The Elbe River was extremely loaded by heavy metals and arsenic during the second half of the 20th century as a consequence of intensive chemical industry activities. To assess long-term contamination trends in the Elbe after socio-economical changes in 1989, selected macroinvertebrate species were used to identify biotic accumulation levels in the period from 1993 to 2005. Concentrations of As, Cd, Pb, and Hg were determined in Asellus aquaticus (Crustacea), Bythinia tentaculata (Mollusca), Erpobdella spp. (Hirudinea), and larvae of the family Chironomidae (Diptera) over a 3-year period along the longitudinal profile of the Czech part of the Elbe River and the lower Vltava (the main Elbe tributary). In this study, new evaluative approaches were used to assess these multiparameter data sets. The results showed that the most contaminated Czech sites were located in the industry-heavy middle Elbe region between Valy and Obříství. Surprisingly high concentrations of As, Cd, and Pb were observed in the upper stretch near the headwaters. Probably as a consequence of former mining activities and the metallurgic and glass industries, the lower Vltava was loaded with Cd and Pb. Despite substantial improvements in the water quality of the Elbe River within the studied time period, we found no general decreasing trend of As, Cd, and Pb in macrozoobenthos. A significant decrease of Hg was revealed, however, occurring already between 1993 and 1996. The results of interspecies comparisons showed that the investigated elements were most accumulated by chironomid larvae and Erpobdella spp.