OMIP‐114: A 36‐Color Spectral Flow Cytometry Panel for Detailed Analysis of T Cell Activation and Regulation in Small Human Blood Volumes

• Published in: Cytometry. Part A Vol. 107; no. 6; pp. 364 - 371
• Main Authors: Thieme‐Ehlert, Marie‐Theres, Jacobs, Thomas, Brandi, Johannes, Mackroth, Maria Sophia
• Format: Journal Article
• Language: English
• Published: Hoboken, USA John Wiley & Sons, Inc 01.06.2025; Wiley Subscription Services, Inc
• Subjects: B-Lymphocytes - immunology; Biomarkers; Blood; Cell activation; Color; Flow cytometry; Flow Cytometry - methods; Helper cells; Humans; Immune checkpoint; Immunological memory; Immunophenotyping - methods; Infectious diseases; Inflammatory diseases; Killer Cells, Natural - immunology; Lymphocyte Activation - immunology; Lymphocytes; Lymphocytes B; Lymphocytes T; Malaria; Malaria - blood; Malaria - immunology; Memory cells; Peripheral blood mononuclear cells; Subpopulations; T-Lymphocyte Subsets - immunology; T-Lymphocytes - immunology; Vector-borne diseases
• ISSN: 1552-4922; 1552-4930; 1552-4930
• DOI: 10.1002/cyto.a.24937

ABSTRACT This 36‐color flow cytometry panel is designed to characterize multiple lymphocyte compartments, with a focus on T cells, their memory subpopulations, and immune checkpoints in human whole blood samples. In clinical settings, the amount of blood available from patients for scientific research is often limited. This restriction may be further exacerbated when working with samples from small children or in resource‐poor settings—both scenarios commonly encountered in malaria and infectious disease research. Accordingly, this panel is designed to maximize the information that can be obtained from as little as 200 μL whole blood using flow cytometry. This panel allows a phenotypic characterization of the main subpopulations within T cells, as well as B cells and NK cells. It includes markers for the analysis of memory subpopulations, regulatory T cell subsets, and T follicular helper cells. Furthermore, surface and intracellular markers for activation and differentiation, effector functions, exhaustion, and immune checkpoints are included, allowing detailed characterization of the main lymphocyte subsets, in particular T cells. This panel was optimized for the analysis of fresh human blood samples obtained from malaria patients, but it may be adapted to the analysis of isolated PBMC or tissue samples, as well as samples from patients with other infectious or inflammatory diseases.