Five marine fish pathogens, including Vibrio anguillarum, Aeromonas salmonicida, V.parahaemolyticus, V. harveyi, Shewanella putrefaciens were previously isolated and identified. In this study,virulence of these pathogens were determined by artificial infection experiments. The results showed that LD50 value of the five bacteria against turbot Scophthalmus maximus were 102~107CFU/fish. The polyclonal antibodies against them were produced from five New Zealand white rabbits respectively,and the titers of the polyclonal antibodies were 1∶2 048 000, 1∶16 000, 1∶16 000, 1∶1 024 000, 1∶128 000 respectively. The polyclonal antibodies were used in indirect ELISA as primary antibody, and the goat anti-rabbit IgG-HRP was used as a secondary antibody. The indirect ELISA method of rapid detection of five pathogens was developed. The sensitivity of the serum was tested, and the lowest concentrations of five pathogens that can be detected were 104, 108, 107, 105, and 106 cell/ml, respectively. Obvious cross reactions were present between three strains of vibrio and the antisera. By using the developed indirect ELISA method, 13 bacterial strains isolated from the infected fish were detected. One strain from the flounder and black rockfish Sebastodes fuscescens samples in the area of Jiaonan, Shandong Province, was positive for S. putrefaciens and two were positive for V. harveyi; one strain from Cynoglossus semilaevis in the area of Wudi, Shandong Province, was positive for both V. parahaemolyticus and V. harveyi. This result is consistent with the result of 16S rDNA method.