DAO 39:231-236 (2000) - doi:10.3354/dao039231
An antigen-capture enzyme-linked immunosorbent assay (ELISA) to detect isometamidium chloride in Oncorhynchus spp.
B. F. Ardelli, P. T. K. Woo*
ABSTRACT: An antigen-capture enzyme-linked immunosorbent assay (ELISA) was developed to detect and measure isometamidium chloride in the plasma of Oncorhynchus tshawytscha and O. mykiss. Isometamidium-ovalbumin conjugate and anti-isometamidium antibodies were used to coat polystyrene plates. The peroxidase saturation technique was used to optimize 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 anti-isometamidium antibodies to coat plates was at pH 7.3 and a 1:1000 dilution (0.0012 mg ml-1 protein). The ELISA was sensitive as it detected 0.0006 mg ml-1 of isometamidium in fish plasma. Isometamidium diluted with saline could not be detected at concentrations less than 0.05 mg ml-1 . The results indicate that this ELISA is much more sensitive when isometamidium is bound to plasma than unbound isometamidium in saline.
KEY WORDS: Antigen-capture ELISA · Isometamidium · Oncorhynchus
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