Importance of storage conditions for the stability of zinc- and cadmium-induced metallothionein

We examined the storage stability of metallothionein (MT), a cysteine-rich protein that has diagnostic potential as a cancer marker and in the assessment of Zn status and heavy-metal toxicity. MT was rapidly degraded in samples of rat whole liver at −20°C or −70°C. MT in supernatants from heat-treated rat liver homogenates stored as 1 : 5 dilutions of liver from Zn- or Cd-induced rats were stable (recovery >98%) for 100 d at temperatures of −70°C and −196°C but not at −20°C, regardless of the presence of dithiothreitol (DTT) or argon. The variability of MT measurement by the 109Cd-hemoglobin affinity assay was however greatest in samples from Zn-induced rats stored without DTT. The integrity of the MT protein in supernatants of heat-treated homogenates stored for 100 d was demonstrated by Sephadex G-75 chromatography. When heat-treated supernatants were stored as dilute solutions (1 : 125 of liver), MT was unstable regardless of treatment or storage temperature. Our findings show that liver MT is stable for at least 4 mo as a supernatant of a heat-treated homogenate (1 : 5 dilution of liver) when stored at or below −70°C and in the presence of DTT.