Multiplexed and scalable cellular phenotyping toward the standardized three-dimensional human neuroanatomy

Abstract

The advent of three-dimensional histological methods has advanced studies of cellular-resolution anatomy of the brain. The use of whole-mount staining and tissue clearing has advanced systems-level identification of cells underlying brain functions in mouse models. However, application of these methods to studies of human brains has been difficult due to their structural variability and the lack of standardized quantitative metrics. Here we report a rapid and scalable staining/imaging technique, termed mFISH3D, that enables single-cell-resolution imaging of mRNAs of more than ten genes in a large mammalian brain. To apply mFISH3D to postmortem human cerebral cortex, we have reconstructed morphogenic tracks of cortical growth, and used the tracks to provide a framework for quantitative assessment of cytoarchitecture. The workflow enabled the objective quantification of biological heterogeneity among cortical regions. We propose these techniques for standardization of 3D histology of the human cortex to provide reproducible measurements of cell-type-specific neuroanatomy.

Tissue clearing method in visualization of cancer progression and metastasis

“…Although some clearing methods are not compatible for retaining mRNA, optimized tissue-clearing methods can capture mRNA signals with 3D images using mFISH (single-molecule fluorescent in situ hybridization), which is a basic method for some spatial transcriptomic technologies ( 20 , 55 , 69 , 70 ). Heinz and Murakami recently reported that their workflow “mFISH3D” enables mRNAs to be visualized using clearing with high resolution in mouse and human brains ( 71 ). In terms of proteomics, Ertürk’s group recently succeeded in spatial mass spectrometry from small portion of cleared samples ( 72 ).…”