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Characterization of Human Monocyte Subsets by Whole Blood Flow Cytometry Analysis
Published on: October 17, 2018
Evaluation of a 12-color flow cytometry panel to study lymphocyte, monocyte, and dendritic cell subsets in humans
Patrick Autissier1, Caroline Soulas, Tricia H Burdo
1Department of Biology, Boston College, Chestnut Hill, Massachusetts 02467, USA.
Insights
A novel 12-color flow cytometry panel enables simultaneous identification of diverse human immune cells, including lymphocytes, monocytes, and dendritic cell (DC) subsets. This advancement aids in studying immune responses during infections like HIV.
Area of Science:
- Immunology
- Cell Biology
- Cytometry
Background:
- Monitoring immune cell populations like lymphocytes, monocytes, and dendritic cells (DCs) is vital for understanding infectious diseases such as HIV.
- Identifying rare or heterogeneous cell populations, particularly DC subsets, presents challenges in traditional flow cytometry.
Purpose of the Study:
- To develop and validate a single 12-color flow cytometry panel for comprehensive analysis of human lymphocyte, monocyte, and DC subsets.
- To enable simultaneous detection of major circulating immune cell populations in a single analysis.
Main Methods:
- Blood samples from healthy volunteers underwent erythrocyte lysis and were labeled with a 12-antibody cocktail.
- Samples were analyzed using a Becton-Dickinson FACSAria with three lasers.
- Data were compared against lineage-specific panels and analyzed using FlowJo software.
Main Results:
- The 12-color panel successfully identified major subsets: CD4+ and CD8+ T lymphocytes, B cells, NK cells, NKT cells, and monocyte subsets (CD14+/CD16+).
- Five distinct HLA-DR+Lin- subsets were identified: CD34+ stem cells, CD123+ plasmacytoid DCs, and three CD11c+ myeloid DC subsets (CD16+, CD1c+, CD141+).
- This panel allows for simultaneous detection of lymphocytes, monocytes, and all known DC subsets.
Conclusions:
- A single 12-color flow cytometry panel provides a comprehensive approach to analyzing key immune cell populations.
- This method facilitates a broader understanding of immune responses in conditions like HIV infection.
- It aids in better defining the roles of individual cell types in diseases such as Acquired Immune Deficiency Syndrome (AIDS).
Abstract:
Monitoring changes in human immune cell populations such as lymphocytes, monocytes, and dendritic cells (DCs) during infectious diseases like human immunodeficiency virus (HIV) is crucial. However, difficulties to identify rare or heterogeneous cell populations can be limiting. For example, to accurately measure DC subsets, eight flow cytometry parameters are ideal. The aim of this work was to analyze the phenotype of human lymphocyte, monocyte, and DC subsets using a single 12-color flow cytometry panel. After erythrocyte lysis, blood from healthy human volunteers was washed and labeled with a cocktail of 12 antibodies. Samples were analyzed on a Becton-Dickinson FACSAria equipped with three lasers. Data were compared with lineage-specific panels using 5-8 Ab combinations per lineage. Acquired data were analyzed using FlowJo software. Our 12-color panel allows for the identification of the following major subsets of circulating cells in a single tube: CD4+ and CD8+ T lymphocytes, B lymphocytes, NK cells, NKT cells, monocyte subsets (CD14 and/or CD16), and five nonoverlapping HLA-DR+Lin- subsets: CD34+ hematopoietic stem cells, CD123+ plasmacytoid DC, and three subsets of CD11c+ myeloid DC expressing either CD16, CD1c (BDCA-1), or CD141 (BDCA-3). We have developed a single flow cytometry panel that allows for simultaneous detection of the lymphocyte and monocyte cell populations and all known DC subsets. Studying these major players of the immune system in one single panel may give us a broader view of the immune response during HIV infection and the ability to better define the role of individual cell types in Acquired Immune Deficiency Syndrome (AIDS) pathogenesis. (c) 2010 International Society for Advancement of Cytometry.
