Immersion Fixation and Staining of Multicubic Millimeter Volumes for Electron Microscopy-Based Connectomics of Human Brain Biopsies

Overview

Journal Biol Psychiatry

Publisher Elsevier

Specialty Psychiatry

Date 2023 Feb 5

PMID 36740206

Authors

Neha Karlupia

Richard L Schalek

Yuelong Wu

Yaron Meirovitch

Donglai Wei

Alexander W Charney

Brian H Kopell

Jeff W Lichtman

Affiliations

Soon will be listed here.

Abstract

Connectomics allows mapping of cells and their circuits at the nanometer scale in volumes of approximately 1 mm. Given that the human cerebral cortex can be 3 mm in thickness, larger volumes are required. Larger-volume circuit reconstructions of human brain are limited by 1) the availability of fresh biopsies; 2) the need for excellent preservation of ultrastructure, including extracellular space; and 3) the requirement of uniform staining throughout the sample, among other technical challenges. 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 × 2 × 2 mm and up to 37 mm with one dimension ≤2 mm. This method should allow synapse-level circuit analysis in samples from patients with psychiatric and neurologic disorders.

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