This study describes a CD5+ B cell that differs from the majority of the CD5+ B cells from human tonsils. This cell, isolated from in vivo activated B cells, expressed activation markers and featured a CD23-, IgMhigh, IgDlow surface phenotype, responded to T cell-independent type-2 antigens in vitro, and was detected in the subepithelial (SE) areas, the tonsil equivalent of the splenic marginal zone (MZ). Most of the cells utilized unmutated Ig VH genes, although cells with mutated genes also were found, a finding confirmed by single-cell studies. Mutated sequences were more frequent in suspensions enriched for CD27+ cells. Repeated VDJ gene sequences were observed in different molecular clones from the same cell suspension, suggesting in situ expansion. These CD5+ B cells seem to share features with previously characterized tonsil CD5- SE B cells and differ from the majority of tonsil CD5+ B cells, which have the surface phenotype of follicular mantle B cells, lack activation markers, do not respond to T cell-independent antigens, and utilize unmutated VH genes. These data are discussed considering the present views on the origin of B cell subset populations and the relationships between MZ and B1 cells.