Setup Extensions and Modifications
Michael Redd from the Cell Imaging Core Facility at the University of Utah put together an Arduino-based solution to get proper bright-field illumination in an OpenSPIM setup: Brightfield.
OpenSPIM for large samples
Field of view
The original OpenSPIM uses a 2X expander to enlarge the laser beam to 2 mm. For larger samples a larger field of view can be preferable. This means that you will need a larger beam to create a taller light sheet. Change the second beam expander lens to f=75 mm or 100 mm and increase the distance between lenses accordingly.
Immersion objectives with long working distance are hard to find. However, at low NA and moderate magnification, a dry objective will also work; the higher NA, the more spherical aberrations will be present. Keep in mind that when using a dry objective in immersion, the WD will increase (approximately by a factor equal to the RI of the new medium, eg. 17 mm in air will increase to 22.6 mm in water).
Example parts list
The table below lists the parts used by the OpenSPIM for brains setup built by Monika Pawłowska (Nencki Institute of Experimental Biology, Warsaw).
|Manufacturer||Accessibility||Description||File or Link/Model #||Image||Quantity||Price (EUR)|
|Nikon||purchase||4X Nikon Plan Fluorite Imaging Objective, 0.13 NA, 17.2 mm WD, dry||N4X-PF||2 (or 3 for double illumination)||410|
|Thorlabs||purchase||Translating Lens Mount for Ø1" Optics (for the objectives||LM1XY/M||2 (or 3)||125|
|Nikon||purchase||Infinity-Corrected Tube Lens for Plan Fluorite Objectives||ITL200||1||405|
|Monika||self made||This chamber can be 3D printed. For windows use 24x24 mm microscopy cover glasses and glue (eg. two component epoxy)||.stl on Github||1|
|Monika||self made||This holder includes two threaded holes so it's easiest to make from metal (eg aluminum). Holder is mounted on the 4D stage arm with M6 screw. Glass slide with 1 mm thickness can be held with nylon or nylon-tipped screw||drawing on Github step file on Github||1|