Indirect enzyme-linked immunosorbent assay (ELISA) for the detection of antibody in serum of Japanese flounder [Paralichthys olivaceus]

An indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of antibody in serum of Japanese flounder, Paralichthys olivaceus against Edwardsiella tarda. Sera were collected from the flounder which had been injected with formalin-killed E. tarda cells (FKC). The sera were precipitated with a 40% saturated ammonium sulfate solution, and immunoglobulin (Ig) was purified from the precipitates by gel-filtration and ion-exchange chromatography. The purified Ig was administered to a rabbit to prepare anti-flounder Ig serum. Sensitivity of the ELISA was more than 6 times higher than that of formerly used agglutination methods. The flounder were vaccinated by three different methods; intraperitoneal injection of lipopolysaccharide of E. tarda (LPS), immersion in LPS, and immersion in FKC solution. The titers of vaccinated groups except for immersion in LPS were significantly higher than that of a control group (p < 0.05). However, in the challenge test, there were no significant differences in the survival rates between vaccinated and control groups.