The awareness of ciliate importance in freshwater has changed dramatically in the last decades, reflecting methodological attempts. We analysed two decades (1994-2018) of data on the surface (0-3 m) ciliate assemblage in the Slapy reservoir (Vltava River, Czech Republic) during two different nutrient-load defined periods. We identified, quantified, and evaluated the biomass of ciliates in the quantitative protargol-stained preparations. We used water age, nutrients, bacteria, heterotrophic nanoflagellates and rhodomonads as the main explanatory background variables. The sampling median and interquartile range data analysis and a non-metric multidimensional scaling were applied. Ciliates were grouped according to their feeding behaviour. The picoplankton filtering species dominated the assemblages in an annual mean (halteriids and minute strobilidiids followed by peritrichs). Algae hunters Balanion planctonicum, urotrichs and nanoplankton filtering tintinnids were important before the spring phytoplankton peak when a maximum of ciliate biomass reflected mixotrophic nanoplankton filtering pelagostrombidiids. Only there, ciliate biomass tightly followed their quantified prey. Heterotrophic and mixotrophic Askenasia and Lagynophrya were typical raptorial/flagellate hunters; only Mesodinium spp. peaked during autumn. The oligotrophication increased the ciliate assemblage biomass in the surface layer during the stratification in concordance with the PEG model.