Multiplex, quantitative cellular analysis in large tissue volumes with clearing-enhanced 3D microscopy (Cₑ3D)
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Li, Weizhe, Ronald N. Germain, and Michael Y. Gerner. “Multiplex, Quantitative Cellular Analysis in Large Tissue Volumes with Clearing-Enhanced 3D Microscopy (Cₑ3D).” Proceedings of the National Academy of Sciences 114, no. 35 (2017): E7321–30. https://doi.org/10.1073/pnas.1708981114.
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This work was written as part of one of the author's official duties as an Employee of the United States Government and is therefore a work of the United States Government. In accordance with 17 U.S.C. 105, no copyright protection is available for such works under U.S. Law.
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Abstract
Organ homeostasis, cellular differentiation, signal relay, and in situ function all depend on the spatial organization of cells in complex tissues. For this reason, comprehensive, high-resolution mapping of cell positioning, phenotypic identity, and functional state in the context of macroscale tissue structure is critical to a deeper understanding of diverse biological processes. Here we report an easy to use method, clearing-enhanced 3D (Cₑ3D), which generates excellent tissue transparency for most organs, preserves cellular morphology and protein fluorescence, and is robustly compatible with antibody-based immunolabeling. This enhanced signal quality and capacity for extensive probe multiplexing permits quantitative analysis of distinct, highly intermixed cell populations in intact Cₑ3D-treated tissues via 3D histo-cytometry. We use this technology to demonstrate large-volume, high-resolution microscopy of diverse cell types in lymphoid and nonlymphoid organs, as well as to perform quantitative analysis of the composition and tissue distribution of multiple cell populations in lymphoid tissues. Combined with histo-cytometry, Cₑ3D provides a comprehensive strategy for volumetric quantitative imaging and analysis that bridges the gap between conventional section imaging and disassociation-based techniques.