Enduring protection from pathogens depends on generation of long-lived memory and antibody-secreting plasma cells in germinal centers (GC). The GC is segregated into a dark zone, where B cells proliferate and mutate their antibodies, and a light zone (LZ) where T cells select B cells for clonal expansion. However, the selection mechanism for GC-egress and the precise departure site remain unclear. Using light-sheet fluorescence microscopy for large-scale imaging of intact lymph nodes, we demonstrate that individual GCs generate highly heterogeneous numbers of departing cells. Combining whole-organ imaging, photo-labeling and single-cell-RNA sequencing revealed that B cells depart GCs through the LZ in a transitional differentiation state. Furthermore, we show that selection for GC-egress is dictated by intrinsic-B cell receptor affinity. Our findings provide a unifying model for the role of T cells in expansion of clones available for GC-egress, and the requirement for intrinsic B cell functions for antibody-mediated immunity.
