Recent data have shown that Dobrava (DOB) hantavirus is the cause of severe haemorrhagic fever with renal syndrome (HFRS) in central and eastern Europe. To determine whether serological assays need to be based on the homologous viral antigen rather than on closely related hantavirus antigens, acute and convalescent sera from patients with HFRS collected in former Yugoslavia were examined for IgM and IgG to three hantavirus antigens; DOB, Hantaan (HTN) and Puumala (PUU). Focus reduction neutralization test was included for comparison and confirmation of the enzyme-linked immunosorbent assay (ELISA) results. Although the results showed that the cross-reactivity was high between these three antigens during the acute phase of the disease, one of 155 patients serum samples reacted only in the DOB antigen-based IgM assay. The evaluation of IgG reactivities revealed that a DOB antigen-based IgG ELISA has to be used in sero-epidemiological studies; 7.1% (11/155) of the acute phase/early convalescent sera and 12.5% (2/16) of the late convalescent sera, respectively, reacted only with the homologous DOB antigen.