OMIP‐94: Twenty‐four‐color (thirty‐marker) panel for deep immunophenotyping of immune cells in human peripheral blood

• Published in: Cytometry. Part A Vol. 103; no. 9; pp. 695 - 702
• Main Authors: Pierzchalski, Arkadiusz, Zenclussen, Ana C., Herberth, Gunda
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.09.2023
• Subjects: Blood; CD27 antigen; CD38 antigen; CD4 antigen; CD45RA antigen; CD57 antigen; CD69 antigen; CD8 antigen; Cell differentiation; Color; Cytotoxicity; Dendritic cells; Eosinophils; Immune system; Invariants; L-selectin; Lasers; Leukocytes (basophilic); Leukocytes (eosinophilic); Leukocytes (mononuclear); Leukocytes (neutrophilic); Lymphocytes; Lymphocytes B; Lymphocytes T; Lymphoid cells; Monocytes; Natural killer cells; Peripheral blood mononuclear cells; Phenotypes; Surface markers; OMIP; whole blood; immune cell subsets; high-dimensional flow cytometry; broad immunophenotyping; spectral cytometry
• ISSN: 1552-4922; 1552-4930; 1552-4930
• DOI: 10.1002/cyto.a.24766

This newly established 24‐color (30‐marker) panel focuses on the characterization of the main human immune cell subtypes and was optimized for the analysis of human whole blood using a full spectrum flow cytometer. The panel covers all main leukocyte populations: neutrophils, eosinophils and basophils, monocytes (with additional subsets), dendritic cells, innate lymphoid cells and lymphocytes. As for lymphocytes, this panel includes CD4+ T helper, Treg cells, and CD8+ cytotoxic T cells. Further T cells subsets are included with special focus on invariant T cells: γδ T cells (including δ2TCR variant), invariant NKT cells and MAIT (mucosal‐associated invariant T cells) cells. Additionally, total B cells (including Bregs and plasmocytes), NK cells, and NKT cells are included. For the overall check of activation status of the analyzed immune cells we used HLA‐DR, CD38, CD57, CD69, PD‐1, and CD94. In addition, we used CD62L, CD45RA, CD27, and CD39 to describe the differentiation status of these cells. The panel was designed to maximize the information that can be obtained from surface markers in order to avoid the need for fixation and permeabilization steps. The presented multimarker panel offers the possibility to discover new immune cell subtypes which in patients and in cohort studies may lead to the identification of altered immune phenotypes and might give a link to immune system based or to certain other diseases. This panel was developed for a full spectrum flow cytometer equipped with a minimum of three lasers. We developed this panel using healthy human fresh blood, however it was also successfully used for staining of isolated human peripheral blood mononuclear cells (PBMC).