Laser particle barcoding for multi-pass high-dimensional flow cytometry

Kwok, S. J.; Forward, S.; Fahlberg, M. D.; Cosgriff, S.; Lee, S. H.; Abbott, G.; Zhu, H.; Minasian, N. H.; Vote, A. S.; Martino, N.; Yun, S. H.

doi:10.1101/2022.06.03.494697

bioRxiv · 10.1101/2022.06.03.494697

Laser particle barcoding for multi-pass high-dimensional flow cytometry

Kwok, S. J.·Forward, S.·Fahlberg, M. D.·Cosgriff, S.·Lee, S. H.·Abbott, G.·Zhu, H.·Minasian, N. H.·Vote, A. S.·Martino, N.·Yun, S. H.

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Abstract

Flow cytometry is a standard technology in life science and clinical laboratories used to characterize the phenotypes and functional status of cells, especially immune cells. Recent advances in immunology and immuno-oncology as well as drug and vaccine discovery have increased the demand to measure more parameters. However, the overlap of fluorophore emission spectra and one-time measurement nature of flow cytometry are major barriers to meeting the need. Here, we present multi-pass flow cytometry, in which cells are tracked and measured repeatedly through barcoding with infrared laser-emitting microparticles. We demonstrate the benefits of this approach on several pertinent assays with human peripheral blood mononuclear cells (PBMCs). First, we demonstrate unprecedented time-resolved flow characterization of T cells before and after stimulation. Second, we show 33-marker deep immunophenotyping of PBMCs, analyzing the same cells in 3 back-to-back cycles. This workflow allowed us to use only 10-13 fluorophores in each cycle, significantly reducing spectral spillover and simplifying panel design. Our results open a new avenue in multi-dimensional single-cell analysis based on optical barcoding of individual cells.

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Kwok, S. J., Forward, S., Fahlberg, M. D., Cosgriff, S., Lee, S. H., Abbott, G., Zhu, H., Minasian, N. H., Vote, A. S., Martino, N., Yun, S. H.. 2022-06-04. Laser particle barcoding for multi-pass high-dimensional flow cytometry. https://doi.org/10.1101/2022.06.03.494697

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