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Deep phenotyping ILCs and T cells: A comparative analysis protocol for conventional and spectral flow cytometry
Jadie Acklam1, Sukhveer K Mann2, Karen G Hogg2
1Centre for Blood Research, York Biomedical Research Institute, Department of Biology, University of York, Heslington, York YO10 5DD, UK; NIHR, Leeds-York Biomedical Research Centre, Leeds, UK.
Here, we present a protocol for identifying human innate lymphoid cells (ILCs) and T cells using a 19-plex flow cytometry panel. We describe steps for titrating antibodies, preparing single-stain controls and samples, and staining cells for human peripheral blood and bone marrow samples. We then detail procedures for transferring a flow cytometry panel optimized for conventional cytometry to spectral flow cytometry analysis. This protocol facilitates correlation with murine models, maximizes measurable parameters, resolves complex fluorophore combinations, and increases panel flexibility.
Here, we present a protocol for identifying human innate lymphoid cells (ILCs) and T cells using a 19-plex flow cytometry panel. We describe steps for titrating antibodies, preparing single-stain controls and samples, and staining cells for human peripheral blood and bone marrow samples. We then detail procedures for transferring a flow cytometry panel optimized for conventional cytometry to spectral flow cytometry analysis. This protocol facilitates correlation with murine models, maximizes measurable parameters, resolves complex fluorophore combinations, and increases panel flexibility.
