M2e5x VLP-supplemented vaccination induces M2e-specific antibodies and enhanced levels of antibodies to heterosubtypic viral antigens. BALB/c mice (N = 18) were immunized with split vaccine alone (Split) or split vaccine supplemented with M2e5x VLPs (Split+M2e5x). Sera were collected 3 weeks after prime (P) and boost (B) vaccination. The IgG level was detected using M2e peptide or influenza A viruses, A/California/04/09 (A/Cali, H1N1), A/Philippines/2/82 (A/Phil, H3N2), and reassortant A/Vietnam/1203/2004 (rgH5N1) as an ELISA coating antigen for antibody detection. (a) IgG antibody responses to M2e peptide. (b) IgG antibody responses to influenza A/Cali virus. (c) Comparison of IgG antibody responses to influenza A/Phil (H3N2) virus. (d) Comparison of IgG antibody responses to reassortant A/Vietnam/1203/2004 virus (rgH5N1). Asterisk indicates significant differences between sera of split vaccine or split vaccine supplemented with M2e5x VLP (*P < 0.05, **P < 0.01). Error bars indicate mean ± SEM. Split: influenza split vaccine (2009 H1N1 pandemic strain), Split+M2e5x: split vaccine supplemented with M2e5x VLPs. ELISA, enzyme-linked immunosorbent assay; M2e, M2 ectodomain; VLP, virus-like particles.