Developing universal influenza vaccines (UIVs) capable of eliciting broad protection remains a critical public health priority. Neuraminidase (NA), a relatively conserved surface antigen of influenza viruses, is a promising target for UIV design. Here, we produced full-length trivalent NA proteins via a baculovirus-insect cell expression system, matching the WHO-recommended strains (H1N1, H3N2, and B/Victoria) for the 2024-2025 season. Immunogenicity evaluation of N2NA (derived from H3N2) in mice demonstrated that nanoemulsion (NE) adjuvant significantly enhanced NA-inhibiting (NI) antibody titers, CD4 T cell responses, and the numbers of IL-2-, IL-4-and IFN-γ-secreting lymphocytes compared to the unadjuvanted vaccine. Monovalent N1 NA and BV NA with NE adjuvant similarly induced stronger NI antibody responses and antiviral protection. Furthermore, the NE-adjuvanted trivalent NA vaccine (N1+N2+BV) elicited robust humoral and cellular immunity and provided effective protection in mice against two H1N1, two B/Victoria, and one H5N1 strain. Collectively, these findings support the efficacy of a multivalent NA-based vaccine strategy and highlight its strong potential as a candidate UIV.