP5

Stage	Leica DM 5000, Upright
Illumination	HXP 120V
Lasers	405nm 458nm, 477nm, 488nm, 514nm 543nm 633nm
Objectives	HCX PL FLUOTAR 10x/0.30. HC PL APO 20x/0.70 imm corr. HCX PL APO CS 40x/ 1.25-0.75 Oil. HCX PL APO 63x/1.40-0.60 Oil. HCX PL APO 100x/1.40-0.70 Oil CS.
Stage	Motorized XY Galvo Z
Contrast	DIC
CO2 \| T° control	No
Detector type	3x PMT Resonnant scanner
Optional modules	Multiposition and Tiling
Software	LAS AF
Room	CLF-213a

Zeiss LSM 800 P2 Lab [SPECIAL AUTHORIZATION REQUIRED]

This particular setup being placed in a P2 Lab, a special authorization is required to access it. Contact the technical manager for further information

Stage	Zeiss AxioObserver, Inverted
Illumination	HXP 120V
Lasers	405nm 488nm 561nm 640nm
Objectives	Plan Apochromat 10x / 0.45 NA DIC II Plan Apochromat 20x / 0.80 DIC II Plan Apochromat 40x / 0.95 DIC III Plan Apochromat 40x / 1.30 NA / oil immersion DIC II Plan Apochromat 63x /1.40 NA / oil immersion DIC II
Stage	Motorized XY Definite Focus
Contrast	DIC
CO2 \| T° control	Yes Full enclosure
Detector type	3x GaAsP
Optional modules	Multiposition and Tiling
Software	Zen Blue 2.6
Room	P2 Lab (F, 4th floor)

Slide Scanner

A slide scanner allows you to digitize full microscopy slides in an automated manner. This is a high throughput system able to process around a hundred of slides in one batch.

The NanoZoomer S60 slide-scanner can work as an intuitive microscope designed for slides or as a regular slide-scanner. The way a slide scanner works implies that designing a specific profile for each of your sample type is needed.
This profile is very dependent of the way you have prepared your slides. Since this processe is time-consuming, it is only useful if you have at least a dozen or more slides of the exact same type to acquire (1-2 hours with testing).

Here are some general guidelines before thinking of using the slide scanner for your experiments:

*SCAN PROFILE*
*If the samples vary in any one of these categories:* Thickness of the slices Layout of the cuts Size of the cuts Intensity / color of the staining *Fluorescence specific:* Number of fluorescent dyes target of the staining used as reference *The profile will have to be updated* and saved as a new variant or it will probably fail (not focused, over/underexposed, stitching failing).
*ACQUISITION TIME*
*The acquisition time depends on:* Number of sample / blade Sample size / blade Lens used Exposure time (not important in BF) Z Stack or not and EDF(extend focus) or not Accuracy of the focus map Each of these parameters can greatly vary the acquisition time, which can range from a few minutes to an hour or more per slide. If you want to change the objective used for the acquisition, you will also need to update most of the previous parameters of the scan profile.

SCAN PROFILE

If the samples vary in any one of these categories:

Thickness of the slices
Layout of the cuts
Size of the cuts
Intensity / color of the staining

Fluorescence specific:

Number of fluorescent dyes
target of the staining used as reference

The profile will have to be updated and saved as a new variant or it will probably fail (not focused, over/underexposed, stitching failing).

ACQUISITION TIME

The acquisition time depends on:

Number of sample / blade
Sample size / blade
Lens used
Exposure time (not important in BF)
Z Stack or not and EDF(extend focus) or not
Accuracy of the focus map

Each of these parameters can greatly vary the acquisition time, which can range from a few minutes to an hour or more per slide.
If you want to change the objective used for the acquisition, you will also need to update most of the previous parameters of the scan profile.

Hamamatsu NanoZoomer S60

Brochure available here for more information : https://www.hamamatsu.com/resources/pdf/sys/SBIS0043E_NanoZoomers.pdf

Illumination	See filters below
Filters	DAPI FITC – AF488 Cy3 (or TxRed) Cy5 Cy7 – AF750 (unique at the CIF-Epalinges at the moment)
Objectives	The NanoZoomer S60 uses a 40x objective allowing for multiple magification through pixel binning.
Stage	Motorized
Contrast	Brightfield, Fluorescence
CO2 \| T° control	No
Cameras	Color camera Color CMOS camera Black and White Camera sCMOS Orca Flash LT Plus
Optional modules
Software	NDP.view2
Room	CLF-212

Widefield | Fluorescence

The majority of fluorescence microscopes, especially those used in the life sciences, are of the epifluorescence design shown in the diagram. Light of the excitation wavelength illuminates the specimen through the objective lens. The fluorescence emitted by the specimen is focused to the detector by the same objective that is used for the excitation which for greater resolution will need objective lens with higher numerical aperture. Since most of the excitation light is transmitted through the specimen, only reflected excitatory light reaches the objective together with the emitted light and the epifluorescence method therefore gives a high signal-to-noise ratio. The dichroic beamsplitter acts as a wavelength specific filter, transmitting fluoresced light through to the eyepiece or detector, but reflecting any remaining excitation light back towards the source.

