An Analysis of Phenotyping and Gating Strategies for B Cells in Human Peripheral Blood

The purpose of this experiment was to design a 9-marker, 8-colour antibody panel and gating strategy for a conventional 3-laser (violet-blue-red) flow cytometer to phenotype B cells from peripheral blood mononuclear cells (PBMCs) in humans. The panel was designed using a combination of B cell lineage markers and markers to identify specific B cell subsets.
Conclusion:
In this experiment, we present a phenotyping panel and optimized gating strategy to identify the following B cell subsets from human PBMCs: Transitional B cells, Naïve B cells, Antibody Secreting Cells (ASC), Unswitched Memory B cells, Switched Memory B cells, IgD Only Memory B cells, IgM Only Memory B cells, Double Negative 1 B cells (DN1) and Double Negative 2 B cells (DN2). Our work demonstrates the importance of choosing a gating strategy to accurately report on the prominent B cell subsets present in human peripheral blood. Daily instrument QC, use of application settings (on BD FACSDiva) and FMO controls for appropriate markers.