Lightsheet - MesoSPIM (Irchel)

The mesoSPIM (mesoscale selective plane illumination microscopy) is the ideal instrument to quickly bridge scales from the µm- to the cm-level, which enables it to serve as an excellent tool for detailed three-dimensional anatomical investigations in many biological and medical fields (Voigt et al., 2019, Vladimirov et al, 2023).

Main features (upgrade 2023):

• Compatibility with all clearing techniques: Clarity, iDISCO, CUBIC, PEGASOS, ECI, etc.
• Large field-of-view, ranging from 1.5 to 15 mm using a set of Mitutoyo Plan Apo objectives.
• Large-sensor camera Hamamatsu Orcal Lighting (29 mm diagonal)
• Dual-sided excitation. Optimized for fast screening of whole mouse brain samples, yielding relatively small datasets (10-14 GB/brain/color) near-isotropic sampling (2-4 µm in X,Y & Z) due to an axially swept lightsheet which leads to uniform z-resolution across the entire field-of-view.
• Modular sample holders and quick sample exchange.
• Large travel range (45 x 45 x 100 mm) to allow imaging of very large samples.

For further information: MesoSPIM.org

University Zurich, Irchel Campus, Room Y44-J-33.

Follow this link to apply for an introduction to the microscope.

Excitation path

The mesoSPIM excitation path is based on axially scanned light-sheet microscopy (ASLM). The waist of the light-sheet is translated through the sample in synchrony with the rolling shutter of the camera. The waist is translated with electrically tunable lenses by Optotune.

Light Sources and Lasers

The iChrome MLE comprises 3 diode lasers and 1 DPSS laser fully integrated in one compact housing.

Lasers

• 405nm (100mW)
• 488nm (100mW)
• 561nm (100mW)
• 640nm (100mW)

Detection path

The mesoSPIM detection path consists of five long-working distance air plan apochromat objectives:

1. Mitutoyo M Plan Apo 2x/0.055
2. Mitutoyo M Plan Apo 5x/0.14
3. Mitutoyo M Plan Apo 7.5x/0.21
4. Mitutoyo M Plan Apo 10x/0.28
5. Mitutoyo M Plan Apo 20x/0.28(t3.5) (corrected for 3.5 mm thick glass)

The microscope offers lateral (XY) resolution of 1.5-2.6 µm (depending on the objective) and axial (Z) resolution  of 3.5 µm for all objectives, across a large field of view thanks to new-generation sCMOS camera Hamamatsu Orca Lightning.

Emission (detection) filters

• Quadband filter 405/488/561/640
• 520/35
• 590/36

Camera

Hamamatsu Orca Lightning sCMOS:

1. image size: 4608 x 2592 px (12 MP)
2. sensor pixel size: 5.5 µm,
3. sensor dimensions (h,w): 25.34 x 14.25 mm (diagonal 29 mm).

Pixel size

The effective pixel size in acquired image depends on objective magnification:

Pixel_size(image) = Pixel_size(sensor)/Magnification

Field of view

Field of view can be calculated from the sensor size and objective magnifications by the formula:
FOV(h,w) = Sensor(h,w)/Magnification

Software

The microscope software, mesoSPIM-control, is developed by us. It is open-source and written in Python. It allows users to specify sequences of z-stacks using a table-based acquisition manager. The software can also be used to acquire large-scale tiling acquisitions.

Depending on your sample, we have a variety of holders and imaging cuvettes.

Examples of samples recently imaged with mesoSPIM

Image source: Vladimirov et al, 2023. The Benchtop mesoSPIM: a next-generation open-source light-sheet microscope for large cleared samples.

• For initial tests and the first experiments, the ZMB will serve users with the needed experimental accessories such as cuvettes and immersion solutions. If they become regular user of the microscope, they have to organize the accessories by themselves.

Literature and Links

Vladimirov et al, 2023. The Benchtop mesoSPIM: a next-generation open-source light-sheet microscope for large cleared samples.

Voigt et al., 2019. The mesoSPIM initiative: open-source light-sheet microscopes for imaging cleared tissue, Nature Methods 16, 1105-1108.

MesoSPIM.org

mesoSPIM-control

Further information (internal UZH use only)

Follow this link for further background information, documents and links.

Responsible Persons

If you have questions about the device please contact the responsible person.

Make sure to acknowledge the Center for Microscopy in your publication to support us.

How to acknowledge contributions of the Center for Microscopy