Zeiss Axiovision Upright

Stage	Zeiss ImagerZ1, Upright
Illumination	HXP 120V
Filter Cubes	DAPI CFP GFP Rhodamin CY5 FRET YellowFP mCherry DsRed
Objectives	EC Plan Neofluar 10X, N.A. 0.3 DIC EC Plan Neofluar 20X, N.A. 0.5 DIC EC Plan Neofluar 40X, N.A. 1.3 Oil DIC Plan Neofluar 63X, N.A. 1.4 Oil DIC EC Plan Neofluar 100X, N.A. 1.3 Oil DIC
Stage	Motorized
Contrast	NC
CO2 \| T° control	No \| No
Cameras	Axiocam MRm Axiocam MRc5
Optional modules	Mosaix (Image stitching)
Software	Axiovision SE64 rel 4.9.1
Room	CLF-212

Zeiss Axiovision Upright B

Stand	Zeiss AxioImager Z1, Upright
Illumination	EBQ100 Isolated
Filters	DAPI BP YFP FITC – GFP Rhodamin – CY3 Alexa 660 CY5.5 DAPI LP
Objectives	EC Plan Neofluar 10x/0.3 NA DIC I EC Plan Neofluar 20x/0.5 NA DIC II EC Plan Neofluar 40x/0.75 NA DIC II EC Plan Neofluar 40x/1.3 NA oil immersion DIC III Plan Neofluar 63x/1.25 NA oil immersion DIC III EC Plan Neofluar 100x/1.3 NA oil immersion DIC III
Stage	Motorized XY
Contrast	DIC
CO2 \| T° control	No
Cameras	Color camera Zeiss Axiocam MRc Black and White Camera Zeiss Axiocam MRm
Optional modules	Mosaix (Tile Scan)
Software	Zeiss Axiovision rel 4.8
Room	CLF-213

Time-Lapse microscopy

Time-lapse microscopy is time-lapse photography applied to microscopy. Microscope image sequences are recorded and then viewed at a greater speed to give an accelerated view of the microscopic process. Before the introduction of the video tape recorder in the 1960s, time-lapse microscopy recordings were made on photographic film. During this period, time-lapse microscopy was referred to as microcinematography. With the increasing use of video recorders, the term time-lapse video microscopy was gradually adopted. Today, the term video is increasingly dropped, reflecting that a digital still camera is used to record the individual image frames, instead of a video recorder.

Zeiss Axiovision Inverted (Time-Lapse)

Stage	Zeiss AxioObserver Z1, Inverted
Illumination	HXP 120V
Filter Cubes	DAPI CFP GFP Rhodamin CY5
Objectives	EC Plan Neofluar 10X, N.A. 0.3 Ph1 EC Plan Neofluar 20X, N.A. 0.5 Ph2 EC Plan Neofluar 40X, N.A. 1.3 Oil DIC Plan Neofluar 63X, N.A. 1.25 Oil DIC EC Plan Neofluar 100X, N.A. 1.3 Oil DIC
Stage	Motorized
Contrast	Phase only
CO2 \| T° control	Yes [Full enclosure]
Cameras	Photometrics CoolSnap HQ2
Optional modules	Multi-position
Software	Axiovision 4.8
Room	CLF-213

Stereomicroscopy

The stereo, stereoscopic or dissecting microscope is an optical microscope variant designed for low magnification observation of a sample, typically using light reflected from the surface of an object rather than transmitted through it. The instrument uses two separate optical paths with two objectives and eyepieces to provide slightly different viewing angles to the left and right eyes. This arrangement produces a three-dimensional visualization of the sample being examined.^[1] Stereomicroscopy overlaps macrophotography for recording and examining solid samples with complex surface topography, where a three-dimensional view is needed for analyzing the detail.

Leica M205-FA

Stand	Leica M205-FA, Upright
Illumination	Transmitted light base RCI, 35W Incident LED illumination Fluorescence light source with fiber optics.
Filters	UV – DAPI GFP 2 – GFP TXR – CY3
Objectives	Plan Apochromat 1x Zoom 20.5x
Stage	Manual
Contrast	Brightfield
CO2 \| T° control	No
Cameras	Color camera Leica DFC300FX R2
Optional modules
Software	LAS AF 6000
Room	CLF-213

Light Sheet Microscopy

Light sheet fluorescence microscopy (LSFM) is a fluorescence microscopy technique with an intermediate-to-high^[1]optical resolution, but good optical sectioning capabilities and high speed. In contrast to epifluorescence microscopy only a thin slice (usually a few hundred nanometers to a few micrometers) of the sample is illuminated perpendicularly to the direction of observation. For illumination, a laser light-sheet is used, i.e. a laser beam which is focused only in one direction (e.g. using a cylindrical lens). A second method uses a circular beam scanned in one direction to create the lightsheet. As only the actually observed section is illuminated, this method reduces the photodamage and stress induced on a living sample. Also the good optical sectioning capability reduces the background signal and thus creates images with higher contrast, comparable to confocal microscopy. Because LSFM scans samples by using a plane of light instead of a point (as in confocal microscopy), it can acquire images at speeds 100 to 1000 times faster than those offered by point-scanning methods.

LaVision Biotec Ultramicroscope II

Laser-capture Microdissection

Laser-capture microdissection (LCM) is a method to procure subpopulations of tissue cells under direct microscopic visualization. LCM technology can harvest the cells of interest directly or can isolate specific cells by cutting away unwanted cells to give histologically pure enriched cell populations. A variety of downstream applications exist: DNA genotyping and loss-of-heterozygosity (LOH) analysis, RNA transcript profiling, cDNA library generation, proteomics discovery and signal-pathway profiling. The total time required to carry out this protocol is typically 1–1.5 h.^[3]

Acturus LCM

Stand	Nikon Eclipse Ti, Inverted
Widefield Illumination	X-Cite 120 PC
Filters	DAPI Alexa 488 Rhodamin
Lasers	Laser cutting Solid state UV Optical tweezer IR laser
Objectives	2x (Overview) 10x 20x 40x
Stage	Motorized XY
Contrast	DIC
CO2 \| T° control	No
Optional modules
Software	Arcturus XT
Room	CLF-213

Image Processing

[Under construction]