The aim of our investigation was to carry out quantitative isolation of Cu-thionein (Cu-Th) in human body fluids (urine, plasma and breast milk) in order to determine the level of exposure to heavy metals. In the experiment covalent affinity chromatography with thiol-disulphide interchange gel (CAC-TDI) was used as a solid phase extraction (SPE) support for preconcentration of Cu-thionein (Cu-Th) protein and Cu bonded with MT from water and human fluids samples. The present paper is a continuation of early experiments on the quantitation of Hg-thionein (Hg-Th), Cd-thionein (Cd-Th) and Zn-thionein (Zn-Th) in human body fluids such as urine, plasma and breast milk.