While two memory compartments, memory B cells and long-lived plasma cells, are thought to contribute to the successful establishment of memory recall responses, the unique roles of each cellular compartment are still unclear. Herein, by tracing influenza anti-hemagglutinin (HA)-specific antibodies in mice, we demonstrate that pre-existing antibodies secreted by long-lived plasma cells are essential for protection from re-infection with the same influenza virus, whereas protection from secondary infection with an antigenically distinct influenza virus requires memory B cell activation. These activated memory B cells were largely specific for the conserved HA stem region, and generated sufficient levels of antibodies for protection from heterologous reinfection. Given that the anti-stem plasmablasts derived from the memory B cells were higher affinity than those from na?ve B cells, our results suggest that maturation of anti-stem memory B cells during primary influenza infection and their subsequent activation are required for protection from re-infection by mutant viruses.