Evaluation of a 12color flow cytometry panel to study lymphocyte, monocyte, and dendritic cell subsets in humans

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 12color 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 BectonDickinson FACSAria™ equipped with three lasers. Data were compared with lineagespecific panels using 5–8 Ab combinations per lineage. Acquired data were analyzed using FlowJo software. Our 12color 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 andor CD16, and five nonoverlapping HLADRLin− subsets: CD34 hematopoietic stem cells, CD123 plasmacytoid DC, and three subsets of CD11c myeloid DC expressing either CD16, CD1c BDCA1, or CD141 BDCA3. 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. © 2010 International Society for Advancement of Cytometry