SREL Reprint #2696




A microscaled mercury saturation assay for metallothionein in fish

Patricia Shaw-Allen, Muriel Elliott, and Charles H. Jagoe

University of Georgia, Savannah River Ecology Laboratory, P.O. Drawer E, Aiken, South Carolina 29802, USA

Abstract: A mercury (Hg) saturation assay for measuring metallothionein (MT) in fish liver was modified by optimizing binding conditions to minimize the mercury and tissue consumed. The revised method uses stable Hg at low concentrations instead of 203Hg. At the reduced Hg concentrations used, MT concentrations in livers homogenized in saline appeared to increase systematically with dilution in both bluegill sunfish (Lepomis macrochirus) and largemouth bass (Micropterus salmoides). This error suggested a binding limitation due to sulfhydryl oxidation or competition for and removal of mercury by non-MT proteins. Homogenizing tissues in trichloroacetic acid (TCA) eliminated the interference. To further evaluate the method, the protocol was tested in the laboratory and field. Metallothionein in bluegill injected with 0.6 mg/kg zinc chloride increased at a rate of 0.03 nmole MT/g liver/h (r2 = 0.53, p = 0.001). Linearity improved when data were corrected for protein content (r2 = 0.74, p < 0.0001). Metallothionein levels in bluegill from a coal ash-contaminated environment were significantly increased over that of hatchery-reared sunfish (F = 20.17, p = 0.0003). The microscaled procedure minimizes concerns related to radioisotope use and waste generation while retaining the high sensitivity of the 203Hg assay.

Keywords: Metallothionein, Mercury saturation, Coal ash, Fish

SREL Reprint #2696

Shaw-Allen, P., M. Elliott, and C. H. Jagoe. 2003. A microscaled mercury saturation assay for metallothionein in fish. Environmental Toxicology & Chemistry 22(9): 2005-2012.

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