Illumination arm

Schemes

Part list

Alignment

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The first lens has to be placed at a distance f₁ from the galvo mirror, being f₁ the focal distance of the lens. The second lens is placed at a distance f₂+f₁ from the first lens. Finally the objective is placed at a distance f₂ from the second lens. In SPIM mode we will place the cylindrical lens between the second lens and the objective lens, at a distance f_cyl from the back focal aperture of the objective, which will still be placed at f₂ from the second lens in the beam expander.

Galvo mirror:

For alignment we will place on the optical bench every component, one by one, following the light pathway. Light coming from the laser is deflected 45º on the first mirror and directed to the 45º Mirror holder. This mirror sends light up to the galvo mirror. Be extremely careful at this point with the laser beam!!!

After the galvo mirror the beam must be parallel to the table at a certain height. This height will be determined by the height of the camera, so make sure that the camera is mounted on the rail, and the beam hits the center of the CCD field of view once is paralel to the table. To control the beam angle we can use the two mirrors (first mirror and 45º mirror mount) and the galvo position. This process can take some time. But patience will be rewarded.

Beam expander:

Then we set the beam expander. The best is to mount the whole piece and make sure that the beam keeps going at the same height and direction that previously. Because the lenses could be easily removed and set back this can be checked as many times as needed. Remember to keep the right distance between the galvo mirror and the first lens!!!.

For the central part of the beam expander we use the SM1E60 lens tube, hold by the tube slip (SM1RC/M). Different tubes are screwed on both side depending on the desired beam magnification but keeping the distances between lenses to 225 mm. The four lenses are inserted on four different SM1E05 lens tubes. The rest of the tubes are used to keep the aforementioned distance between lenses.

Two combinations are possible in our design:

3.5x:

8x:

Objective holder:

The objective holder is mounted on two perpendicular translational stages. This system is really useful to fine adjustment of the light-sheet and to the replacement of the objectives.

For SPIM mode the cylindrical lens is mounted on a SM1E05 lens tube and attached to the objective holder (in grey color in the figure). In DSLM mode we replace this lens tube for an empty one. In that way we keep constant distance between the second lens of the beam expander and the back focal aperture of the objective. Then we place a SM1E10, a SM1E03 lens tubes and a SM1A12 ring to keep 5 cm from the cylindrical lens. To find the conjugate plane of the galvo, we make it move slowly. The position where the beam is steady is the plane where the back focal aperture of the objective must be placed. Setting the galvo a 0 V, the beam should be parallel to the optical bench and following the holes of the table straight. If this is not the case we can make adjustments with the KM100T mount.

Shutter:

Finally, when everything is aligned, we will introduce the shutter, and optionally two diaphragms pinholes before and after the beam expander. This will make easier to detect misalignments and correct them.