Abstract
Electron microscopy (EM) is widely used for studying cellular structure and network connectivity in the brain. We have built a parallel imaging pipeline using transmission electron microscopes that scales this technology, implements 24/7 continuous autonomous imaging, and enables the acquisition of petascale datasets. The suitability of this architecture for large-scale imaging was demonstrated by acquiring a volume of more than 1 mm3 of mouse neocortex, spanning four different visual areas at synaptic resolution, in less than 6 months. Over 26,500 ultrathin tissue sections from the same block were imaged, yielding a dataset of more than 2 petabytes. The combined burst acquisition rate of the pipeline is 3 Gpixel per sec and the net rate is 600 Mpixel per sec with six microscopes running in parallel. This work demonstrates the feasibility of acquiring EM datasets at the scale of cortical microcircuits in multiple brain regions and species.
Highlights
Serial-section electron microscopy has a long history of elucidating brain structure and connectivity1–3
The imaging platform described here uses a standard JEOL 1200EXII 120 kV transmission electron microscopy (TEM) that has been modified with customized hardware and software
The key hardware modifications are: [1] an extended column and custom electron-sensitive scintillator that produce a tenfold increase in the field-of-view with negligible impact on spatial resolution; [2] a single, large-format CMOS camera outfitted with a low distortion lens that reduces image acquisition time to 50–150 ms; [3] a nano-positioning sample stage that offers fast, high-fidelity montaging of large tissue sections; and [4] an advanced reel-to-reel tape translation system that accurately locates each section using index barcodes for random access on the GridTape32
Summary
We have built a parallel imaging pipeline using transmission electron microscopes that scales this technology, implements 24/7 continuous autonomous imaging, and enables the acquisition of petascale datasets. The suitability of this architecture for largescale imaging was demonstrated by acquiring a volume of more than 1 mm of mouse neocortex, spanning four different visual areas at synaptic resolution, in less than 6 months. Such EM systems should provide high degrees of automation and reliability in order to permit unsupervised continuous operation
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