Spatial mapping of protein composition and tissue organization: a primer for multiplexed antibody-based imaging

TL;DR

This paper reviews recent multiplexed antibody-based imaging methods for mapping protein composition and tissue organization, emphasizing their importance for understanding cellular communication in complex tissues.

Contribution

It provides a comprehensive overview of multiplexed antibody imaging techniques, offering guidelines for validation, data generation, and harmonization to advance spatial proteomics.

Findings

Multiple antibody-based imaging strategies exist with distinct detection modes.

Frameworks for validation and data harmonization are essential for reproducibility.

Guidelines support the adoption and standardization of multiplexed tissue imaging.

Abstract

Tissues and organs are composed of distinct cell types that must operate in concert to perform physiological functions. Efforts to create high-dimensional biomarker catalogs of these cells are largely based on transcriptomic single-cell approaches that lack the spatial context required to understand critical cellular communication and correlated structural organization. To probe in situ biology with sufficient coverage depth, several multiplexed protein imaging methods have recently been developed. Though these antibody-based technologies differ in strategy and mode of immunolabeling and detection tags, they commonly utilize antibodies directed against protein biomarkers to provide detailed spatial and functional maps of complex tissues. As these promising antibody-based multiplexing approaches become more widely adopted, new frameworks and considerations are critical for training future users, generating molecular tools, validating antibody panels, and harmonizing datasets. In this perspective, we provide essential resources and key considerations for obtaining robust and reproducible multiplexed antibody-based imaging data compiling specialized knowledge from domain experts and technology developers.