From HeLa to human blood: extending single-cell proteomics to the smallest immune cells

Hendriks, I. A.; Buch-larsen, S. C.; Rykaer, M.; Lechner, M. Y.; Kverneland, A. H.; Arrey, T. N.; Hermanson, D.; Damoc, E.; Olsen, J. V.

doi:10.1101/2025.11.10.687556

bioRxiv · 10.1101/2025.11.10.687556

From HeLa to human blood: extending single-cell proteomics to the smallest immune cells

Hendriks, I. A.·Buch-larsen, S. C.·Rykaer, M.·Lechner, M. Y.·Kverneland, A. H.·Arrey, T. N.·Hermanson, D.·Damoc, E.·Olsen, J. V.

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Abstract

Single-cell proteomics (SCP) holds the promise of decoding cellular heterogeneity at the functional level, yet achieving deep and reproducible proteome coverage from individual cells has remained a formidable challenge. Here, we established a high-sensitivity, label-free SCP platform that surpasses previous limits in depth and reproducibility. By integrating optimized low-input sample processing, refined liquid chromatography, and the Orbitrap Astral Zoom mass spectrometer, our approach routinely quantified over 7,000 proteins per individual HeLa cell, capturing thousands of low-abundance proteins that eluded prior SCP studies. Applied to very small human peripheral blood mononuclear cells (PBMCs), we identified up to 4,000 proteins per cell, including key markers distinguishing monocytes, T cells, and activated lymphocytes within heterogeneous populations, underscoring that single-cell proteomics can now directly elucidate clinically relevant primary samples with both depth and precision.

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Hendriks, I. A., Buch-larsen, S. C., Rykaer, M., Lechner, M. Y., Kverneland, A. H., Arrey, T. N., Hermanson, D., Damoc, E., Olsen, J. V.. 2025-11-11. From HeLa to human blood: extending single-cell proteomics to the smallest immune cells. https://doi.org/10.1101/2025.11.10.687556

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