Techniques and Methods|Articles in Press

Immersion fixation and staining of multi-cubic millimeter volumes for electron microscopy-based connectomics of human brain biopsies

Published:February 03, 2023DOI:


      Connectomics allow mapping of cells and their circuits at the nanometer scale in volumes of about 1 mm3. Given that the human cerebral cortex can be 3 mm in thickness, larger volumes are required. Larger volume circuit reconstructions of human brain, among other technical challenges, are limited by a) availability of fresh biopsies; b) need for excellent preservation of ultrastructure including extracellular space; c) requirement of uniform staining throughout the sample. Cerebral cortical samples from neurosurgical patients are available owing to lead placement for deep brain stimulation. Described here is an immersion fixation, heavy metal staining and tissue processing method that consistently provides excellent ultrastructure throughout human and rodent surgical brain samples of volumes 2 x 2 x 2 mm3 and of volumes up to 37 mm3 with one dimension no greater than 2 mm. This method should allow synapse level circuit analysis in samples from psychiatric and neurological patients.

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