Abstract: A 1 320 bp fragment of NP gene of avian influenza virus (AIV) was cloned and integrated into the genome of T4-z1 phage with SOC-deleted and lysozymedefected by homologous recombination. The recombinant phage was named as T4-z1-NP. The results of SDS-PAGE and Western blot showed that nucleoprotein of AIV was successfully displayed on the surface of the T4-z1-NP. Based on the purified T4-z1-NP, an indirect enzyme-linked immunosorbent assay (T4-NP-ELISA) was developed for detection of antibodies against AIV. Its sensitivity relative to AGP was 100% but the specificity was 87.62%. The detection accuracy rate of T4-NP-ELISA was 91.43%. It could detect antibodies to H5, H7 and H9 subtype AIV, but has no significant reaction to positive serum of IBD, IB, MD, ND and EDS. The sensitive test confirmed that when the positive serum of AI was diluted to 1∶640, the OD450 value was still positive, which suggested it had better sensitivity. The accordance rate between T4-NP-ELISA and commercial IDDEXX ELISA was 968%, when 192 serum samples were detected with them. These results indicated that T4-NP-ELISA could not only detect antibody to AIV, but also provide an alternative technique for diagnosis and control of AI.