An antigen-capture enzyme-linked immunosorbent assay

An antigen-capture enzyme-linked immunosorbent assay (ELISA) was developed to detect and measure isometamidium chloride in the plasma of Oncorhynchus tshawytscha and 0. mykiss. Isometamidium-ovalbunlin conjugate and anti-isometamidium antibodies were used to coat polystyrene plates. The peroxidase saturation technique was used to optirnize the coating antigen concentration; it demonstrated low affinity of the isometamidium-ovalbumin conjugate but high affinity of the anti-isometamidium antibodies for polystyrene surface sites. The optimal conditions of antiisometamidium antibodies to coat plates was at pH 7.3 and a 1:1000 dilution (0.0012 mg ml-' protein). The ELISA was sensitive as it detected 0.0006 mg ml-' of isometamidium in fish plasma. Isometamidium diluted with sallne could not be detected at concentrations less than 0.05 mg ml-I . The results indicate that this ELISA is much more sensitive when isometamidium is bound to plasma than unbound isometamidium in saline.