The EU Water Framework Directive (WFD) requires the biological elements of aquatic ecosystems, including fish abundance, to be monitored. At present hydroacoustic methods provide the most reliable and accurate estimates of many fish stocks but further development and standardization of these methods are necessary if they are to be useful as a monitoring tool. The aim of this paper is to compare acoustic estimates of fish abundance obtained at the operating frequencies 70 kHz (Simrad EK60) and 120 kHz (Simrad EY500), those most often used in inland water fishery acoustics. Measurements were performed in Lake Pluszne, where the epilimnion is occupied mainly by cyprinids and percids, and the hypolimnion is dominated by vendace (Coregonus albula). The frequency–response function was studied using Sonar 5 multi-frequency analysis facilities. It has been shown that, independent of the fish populations, these two frequencies give similar estimates of the fish target strength (TS) and volume backscattering strength (Sv).