Objective To recover broad-neutralizing monoclonal antibodies (BnAbs) from avian influenza A (H5N1)virus infection cases and investigate their genetic and functional features. Methods We screened the Abs repertoires of expanded B cells circulating in the peripheral blood of H5N1 patients. The genetic basis, biological functions, and epitopes of the obtained BnAbs were assessed and modeled. Results Two BnAbs, 2-12D5, and 3-37G7.1, were respectively obtained from two human H5N1 cases on days 12 and 21 after disease onset. Both Abs demonstrated cross-neutralizing and Ab-dependent cellular cytotoxicity (ADCC) activity. Albeit derived from distinct Ab lineages, i.e., VH1-69-D2-15-JH4 (2-12D5) and VH1-2-D3-9-JH5 (3-32G7.1), the BnAbs were directed toward CR6261-like epitopes in the HA stem, and HA2 I45 in the hydrophobic pocket was the critical residue for their binding. Signature motifs for binding with the HA stem, namely, IFY in VH1-69-encoded Abs and LXYFXW in D3-9-encoded Abs, were also observed in 2-12D5 and 3-32G7.1, respectively. Conclusions Cross-reactive B cells of different germline origins could be activated and re-circulated by avian influenza virus. The HA stem epitopes targeted by the BnAbs, and the two Ab-encoding genes usage implied the VH1-69 and D3-9 are the ideal candidates triggered by influenza virus for vaccine development.