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Quantification of metallothioneins by a silver-saturation method

Journal Article · · Toxicol. Appl. Pharmacol.; (United States)
The binding of Ag+ to metallothionein (MT) was investigated, and a Ag-saturation assay was developed for the measurement of MT in tissues. When samples of purified hepatic Zn-MT or Cd-MT were titrated with Ag+ followed by hemolysate-heat treatment, it was found that saturation of MT occurred at 17 to 18 g-atoms Ag+/mol protein. The rank order of potencies of metals to displace Ag+ from 110mAg-labeled Ag-MT was Ag+ greater than Cu2+ greater than Cd2+ greater than Hg2+ greater than Zn2+ at pH 8.5 in 0.5 M glycine buffer. When a Ag-saturation (Ag-hem) assay was compared with a Cd-saturation (Cd-hem) technique, excellent correspondence was obtained in the measurement of MT from various sources including purified Zn-MT (II), MT in human kidney and liver tissue samples, hepatic MT in Cd-injected adult rats, and renal MT in Hg-injected adult rats. However, in cases where significant amounts of Cu-MT were present, such as in the livers of Cu-injected rats or in the kidneys of Cd-injected rats, the Cd-hem assay consistently underestimated the MT concentrations compared to the Ag-hem method. This is attributed to the inability of Cd to displace Cu effectively from MT at pH 8.5. Thus the Cd-hem assay is not recommended for the measurement of metallothioneins containing a high Cu content. Under such conditions, the Ag-hem method seems superior.
Research Organization:
Univ. of Western Ontario, London
OSTI ID:
5615343
Journal Information:
Toxicol. Appl. Pharmacol.; (United States), Journal Name: Toxicol. Appl. Pharmacol.; (United States) Vol. 3; ISSN TXAPA
Country of Publication:
United States
Language:
English