[b]Introduction and objective[/b]. The aim of study was to test the sera of healthy dairy cows by ELISAs, the methods also used in other groups of animals in the Czech Republic, and thus to obtain actual data about [i]N. caninum[/i] and [i]T. gondii[/i] seroprevalence in cattle. [b]Material and methods[/b]. In the Czech Republic, sera from 546 clinically healthy dairy cows ([i]Bos primigenius f. taurus[/i]) aged > 2 years from 49 farms in 7 districts were collected. Sera were tested for [i]Neospora caninum[/i] antibodies by a commercial competitive-inhibition enzyme-linked immunosorbent assay; samples with more than 30% inhibition were considered as positive. The same samples were also analysed for Toxoplasma-specific IgG antibodies by an enzyme-linked immunosorbent assay; samples with more than 50% S/P were considered as positive. [b]Results[/b]. Antibodies against [i]N. caninum[/i] were found only in 3 cows (0.5%) with inhibitions of 47, 78 and 85. Antibodies against [i]T. gondii [/i]were found in 53 cows (9.7%) with S/P ranging from 51% to over 211%; positive animals were found in 4 of 7 districts, with prevalences ranging from 8% – 14%. Indication of mixed infections (concurrent presence of both [i]N. caninum[/i] and [i]T. gondii [/i]antibodies) was not proved. [b]Conclusion[/b]. The results of the study indicate that dairy cows in the Czech Republic have a relatively low seroprevalence for both [i]N. caninum[/i] and [i]T. gondii[/i]. Therefore, natural infection with [i]N. caninum[/i] and [i]T. gondii[/i] seems not to be very common in Czech cattle. These results show actual data about [i]N. caninum[/i] and [i]T. gondii [/i]infection in healthy dairy cattle from the Czech Republic.