The metallothioneins (MT), a family of proteins with relatively low molecular weight (6-7 kDa), are characterised by the intrinsic presence of 20 cysteinyl groups in their structure, which confers unique metal binding properties to the molecule. Since MT are involved in biological roles, quantification of MT remains an important task. To date, a large number of determination methods have been developed. In this paper recent developments, from 1995 to the present, in methodology employed in quantification studies of total MT and MT polymorphism are described. Different fields were taken into consideration, such as (i) separation techniques and hyphenated systems, (ii) electrochemical methods, (iii) immunological methods and (iv) quantification of MT mRNA. The data presented are based on our own and published results. A brief overview of the use of metallothionein as a biomarker is included as a relevant example of the importance of MT quantification. Finally, general problems associated with determination and evaluation of obtained results within the above four topics are mentioned.