Determination of Cd-thionein in biological materials: comparative standard recovery by five current methods using protein nitrogen for standard calibration.

Five methods of metallothionein (MT) estimation in biological materials (Hg-saturation assay, Cd-saturation assay, thiolate group determination, Sephadex G-75 gel chromatography with subsequent Cd determination by Zeeman-AAS, and a radioimmunoassay (RIA) were compared for their ability to recover standard MT(rabbit Cd-MTI) from liver or kidney S9. Uniform molar calibration of all assays was achieved using the known amino acid composition of standard MT and the nitrogen content of the standard MT stock solution as measured after complete Kjeldahl digestion. Known molar amounts of standard MT were added exogenously to rat liver- or kidney-S9 samples and recoveries measured using the methods indicated. In an overall rating, RIA and Cd-saturation assay performed best with recoveries of 97 +/- 12.0 and 105 +/- 9.7%, respectively. On the other hand, the remaining methods either underestimated (thiolate group determination, G-75/Cd-AAS) or overestimated (Hg-saturation assay) the theoretical expectation by up to 44%.