Immunochemical Quantification of Metallothioneins of a Marine Mollusc

Abstract

Mercury-induced, low molecular weight, metal-binding proteins were isolated from the marine mussel Mytilus edulis and used as antigen in the development of antibodies and an enzyme-linked immunosorbent assay (ELISA) for quantification of the proteins. Partial characterization of the isolated protein indicated that its properties were consistent with those of metallothionein (MT). In contrast with most MTs, this protein occurred as an apparent dimer and contained glycine at high levels. Polyclonal antibodies against this protein were produced in goats and purified to an IgG fraction by ammonium sulfate precipitation and DEAE ion-exchange chromatography. Ouchterlony analysis and ELISA showed that these antibodies were cross-reactive with two other charge variants of mercury-induced, metal-binding proteins of M. edulis, but not with rabbit MT. The ELISA was based on an indirect, competitive procedure utilizing a rabbit anti-goat IgG–horseradish peroxidase conjugate as the second antibody. Application of this assay to cytosolic extracts of mussel gills indicated 0.5 μg metal-binding proteins/g wet tissue weight in gills of control mussels and elevated levels up to 1780 μg/g following exposure to mercury, cadmium, or copper for 28 d. Zinc was not effective as an inducer of these proteins.