FAQ 1: How thick should I cut my tissue sections for the optical fractionator probe?

Post-processing tissue thickness is a critical consideration. Probes such as the Optical Fractionator require a tissue thickness that can be broken up optically by the focal point of a high numerical aperture objective. The sections must be thick enough to allow many focal planes through the disector and to allow for guard zones. Typically, tissue between 15-30 microns thick post-processing allows for the loss of upper and lower guard zones while still resulting in several optical Z-planes when using a 1.4 NA objective. Tissue sectioned between 30-60 microns usually gives usable results after staining and dehydration. You should do some experimentation towards developing a staining protocol that does not result in excessive shrinkage in the Z axis.

This FAQ comes from stereology.info, our curated collection of information and references regarding the use of stereology in the biological sciences. Feel free to review the other FAQs there as well. We hope this thread will spark some in-depth discussion on stereology!