Abstract Classifying the heterogeneous human B cell compartment into discrete subsets based on surface phenotype alone remains a major challenge in the field of B cell biology. Our lab has discovered a novel mouse IgM+IgDlow/- B cell subset (BDL) that induces Treg proliferation, making it a potentially significant therapeutic target for autoimmune diseases. But due to their rarity and lack of sufficient cell surface identifiers, a more definitive phenotype is required to track this B cell subset in humans. The objective of these studies was to characterize human IgDlow/- B cells from PB using multiomic CITE-seq analysis. Here, we adapted a standardized gating scheme (CD27xIgD → CD24xCD38) that captures all the major canonical human PB B cell subsets. This scheme allowed us to analyze FACS-purified CD19+IgM+IgDlow/- PB B cells using CITE-seq by targeting these four protein markers and recreate biaxial plots similar to our flow cytometry analyses. It is increasingly understood that mRNA levels within a cell do not correlate well with cell surface protein expression. This represents a major drawback to using a transcriptome-only scRNA-seq approach because any ‘novel’ cell populations discovered based on the upregulation of surface protein transcripts may prove impossible to validate. These studies have established a workflow for a multiomic approach to identify discrete human B cell subsets based on cell surface expression that can be analyzed for transcriptional differences.
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