The nucleoprotein (NP) antigen isolated from influenza A virus by solubilization with Triton X-100 (TX-100) and further electrophoresis with SDS-cellulose acetate membrane gave a single band on SDS-polyacrylamide gel electrophoresis. Rabbit anti-serum hyperimmunized with the NP reacted only against the NP antigen. Moreover, a well-defined single precipitin line was shown between the NP and human sera. These results suggested that the NP was possible to detect anti-NP antibody in human serum. Immuno double diffusion (IDD) and single radial immunodiffusion (SRD) tests using the NP were established to detect the anti-NP antibody in human sera. During an epidemic caused by antigenic drift strain, anti-NP antibody was detected by the IDD test in the cases which did not show any significant rise in HAI titer. During a mixed epidemic caused by the different strain of HA antigen, the infection ratio in mass population was revealed more convenient and sensitive by SRD than HAI. The anti-NP antibody was detected by IDD for long periods of one year or more after infection. These results suggest that the detection of anti-NP antibody is applicable to serologic studies, particularly serologic diagnosis and serologic surveys of influenza infection in mass population.