A high-resolution blood immune cell doublet atlas via imaging spectral cytometry
- 1La Jolla Institute for Immunology
Abstract
Circulating cell-cell complexes (doublets) provide critical insights into in vivo immune interactions. Here, we leverage high-dimensional imaging spectral cytometry to systematically map the landscape of immune doublets in human peripheral blood. By integrating morphometric imaging features with a 22-color spectral panel, we established a gating workflow that cleanly segregates genuine, physically interacting complexes from coincidental transits and platelet contaminants, and resolved 12 highly pure homotypic and heterotypic immune doublet configurations in peripheral blood mononuclear cells (PBMC). At steady state, each doublet population displayed unique abundance, affinity and surface phenotypic profiles relative to circulating singlets. Principal component analysis (PCA) of quantitative imaging features further revealed lineage-specific topologies distinguishing singlet cells, homotypic doublets, and heterotypic pairs. Finally, comparative cohort analysis demonstrated that while doublet frequencies, affinities and surface phenotypes remain comparable between healthy donors and patients with tuberculosis (TB) disease, imaging-derived features uncovered disease-specific alterations in doublets morphology. Specifically, high-dimensional clustering using imaging parameters showed that TB disease significantly enriches for a tightly synapsed T cell-monocyte cluster characterized by high CD3-CD14 pixel correlation. Collectively, this study establishes a robust framework for mapping circulating cell-cell networks and demonstrates the superior sensitivity of imaging features over conventional fluorescence for capturing functional immune interactions in health and disease.
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BibTeX
@misc{kim2026highresolution,
title = {{A high-resolution blood immune cell doublet atlas via imaging spectral cytometry}},
author = {C. Kim and D. Hinz and A. Middlebrook and A. J. Tyznik and L. M. D'Cruz and T. Scriba and B. Peters and J. G. Burel},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.03.749150},
url = {https://www.biorxiv.org/content/10.64898/2026.09.03.749150v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - A high-resolution blood immune cell doublet atlas via imaging spectral cytometry
AU - C. Kim
AU - D. Hinz
AU - A. Middlebrook
AU - A. J. Tyznik
AU - L. M. D'Cruz
AU - T. Scriba
AU - B. Peters
AU - J. G. Burel
PY - 2026
DA - 2026/09/03
PB - bioRxiv
DO - 10.64898/2026.09.03.749150
UR - https://www.biorxiv.org/content/10.64898/2026.09.03.749150v1
AB - Circulating cell-cell complexes (doublets) provide critical insights into in vivo immune interactions. Here, we leverage high-dimensional imaging spectral cytometry to systematically map the landscape of immune doublets in human peripheral blood. By integrating morphometric imaging features with a 22-color spectral panel, we established a gating workflow that cleanly segregates genuine, physically interacting complexes from coincidental transits and platelet contaminants, and resolved 12 highly pure homotypic and heterotypic immune doublet configurations in peripheral blood mononuclear cells (PBMC). At steady state, each doublet population displayed unique abundance, affinity and surface phenotypic profiles relative to circulating singlets. Principal component analysis (PCA) of quantitative imaging features further revealed lineage-specific topologies distinguishing singlet cells, homotypic doublets, and heterotypic pairs. Finally, comparative cohort analysis demonstrated that while doublet frequencies, affinities and surface phenotypes remain comparable between healthy donors and patients with tuberculosis (TB) disease, imaging-derived features uncovered disease-specific alterations in doublets morphology. Specifically, high-dimensional clustering using imaging parameters showed that TB disease significantly enriches for a tightly synapsed T cell-monocyte cluster characterized by high CD3-CD14 pixel correlation. Collectively, this study establishes a robust framework for mapping circulating cell-cell networks and demonstrates the superior sensitivity of imaging features over conventional fluorescence for capturing functional immune interactions in health and disease.
ER -