FlyEM plans to trace neurons throughout the fly brain, spanning regions larger than 100 microns. Because of the limited field of view possible when doing FIB-SEM imaging, we introduced a hot-knife sectioning strategy that involves cutting a specimen into slabs several microns thick. Cutting with a hot-knife ensures minimal damage to the tissue. Each of these small slabs can then be imaged using FIB-SEM.
The advantages of hot-knife sectioning are two-fold. First, it enables imaging over very large spans of neuropil which would otherwise be infeasible. Second, it is possible to speed up dataset acquisition by parallelizing (scaling out) the imaging across several FIB-SEM machines.
Figure: Two adjacent sections resulting from hot-knife sectioning.
Video: Registration between hot-knife sections is almost seamless.
The figure above shows the quality of the hot-knife technique by showing two adjacent sectioned slabs. The quality of the cut enables the two slabs to be registered with only a small amount of data loss permitting the tracing of almost every process. The video reveals the quality of this registration.