Influenza vaccine effectiveness varies annually due to the fast evolving seasonal influenza A(H3N2) strain and egg-derived mutations-both of which can cause a mismatch between the vaccine and circulating strains. To address these limitations, we have developed a hemagglutinin (HA)-based protein-detergent nanoparticle influenza vaccine (NIV) with a saponin-based Matrix-M? adjuvant. In a phase 1 clinical trial of older adults, the vaccine demonstrated broadly cross-reactive A(H3N2) HA antibody responses. Two broadly neutralizing monoclonal antibodies derived from NIV-immunized mice were characterized by transmission electron microscopy (TEM), antibody competition assays, fluorescence-activated cell sorting (FACS) analysis, and protein-protein docking. These antibodies recognize two conserved regions of the head domain, namely the receptor binding site and the vestigial esterase subdomain, thus demonstrating the potential for an HA subunit vaccine to elicit antibodies targeting structurally and antigenically distinct but conserved sites. Antibody competition studies with sera from the phase 1 trial in older adults confirmed that humans also make antibodies to these two head domains and against the highly conserved stem domain. This data supports the potential of an adjuvanted recombinant HA nanoparticle vaccine to induce broadly protective immunity and improved vaccine efficacy.