Significance The development of a universal influenza vaccine is a major public health need globally, and identifying the optimal formulation will be an important first step for developing such a vaccine. Here we show that a two-dose immunization of humans with an inactivated, AS03-adjuvanted H5N1 avian influenza virus vaccine engaged both the preexisting memory and naive B cell compartments. Importantly, we show that the recruited memory B cells after first immunization were directed against conserved epitopes within the H5 HA stem region while the responses after the second immunization were mostly directed against strain-specific epitopes within the HA globular head. Taken together these findings have broad implications toward optimizing vaccination strategies for developing more effective vaccines against pandemic viruses.