ht-MASH: a high-throughput, cost-effective, and robust protocol for microscopic 3D imaging of human angio- and cytoarchitecture in large human brain samples.

The performance of many optical tissue clearing protocols has considerably improved in the last few years, so that now even notoriously difficult specimen such as highly myelinated human brain tissue can be rendered highly transparent. However, optical tissue clearing is still routinely performed on relatively small samples, especially in the case of the human brain. Recent advances in histological tissue processing now allow scaling up the clearing process considerably towards much larger samples. Yet so far, these methods can have considerable drawbacks in their feasibility to be implemented routinely, especially in smaller laboratories. Here, we present an updated version of our MASH protocol, which allows optical tissue clearing of very large human brain tissue samples and labelling of angio- and cytoarchitecture therein. This pipeline is cost-efficient and easy to implement, so that even smaller labs can apply it at scale. At the same time, the use of rapid prototyping using 3D printing to create custom clearing equipment is versatile enough to be adjusted to other optical tissue clearing methods than the one used in this study (e.g., aqueous methods such as CUBIC or other solvent-based methods of the DISCO family), sample sizes or tissue types. Our pipeline has, therefore, the potential to advance optical tissue clearing and labelling of large human tissue samples towards a more robust and routine implementation in the blooming field of 3D histology.