Technology

knifeprism

3Scan’s core technology centers around what is formally named the “Knife Edge Scanning Microscope” (KESM).  Our microscope falls into the category of a serial sectioning microscope, bringing together elements of machine vision, inspection line optics, and microtomography.

What makes our microscope truly unique is the capability to do a relatively large volume of tissue with incredibly high throughput and great resolution.  Currently our prototype is specified to do a cubic centimeter of tissue in 7 hours with a 2 micron resolution, with a maximum volume of ~25 cubic centimeters.  Such capacity greatly fills the gap in the field which exists between high resolution, low volume, 3d techniques such as confocal microscopy or STED microscopy (maximum depth of ~3 microns) and the low resolution, high volume techniques of ultrasound, CT, or MRI scans.

The optical path of our system

A rendering of a tissue slice

To achieve such capabilities our microscope operates in a very particular fashion.  In the KESM system, a modified diamond microtome knife is held fixed with an illumination source shining through the blade.  The sample is then moved into the knife, cutting off 1 micron thick sections.  As each slice is made, it is illuminated by the light exiting the knife and simultaneously scanned by our scanning objective.  The process is repeated across the face of the cube, and after many thousands of slices are made the tissue being examined is completely scanned into the system.

After this the software side of our system steps in, reconstructing from the thousands upon thousands of images a complete 3d model of the tissue investigated.  With the 3d model in hand, many different compression techniques can be used to reduced the large volume of raw data (approx. .4 terabytes per cubic centimeter) to an easily manageable size.  Further, depending on the staining and tissue being imaged, a variety of techniques can be used to artificially color the tissue to provide even more data layered over the 3d construction.  Further in the future we imagine using several different stains to provide yet another dimension of information, as we can easily image multiple fluorescences in each slice.

Finally is the analysis, in which our machine vision techniques can give detailed information about the tissue not previously possible.  By analyzing a variety of mathematical factors of the final model, and by comparing these factors to a library of similar tissues previously imaged, the KESM system can serve as an aid to the researcher in pointing out possible interesting phenomenon found through data mining that researcher may not have picked up on during first inspection.

The Knife-Edge Scanning Microscope was originally developed in the Brain Networks Laboratory at Texas A&M.

For information about tissue preparation consult our methods paper on the Journal of Visualized Experiments, Specimen Preparation, Imaging, and Analysis Protocols for Knife-edge Scanning Microscopy.

3Scan © 2014