Compared accuracy of two enzyme linked immunosorbent assay (ELISA) methods performed with polyclonal antibodies to measure immunoglobulin levels inPlatichthys flesusL. 1758 (Flounder)

Abstract Polyclonal antibodies (PAbs) directed against flatfish ( Platichthys flesus L.) immunoglobulins (IgM) were prepared and characterised for use in immuno-assays. IgG antibodies produced in two rabbits reacted specifically with flatfish IgM. A double-antibody sandwich enzyme-linked immunosorbent assay (DS-ELISA) and a competition ELISA (C-ELISA) have been developed to measure flatfish immunoglobulin concentration in serum. The DS-ELISA employs biotinylated and non-biotinylated Ig, while C-ELISA uses only one biotinylated Ig. The C-ELISA method gave an average value of 2·630±0·067 mg ml −1 for flounder IgM and the DS-ELISA 4·010±0·072 mg ml −1 . Total serum protein titration was determined by the Bradford method and values of 20% and 30·4% IgM in the total serum protein were calculated using the C-ELISA and DS-ELISA, respectively. A comparative study of these two methods reveals that both can be used. However, the C-ELISA seems to be more accurate and easier to use. It is concluded that the ELISA technique using PAbs is a potential method to measure flounder IgM and could be helpful in the assessment of the immune status in flatfish.