A complete pipeline for high‐plex spatial proteomic profiling and analysis of neural cell phenotypes on the CosMx™ Spatial Molecular Imager and AtoMx™ Spatial Informatics Platform
Authors: Rosenbloom Alyssa , Bonnett Shilah , Conner Mark , Phan‐Everson Tien , Lewis Zachary , Ong Giang , Liang Yan , Brown Emily , Pan Liuliu , Wardhani Aster , Korukonda Mithra , Brown Carl , Dunaway Dwayne , Zhao Edward , McGuire Dan , Woo Sangsoon , Filanoski Brian , Meredith Rhonda , Chantranuvatana Kan , Birditt Brian , Yi Hye Son , Piazza Erin , Reeves Jason , Kang Christine , Geiss Gary , Beechem Joseph M. 
Published: December 2023

Background

The brain is complex and heterogeneous where cell function and cell‐to‐cell communication are critical for rapid and accurate performance. The ability to explore protein‐driven activities at high resolution within spatial context of their immediate environment is critical to gain comprehensive pictures of brain development, activity, aging, disease or dysfunction, and inflammatory responses. Many existing approaches for high‐plex single cell spatial proteomics face issues around simplicity, speed, scalability, and big data analysis.

Method

Here, we present an integrated workflow from sample through analysis that addresses key concerns around high plex proteomics. The CosMx™ Spatial Molecular Imager and AtoMx™ Spatial Informatics Platform comprises an end‐to‐end workflow that efficiently handles highly multiplex protein analysis at plex sizes exceeding 68 targets. The CosMx protein assays uses oligonucleotide conjugated antibodies, detected using universal, multi‐analyte CosMx readout reagents. The CosMx Mouse Neural Cell Typing and Alzheimer’s Pathology panel is optimized to comprehensively profile neural cell lineages across the brain as well as the progression of Alzheimer’s disease (AD), including specific antibodies for humanized mouse AD models. The AtoMx spatial informatics platform provides full analysis support, including whole‐slide image viewer, and methods for performing built‐in or fully customizable analyses for cell typing, ligand‐receptor analysis, neighborhood analysis and spatial differential expression.

Result

The CosMx protein assay reagents were validated on FFPE adult mouse brain, mouse embryo, and Alzheimer’s positive human brain. We used the CosMx Mouse Neural Cell Typing and Alzheimer’s Pathology panel with the CosMx Spatial Molecular Imager to identify multiple neuronal subtypes, different reactive states of astrocytes and microglial, cell degeneration and proliferation. Single cell exploration of mitochondria showed distinct patterning of key immune targets based on their immediate microenvironment.

Conclusion

CosMx SMI is a high‐plex spatial multi‐omics platform that enables detection of > 68 proteins at subcellular resolution. In combination with the high‐plex CosMx Mouse Neural Cell Typing and Alzheimer’s Pathology panel, we present a flexible and scalable informatics platform, a robust solution for comprehensive neural and disease phenotyping that captures the complexity of neuronal and glial cellular activity with full spatial context.

FOR RESEARCH USE ONLY. Not for use in diagnostic procedures.